Faculty of Pharmacy - Mahidol University

Mahidol University Annual Research Abstracts, Vol. 38 289 

AFLATOXIN M 1 CONTAMINATIN IN RAW 

MILK WITHIN THE CENTRAL REGION OF 

THAILAND (NO. 0718) 

Nongluck Ruangwises 1 , Suthep Ruangwises 2 

1 Department of Pharmaceutical Chemistry, Faculty of 

Pharmacy, Mahidol University, Thaialnd. E-mail : 

pynrw@mahidol.ac.th; 2 Department of Veterinary Public 

Health, Faculty of Veterinary Science, Chulalongkorn 

Univesity, Thailand. 

Aflatoxin M 1 (AFM 1 ) was found in all of the 240 raw 

milk samples collected from milk tanks of 80 dairy farms at a 

collecting center in the central region of Thailand. Milk samples 

from individual farms were collected in three seasons. The average 

concentration of AFM 1 in milk samples collected in winter (0.089 

+ 0.034 μg/L) was significantly higher than those in rainy season 

(0.071 + 0.028 μg/L) and summer (0.050 + 0.021 μg/L). The 

present study suggests that regulatory limits for AFM 1 are needed 

to regulate and ensure the quality of raw milk and milk products 

in Thailand. 

Key words : aflatoxin M 1 , raw milk, HPLC 

(Bulletin of Environmental Contamination and Toxicology. 85(2), 

195-8, 2010.) 

This study was financially supported in part by a 2005-2006 Fiscal 

Research Grant. 

SCREENING TEST FOR ENROFLOXACIN 

RESIDUES IN CHICKEN MEAT BY PLANAR 

CHROMATOGRAPHY (NO. 0719) 

Jerawan Tansiri 1 , Nongluck Ruangwises 1 , Piyanuch 

Rojsanga 1 and Tasanee Chokcharoenrat 2 


Pharmacy, Mahidol University, Thailand; 2 Department of 

Medical Sciences, Ministry of Public Health Thailand. E-mail 

: pynrw@mahidol.ac.th 

Enrofloxacin is an antibiotic widely used in foodproducing 

animals such as pork, beef, chicken, fish, and shrimp, 

especially chicken and egg products. Antibiotic contamination 

causes decrease in meat quality and effects on human health. 

Screening test for enrofloxacin residues in chicken meat by high 

performance thin layer chromatography have been developed. 

Appropriate solvents are used for extracting the residues from 

samples before analyzing by high performance thin layer 

chromatography which can distinguish the analyte from other 

substances. The detection limit is 100 μg/kg in chicken meat. 

The sensitivity and specificity are 90, 100% respectively. The 

other confirmatory methods such as liquid chromatography mass 

spectrometry was used to analyze the same samples by comparison 

with high performance thin layer chromatography screening test. 

The coincidental results were obtained in all samples. Chicken 

meat samples from various sources around Bangkok were sampled 

and tested by high performance thin layer chromatography 

screening test. From 116 samples of chicken meat, only one 

sample was out of the limit of enrofloxacin residue (100 μg/kg). 

Key words : Enrofloxacin, chicken meat, planar chromatography 

(Poster presentation In Pure and Applied Chemistry Internation 

Conference 2010: Challenges in Chemistry for Sustainable 

Development Jan 21-23, 2010, Sunee Grand Hotel and 

Convention Center, Ubon Ratchathani, Thailand.) 

This study was financially supported by National Research 

Council and Mahidol University, Bangkok, Thailand. 

A SENSITIVE NON-AQUEOUS CAPILLARY 

ELECTROPHORESIS-MASS SPECTROMETRIC 

METHOD FOR MULTIRESIDUE ANALYSES 

OF B-AGONISTS IN PORK (NO. 0720) 

Oraphan Anurukvorakuna, Wolfgang Buchbergerb, Markus 

Himmelsbachb, Christian W. Klampelb and Leena 

Suntornsuk 

Department of Pharmaceutical Chemistry, Facult of 

Pharamcy, Mahidol University, Bangkok 10400, Thailand. Email 

: pyll@mahidol.ac.th 

Non-aqueous capillary electrophoresis-mass 

spectrometry (NACE-MS) was developed for trace analyses of 

β-agonists (i.e. clenbuterol, salbutamol and terbutaline) in pork. 

The NACE was in 18 mM ammonium acetate in methanolacetonitrile-glacial 

acetic acid (66: 33:1, v/v/v) using a voltage 

of 28 kV. The hyphenation of CE with a time-of-fight MS was 

performed by electrospray ionization interface employing 5 mM 

ammonium acetate in methanol-water (80 : 20, v/v) as the sheath 

liquid at a flow rate of 2 μL/min. Method sensitivity was enhanced 

by a co-injection technique (combination of hydrodynamic and 

electrokinetic injection) using a pressure of 50 mbar and a voltage 

of 10 kV for 12 s. The method was validated in comparison with 

HPLC-MS-MS. The NACE-MS procedure provided excellent 

detection limits of 0.3 ppb for all analytes. Method linearity was 

good (r 2 > 0.999, in a range of 0.8 – 1000 ppb for all analytes). 

Precision showed > RSD s of < 17.7%. Sample pre-treatment was 

carried out by solid-phase extraction using mixed mode reversed 

phase/cation exchange cartridges yielding recoveries between 69 

and 80%. The NACE-MS could be successfully used for the 

analysis of β-agonists in pork samples and result showed no 

statistical differences from the values reported by the Ministry 

of Public Health, Thailand using HPLCMS-MS method. 

Key words : clenbuterol, salbutamol, terbutaline 

(Biomedical Chromatography. 2010, 24, 588-599) 

(The Thailand Research Fund, The Royal Golden Jubilee Ph.D. 

Program) 

RAPID ANALYSIS OF ALKYLPHOSPHONATE 

DRUGS BY CAPILLARY ZONE ELECTRO- 

PHORESIS USING INDIRECT ULTRAVIOLET 

DETECTION (NO. 0721) 

Brompoj Purthiwanasan, Leena Suntornsuk

290 

Department of Pharmaceutical Chemistry, Faculty of 

Pharmacy, Mahidol University, Bangkok 10400, Thailand. Email 

: pylll@mahidol.ac.th 

A rapid capillary electrophoretic method for the 

analysis of three alkylphosphonate drugs (i.e. fosfomycin 

disodium (FOS), clodronate disodium (CLO) and alendronate 

sodium (ALN)) was developed by using multiple probe BGE and 

indirect UV detection. BGE containing 30 mM benzoic acid, 

5mM salicylic acid and 0.5 mM CTAB (pH 3.8), temperature of 

301C, applied voltage of 30 kV and detection at 220 nm provided, 

baseline separation of all analytes (resolution ® 4.2) in 3.2 min. 

EOF reversal by addition of CTAB and negative voltage polarity 

leading to the co-EOF flow and short analysis time. Two probe 

BGE greatly improved peak symmetry. The method showed good 

linearity (r 2 40.999 in ranges of 20-1000 mg/mL for FOS, 100- 

1000 mg/mL for CLO and 100-750 mg/mL for ALN) repeatability 

(RSDo2.15%), recovery (99.3-101.1%)and sensitivity (LODo50 

mg/mL). Freshly prepared BGE and sample solutions are essential 

for the method precision and accuracy. This new method can be 

utilized for routine analysis of FOS, CLO and ALN in dosage 

forms because of its efficiency, reliability, speed and simplicity. 

Key words : Alendronae, Clodronate, Fosfomycin, Indirect UV 

detection. 

(Jounral of Seperation Science. 2010, 33, 228-234.) 

(The National Research Council of Thailand (grant no. PK 2551- 

24) 

RECENT ADVANCES OF CAPILLARY 

ELECTROPHORESIS IN PHARMACEUTICAL 

ANALYSIS (NO. 0722) 

Leena Suntornsuk 




This review covers recent advances of capillary 

electrophoresis (CE) in pharmaceutical analysis. The principle, 

instrumentation, and conventional modes of CE are briefly 

discussed. Advances in the different CE techniques (non-aqueous 

CE, microemulsion electrokinetic chromatography, capillary 

isotachophoresis, capillary electrochromatography, and 

immunaffinity CE), detection techniques (mass spectrometry, 

light-emitting diode, fluorescence, chemiluminescence, and 

contactless conductivity), on-line sample pretreatment (flow 

injection) and chiral separation are described. Applications of 

CE to assay of active pharmaceutical ingredients (APIs), drug 

impurity testing, chiral duurg separation, and determination of 

APIs in biological fluids published from 2008 to 2009 are 

tabulated. 

Key words : Capillary electrophoresis, Active pharmaceutical 

ingredient, Impurity, Chiral separation 

(Analytical bioanalytical Chemistry. 2010, 398, 29-52) 

Faculty of Pharmacy 

SIMULTANEOUD DETERMINATION OF R-(-), 

S-(+)-BACLOFIN AND IMPURITY A BY 

ELECTROKINETIC CHROMATOGRAPHY 

(NO. 0723) 

Leena Suntornsuk, Suphutcharasa Ployngam 




A rapid method for he simultaneous analysis of R-(-)- 

, S- (+)-baclofen and impurity A, (4RS)-4-(4-chorophenyl) 

pyrrolidin-2-one, by electrokinetic chromatography was 

established. The optimizedcondition was in 100 mM sodium 

borate buffer (pH 9.9) containing 18mM β-cyclodextrin (CD) 

and 1% (v/v) CAN using a fused silica capillary dynamically 

coated with polyethylene oxide (PEO), with an effective length 

of 56 cm and an inner diameter of 50 mm, hydrodynamic injection 

at 50 mbar for 6 s, temperature of 45 o C, applied voltage of 27 

kV an UV detection at 220 nm. Baseline separation of all analytes 

was achieved within 9 min (R s > 2.7) with the migration order of 

impurity A, S-(+)- and R-(-)-baclofen. The method showed good 

linearity (r 2 > 0.999 in a range of 5-50 μg/mL for impurity A and 

50-500 μg/mL for baclofen enantimores), precision (%RSD s < 

3.37%) and recoveries (100.3% for R-(-)-baclofen, 101.6% for 

S-(+)-baclofen and 96.1% for impurity A). Detection and 

quantitation limits were 10 and 30 μg/ml for both enantiomers, 

and 2 and 5 μg/mL for the impurity, respectively. The method 

was efficient for the determination of baclofen enantiomers and 

impurity A in pharmaceutical raw material and formulations due 

to its reliability, speed and simplicity. 

Key words : Baclofen enantiomers, Impurity A, Cyclodextrin 

(Journal of Pharmaceutical and Biomedical Analysis. 2010, 51, 

541-548) 

(Thailand Research Fund (MAG-OSMEP505S178) 

CHIF-BASED APPLICATION ORIENTED 

CAPILLARY ELECTROPHORESIS ANALY- 

SERAS A GENERIC CE PLATFORM : ANALYSIS 

OF FLUORESCENT DYES BY NACE AND THE 

POTENTIAL FOR DERIVATISED BIOANALYTES 

WITH RED LIF DETECTION (NO. 0724) 

Nantana Nuchtavorn 1,2 , Petr Smejkal 1 , Michael Bredmoe 2 , 

Marketa Ryvoloval 1 , Fritz Bek 3 , Frantisek Foret 1 , Rosanne 

Guijt 1 , Leena Suntornsuk 4 , Mirek Mackal 1,2 

1 National Centre for Sensor Research, Irish Separation 

Science, Cluster and School Chemical Sciences, Dublin City 

University, Dublin 9, Ireland; 2 Australian Centre for Research 

on Separation Science, School of Chemistry, University of 

Tasmania, GPO Box 252-75, Hobart, Tasmania 7001, 

Australia; 3 Agilent Technologies, P.O.Box 1280, 76337, 

Waldbronn, Germany; 4 Department of Pharmaceutical 

Chemistry, Faculty of Pharmacy. Mahidol University, 

Bangkok 10400, Thailand. E-mail : pyll@mahidol.ac.th


CE separations of fluorescent dyes (i.e. methylene blue 

(MB), nile blue A (NB), brilliant cresyl blue (BC), toluidine blue 

(TB) and brilliant green (BG)) were performed on a commercial 

chip based-CE, Agilent 2100 Bioanalyzer using DNA microchips 

which is a glass chip with a 14 mm separation channel. These 

dyes are all blue in colour except BG and can be detected by red 

laser with the excitation wavelength at 635 nm. Non-aqueous 

CE (NACE) is a versatile method, provides different separation 

selectivity and solubility for a wide range of analytes from 

different organic solvents. Various solvents (i.e. ACN, DMSO, 

DMF, Acetone, EtOH and MeOH) were evaluated for the 

separation of these dyes2, however in this work. DMSO was 

selected as a suitable solvent to prepare ammonium acetate 

electrolyte, due to the non-volatility and compatibility of 

microchips. The linearities (r 2 ) were in the range of 0.9739-0.9988 

over 5-28 μM, 1.56-50 nM, 5-75 nM, 10-100 μM and 25-200 

μM for MB, NB, BC, TB and BG, respectively. The limits of 

detectin were 90 nM, 1.4 nM, 2 nM, 1 μM and 2.9 μM for MB, 

NB, BC, TB and BG, respectively. Due to the most sensitivity of 

NB among these fluorescent dyes, it should be potential 

fluorescent derivatisation reagent that would be suitable for the 

derivatisation of bioanalytes. Preliminary results will be shown 

examples of utilizations of NB in particular as a fluorescence tag 

for carboxylic acids and of fluorescent dyes in general for staining 

cells. 

(34 th International Symposium on Capillary 

Chromatography(ISCC), Riva Del Garda, Italy, 1-4 June 2010, 

poste presentation R06,p.40) 

ANALYSIS OF FLUORESCENT DYES BY 

NON-AQUEOUS CE ON A CHIP AND THE 

POTENTIAL FOR STAINED MICROOR- 

GANISMS WITH RED LIF DETECGTION 

(NO. 0725) 

Nantana Nuchtavor 1,2 , Petr Smejkal 1 , Michael Bredmore 2 , 

Marketa Ryvoloval 1 , Fritz Bek 3 , Frantisek Forest 1 , Rosanne 

Guijt 1 , Leena Suntornsuk 4 , Mirek Mackal 1,2 

1 National Centre for Sensor Research, Irish Separation 

Science Cluster and School of Chemical Sciences, Dublin City 

University, Dublin 9, Ireland; 2 Australian Centre for Research 

on Separation Science, School of Chemistry, Unversity of 

Tasmania, GPO Box 252-75, Hobart, Tasmania 7001, 

Australia; 3 Agilent Technologies, P.O. Box 1280, 76337 

Waldbronn, Germany; 4 Department of Pharmaceutical 

Chemistry, Faculty of Pharmacy, Mahidol University, 

Bangkok 10400, Thailand. 

CE separations of fluorescent dyes (i.e. methylene blue 

(MB), toluidine blue (TB), nile blue A (NB) and brilliant cresyl 

blue (BC)) were performed on commercial chip based-CE, Agilent 

2100 Bioanalyzer using DNA microchips which is a glass chip 

with a 14 mm separation channel. These dyes are all blue in colour 

and can be detected by red laser with the excitation wavelength 

at 635 nm. Non-aqueous CE (NACE) is a versatile method, 

provides different separation selectivity and solubility for a wide 

range of analytes from different organic solvents. Various solvent 

(i.e. DMSO, MeOH and ACN) were evaluated for the separation 

of these dyes1, however DMSO was selected as a suitable solvent 

to prepare ammonium acetate electrolytes, due to the nonvolatility 

and compatibility of microchips. The linearities (r 2 ) were 

in the range of 0.9739-0.9988 over 5-28 μM, 10-100 μM, 1.56- 

50 nM and 5-75 nM, for MB, TB, NB and BC, respectively. The 

limits of detection were 90 nM, 1 mM, 1 μM 1.4 nM, 2 nM and 

for MB, TB, NB and BC, respectively. Due to the most sensitivity 

of NB among these fluorescent dyes, it was utilized in particular 

as a fluorescent dye for staining microorganisms. A spacer method 

2, 3, was used for sample injection using a dilute cetyltrimethyl 

ammonium bromide to sweep microorganisms out of the sample 

zone and a small plug of blocking agent to negate the cells’ 

mobility and focus them into a narrow zone. This method shows 

the potential for the test of bacterial/fungal contamination using 

a chip-based CE. 

Key words : Microchip-based capillary electrophoresis, Laser 

induced fluorescence detection, Dyes, Microorganisms 

(2 nd REMSEA 2010: Regional Electrochemistry Meeting of South- 

East Asia on Applied Electrochemistry for Modern Life, Maha 

Chulalongkorn Building, Chulalongkorn University, Bangkok, 

Thailand, 16-19 November 2010, poster presentation PA28, p. 

133) 

BIOLOGICAL ACTIVITIES OF EXTRACTS 

FROM ANDAMAN SEA SPONGES, THAILAND 

(NO. 0726) 

Patchara Pedpradab 1 , Wanlapa Molex 2 , Veena Nukoolkarn 3 , 

Udomsak Darumas 4 

1 Department of Marine Science, Rajamangala University of 

Technology Srivijaya, 92150 Trang, Thailand, E-mail: 

agelas2004@gmail.com; 2 Department of Marine Science, 

Rajamangala University of Technology Srivijaya, 92150 

Trang, Thailand; 3 Department of Pharmacognosy, Faculty 

of Pharmacy, Mahidol University, 10400, Bangkok, Thailand, 

E-mail: pyvnk@mahidol.ac.th; 4 Department of Biology, 

Faculty of Sciences, Walailuk University, Tasala, 80161 

Nakhonsrithammarat, Thailand 

Thirty six organic extracts were prepared from 

eighteen marine sponges collected from the Andaman Sea, 

Thailand. The extracts were examined for anti-malaria, anti- 

Microbacterium tuberculosis, anti-herpes simplex virus, 

antimicrobial, anti-acetylcholinesterase enzyme and cytotoxic 

activities. Four extracts showed anti-M. tuberculosis, one antimalarial, 

twenty four antimicrobial and one extract exhibited 

cytotoxic activity. However, anti-acetylcholinesterase enzyme and 

anti-herpes simplex virus type 1 (HSV-1) activities were not 

recorded. Dichloromethane extracts prepared from Axinyssa sp., 

Halichondria sp. and Chondrosia reticulata exhibited potential 

anti-M. tuberculosis at MIC 50, 100 and 200 mg/mL, respectively. 

The Hexane part of Phakellia ventilabrum extract showed antimalarial 

activity (MIC= 2.8 mg/mL) while the dichloromethane 

extract showed anti-M. tuberculosis and cytotoxic activity with 

MIC 200 and IC 50 7.1 mg/mL, respectively. Antimicrobial activity 

was found in both the hexane and dichloromethane parts of 

extracts. 

Key words: Andaman Sea, Biological activity, Marine sponges

292 

SCREENING OF BIOACTIVE COMPOUNDS 

FROM MARINE MICROORGANISMS 

TARGETING BACTERIAL CELL DIVISION 

PROTEIN FtsZ (NO. 0727) 

Phennapa Charoenwiwattanakij 1 , Jaturong Pratuangdejkul 2 , 

Krit Thirapanmethee 3 , Montree Jaturanpinyo 4 , Chiti 

Thawai 5 , Khanit Suwanborirux 6 , and Veena Nukoolkarn 7 

1 Department of Pharmacognosy, Faculty of Pharmacy, 

Mahidol University, Sri-ayudhya road, Bangkok 10400, 

Thailand, E-mail: yoyo_ws@yahoo.com ; 2 Department of 

Microbiology, Faculty of Pharmacy, Mahidol University, Sriayudhya 

road, Bangkok 10400, Thailand, E-mail: 

pyjpd@mahidol.ac.th 3 Department of Microbiology, Faculty 

of Pharmacy, Mahidol University, Sri-ayudhya road, Bangkok 

10400, Thailand, E-mail: pyktr@mahidol.ac.th 4 Department 

of Manufacturing Pharmacy, Faculty of Pharmacy, Mahidol 

University, Sri-ayudhya road, Bangkok 10400, Thailand, Email: 

pymjt@mahidol.ac.th 5 Department of Applied Biology, 

Faculty of Science, King Mongkut’s Institute of Technology 

Ladkrabang, Bangkok, Thailand, E-mail: 

cthawai@yahoo.com 6 Faculty of Pharmaceutical Sciences, 

Chulalongkorn University, Bangkok 10330, Thailand, E-mail: 

skhanit@chula.ac.th 7 Department of Pharmacognosy, Faculty 


10400, Thailand, E-mail: pyvnk@mahidol.ac.th 

The essential bacterial cell division protein FtsZ has 

been recognized as an attractive target for new antibiotic agents 

but the target remains underexploited. In our study, various marine 

Actinobacteria were collected, cultured and extracted as well as 

tested for identifying FtsZ inhibitory activity. The antibacterial 

activity was characterized by using disc diffusion method. The 

results showed that Actinomycetes isolated AN6-2 and S08-2 

exhibited good activity against S. aureus, B. subtilis, P. 

aeruginosa, and E. coli. The inhibition zones of AN6-2 were 

2.4, 2.0., 1.1 and 1.57 cm while that of S08-02 were 2.61, 3.2, 

1.31 and 1.51 cm, respectively. To determine on-target activity, 

the morphometric analysis of ftsz-overexpressed E. coli was 

observed under electron microscopy. Both Actinomycetes isolated 

AN6-2 and S08-2 extract showed remarkable results with 

expressing filamentation of the bacilli due to continued shortterm 

growth in the absence of cell division. Effect on Z-ring 

characteristic of E. coli cells bearing an FtsZ-GFP fusion was 

studied by confocal microscopy. In the absence of the AN6-2 

extract, the typical Z ring pattern was observed at the mid-cell. 

Addition of AN6-2 extract resulted in cell elongation with 

dissipated Z-ring morphology. These results indicated that marine 

Actinobacteria is the promising sources for FtsZ inhibitor. The 

isolation and structure elucidation of the active compound are 

under investigation. 

Key words: FtsZ, FTsZ inhibitors, Marine microorganisms 

BACTERIAL CELL DIVISION PROTEIN (FtsZ) 

INHIBITORS FROM THAI MEDICINAL PLANTS 

(NO. 0728) 

Krit Thirapanmethee 1 , Veena Nukoolkarn 2 , Montree 

Jaturanpinyo 3 , and Jaturong Pratuangdejkul 4 

1 Department of Microbiology, Faculty of Pharmacy, Mahidol 


pyktr@mahidol.ac.th; 2 Department of Pharmacognosy, 

Faculty of Pharmacy, Mahidol University, Sri-ayudhya road, 

Bangkok 10400, Thailand, E-mail: pyvnk@mahidol.ac.th; 

3 Department of Manufacturing Pharmacy, Faculty of 

Pharmacy, Mahidol University, Sri-ayudhya road, Bangkok 

10400, Thailand, E-mail: pymjt@mahidol.ac.th; 4 Department 

of Microbiology, Faculty of Pharmacy, Mahidol University, 

Sri-ayudhya road, Bangkok 10400, Thailand, E-mail: 

pyjpd@mahidol.ac.th 

As the less effective antibiotics and the emergence of 

multi-drug-resistant bacteria continue to develop, these become 

more significant problems, both in hospitals and in the 

community. To overcome the rising antibiotic resistance of 

important human pathogens, there is an urgent need to discover 

novel antibacterial targets and novel drug with new mechanisms 

of action. We currently interest to study protein that plays a crucial 

role in bacterial cell division. To this end, we focus our attention 

on the bacterial protein FtsZ due to its major role in formation of 

Z-ring that is essential for bacterial cell division and viability. 

FtsZ (filament forming temperature – sensitive mutant Z), a 40.3 

kD tubulin homolog, is an essential cell division protein. It 

polymerized in a GTP – dependent manner to form a Z – ring at 

the mid – cell, which act as a framework for recruiting other cell 

division proteins. There are many studies reported that 

perturbation of FtsZ assembly result in inhibition of bacterial 

proliferation. Therefore, FtsZ is a very promising target for the 

new class of antimicrobial agent. Berberine, a isoquinolone 

alkaloid obtained from various species of plants such as Berberis 

dawinii, Coscinium fenestratum, and Coscinium wallichianum. 

Plants containing berberine have been used as antimicrobial, 

stomachic and bitter tonic in traditional Indian and Chinese 

medicine. It exhibited antimicrobial activity against the variety 

of bacteria, fungi, protozoa, helminthes, and virus. Although, there 

are several studies on antimicrobial activity of berberine, but the 

mechanism of action have not been reported yet. The objective 

of this study is to elucidate the antimicrobial mechanism of 

berberine as an FtsZ inhibitor. Molecular modeling studies were 

carried out in order to gain structural information of interaction 

between E. coli-FtsZ and berberine. 

Key words: FtsZ, Berberine, GTPase inhibitor 


ASSESSMENT OF LIPOSOMES FOR DELIVERY 

OF CINNAMALDEHYDE, A FtsZ INHIBITOR 

(NO. 0729) 

Nevadee Somkitsiri 1 , Jaturong Pratuangdejkul 2 , Krit 

Thirapanmethee 3 , Veena Nukoolkarn 4 , Satit 

Puttipipatkhachorn 5 , and Montree Jaturanpinyo 6 



10400, Thailand, E-mail: pymjt@mahidol.ac.th; 2 Department 



pyjpd@mahidol.ac.th; 3 Department of Microbiology, Faculty 


10400, Thailand, E-mail: pyktr@mahidol.ac.th; 4 Department


of Pharmacognosy, Faculty of Pharmacy, Mahidol University, 


pyvnk@mahidol.ac.th; 5 Department of Manufacturing 

Pharmacy, Faculty of Pharmacy, Mahidol University, Sriayudhya 


pyspt@mahidol.ac.th; 6 Department of Manufacturing 

Pharmacy, Faculty of Pharmacy, Mahidol University, Sriayudhya 


pymjt@mahidol.ac.th 

Cinnamaldehyde, an aromatic compound extracted 

from the bark of cinnamon tree, has been recently recognized as 

a potent antibiotic with novel mechanisms for inhibiting essential 

cell division protein (FtsZ protein) in bacteria. Since this protein 

is found only in bacteria, its inhibitor has been attractive as a 

new challenge for many researchers to develop a potential antimicrobial 

agent. However, use of cinnamaldehyde as a therapeutic 

agent is limited by its poor aqueous solubility. This issue can be 

practically overcome by encapsulation within liposomes. This 

study was thus aim to investigate the physicochemical properties 

of liposomal formulations containing cinnamaldehyde. 

Key words: Cinnamaldehyde, Liposome, Drug delivery 

SCREENING OF BIOACTIVE COMPOUNDS 

FROM NATURAL PRODUCTS TARGETING 

BACTERIAL CELL DIVISION PROTEIN FtsZ 

(NO. 0730) 

Phennapa Charoenwiwattanakij 1 , Jaturong Pratuangdejkul 2 , 

Krit Thirapanmethee 3 , Montree Jaturanpinyo 4 , and Veena 

Nukoolkarn 5 



Thailand, E-mail: yoyo_ws@yahoo.com 2 Department of 



pyjpd@mahidol.ac.th 3 Department of Microbiology, Faculty 


10400, Thailand, E-mail: pyktr@mahidol.ac.th 4 Department 



pymjt@mahidol.ac.th 5 Department of Pharmacognosy, 


Bangkok 10400, Thailand, E-mail: pyvnk@mahidol.ac.th 

The increasing public health problem due to the 

emergence and spread of drug-resistant bacteria as well as the 

rising numbers of infectious patients leads to the need to search 

for new antibacterial agents. After the discovery of penicillin, 

the search for antibacterial agents focused on microorganisms. 

One source of them is from marine microorganisms which 

produces various kinds of bioactive substances. FtsZ, prokaryotic 

cytoskeleton protein, is one of the novel antibacterial targets. In 

cytokinesis, FtsZ is the first protein at mid cell and working as 

GTPase to undergo cytokinesis process by forming FtsZ ring. It 

is a tubulin homolog which is conserved in prokaryotic cells. 

This suggests that FtsZ can be a specific target for antibacterial 

agents. To discover new FtsZ inhibitors, various substances from 

nature are investigated, such as, berberine, cinnamaldehyde, 

curcumin, etc. These compounds gave positive results as FtsZ 

inhibitors. The other source of active compounds, 

microorganisms, is in the interest and will be investigated. We 

expect that our discovery targeting on FtsZ protein will lead to 

the new generation of compounds with significant therapeutic 

value for antibiotic drugs in the future. 

Key words: FtsZ, FTsZ inhibitors, Marine microorganisms 

THAI MARINE ORGANISMS : POTENTIAL 

SOURCES FOR ACETYLCHOLINESTERASE 

INHIBITORS (NO. 0731) 

Kornkanok Inkaninan 1 , Roosanee Langjae 2 , Anuchit 

Plubrukarn 3 , Yutthapong Sangnoi 4 , Akkharawit Kanjanaopas 

5 , Veena S Nukoolkarn 6 , and Khanit Suwanborirux 7 

1 Faculty of Pharmaceutical Sciences, Naresuan University, 

Phitsanulok 60500, Thailand, E-mail: k_ingkaninan@ 

yahoo.com; 2 Faculty of Pharmaceutical Sciences, Prince of 

Songkla University, Songkla 90112, Thailand; 3 Faculty of 

Pharmaceutical Sciences, Prince of Songkla University, 

Songkla 90112, Thailand, E-mail: anuchit.pl@psu.ac.th, 

chatchai.w@psu.ac.th, akkharawit.k@psu.ac.th,; 4 Faculty of 

Agro-Industry, Prince of Songkla University, Songkla 90112, 

Thailand; 5 Faculty of Pharmaceutical Sciences, Prince of 

Songkla University, Songkla 90112, Thailand, E-mail: 

akkharawit.k@psu.ac.th; 6 Faculty of Pharmacy, Mahidol 

University, Bangkok 10400, Thailand, E-mail: 

pyvnk@mahidol.ac.th; 7 Faculty of Pharmaceutical Sciences, 

Chulalongkorn University, Bangkok 10330, Thailand, E-mail: 

skhanit@chula.ac.th 

Acetylcholinesterase (AChE) inhibitors have gained 

interest due to their application for the treatment of Alzheimer’s 

disease. In order to search for new AChE inhibitors, marine 

organisms and micro-organisms in Thailand were collected and 

tested for AChE inhibitory activity using Ellman’s colorimetric 

methid in 96-welled microplates. The studies revealed some 

interesting AChE inhibitors. Petrosamine, a pyridoacridine 

alkaloid from a Thai marine sponge, Petrosia sp. showed potent 

inhibitory activity on AChE. Moreover, a new stigmastane-type 

steroidal alkaloid, 4-acetoxy-plankinamine B isolated from a 

sponge, Corticium sp. and marinoquinoline A 3 , a pyrole derivative 

isolated from a novel marine gliding bacterium, Rapidithrix 

thailandica showed a high activity expressed by the concentration 

that inhibited 50% of AChE activity (IC 50 ) in a micromolar range. 

Key words: Aetylcholinesterase inhibitor, Marine organisms, 

Petrosamine 

IN SILICO SCREENING AND STRUCTURAL 

BASED APPROACHES FOR NEW TARGET OF 

ANTIBACTERIAL AGENTS (NO. 0732) 

Jaturong Pratuangdejkul 1 , Kosit Sheranaravenich 2 , 

Phennapa Charoenwiwattanakij 3 , Veena Nukoolkarn 4 , Krit 

Thirapanmethee 5 , and Montree Jaturanpinyo 6 



pyjpd@mahidol.ac.th; 2 Department of Microbiology,

294 


Bangkok 10400, Thailand, E-mail: kkosit@hotmail.com; 



Thailand, E-mail: yoyo_ws@yahoo.com; 4 Department of 

Pharmacognosy, Faculty of Pharmacy, Mahidol University, 


pyvnk@mahidol.ac.th; 5 Department of Microbiology, Faculty 


10400, Thailand, E-mail: pyktr@mahidol.ac.th; 6 Department 



pymjt@mahidol.ac.th 

As the less effective antibiotics and the emergence of 

multi-drug-resistant bacteria continue to develop, these become 

more significant problems, both in hospitals and in the 

community. To overcome the rising antibiotic resistance of 

important human pathogens, there is an urgent need to discover 

novel antibacterial targets and novel leads with new mechanisms 

of action. Beside the conventional strategies i.e. whole-cell 

screening of natural products and molecules that exhibit 

antibacterial activities, recent advances in target identification 

and assay development have resulted in a flood of target-driven 

new antibiotic discovery. Many new structures dwell in bacteria 

have been studied by structure–genomics initiatives, delivering 

a wealth of new targets to be considered. To reinforce our 

discovery of novel antibiotics, in silico screening upon structural 

based approaches have yielded, placing particular emphasis on 

screens capable of identifying new leads involved in such 

processes as virulence factors, novel targets of essential metabolic 

pathways, underexploited targets that reside in bacterial cell, etc. 

Using the innovative tools of computational structure-based 

screening and design, antibacterial discovery must exploit these 

targets to harvest the next generation of antibacterial treatments 

and to accelerate the process of drug discovery. Owing to the 

need of new antibacterial therapy to overcome the problem of 

antibiotic resistance and the emergence or re-emergence of 

infections diseases, we currently interest to study protein that 

play a crucial role in bacterial cell division. To this end, we focus 

our attention on the bacterial protein FtsZ due to its major role in 

formation of Z-ring that is essential for bacterial cell division 

and viability. Although FtsZ is a homologue of eukaryotic 

cytoskeleton protein tubulin, but it is conserved in bacteria and 

does not appear in human cell. Thus, FtsZ provides as a new 

target for screening of specific FtsZ-inhibitors that can be 

developed as an antibacterial drug without any perturbation on 

human host cell. Up to now, three dimensional (3D) structures of 

FtsZ in different species (Aquifex aeolicus, Bacillus subtilis, 

Methanococcus jannaschii, Pseudomonas aeruginosa and 

Mycobacterium tuberculosis) and nucleotide states (GTP, GDP 

and empty) have been determined using X-ray crystallography. 

Structural insights into the conformational variability of FtsZ have 

been previously discussed and proposed the idea for straight and 

curved conformations of FtsZ protofilaments in order to describe 

mechanical role in Z-ring of cell division 1 . Moreover, the available 

FtsZ structures have also been used for understanding the 

mechanism of FtsZ-inhibitors at atomic level by applying 

molecular modeling techniques. Accordingly, the aim of our 

research is to perform molecular modeling study of FtsZ and its 

inhibitors screened from natural substances which show inhibition 

of bacterial proliferation. In the present work, Escherichea coli 

were used as bacterial model for study FtsZ-inhibitory activity 


of berberine, the natural compound isolated from Coscinium 

wallichianum. Molecular modeling studies were carried out in 

order to gain structural information of interaction between E. 

coli-FtsZ and berberine. 

Key words: Homology modeling, FtsZ, berberine 

IN SILICO STUDY OF INHIBITOR IN MYCO- 

BACTERIUM TUBERCULOSIS FTSZ : A 

STRUCTURAL INSIGHT INTO THE BINDING 

INTERACTION USING MOLECULAR DOCKING 

AND MOLECULAR DYNAMICS (NO. 0733) 

Kosit Sheranaravenich 1 , Veena Nukoolkarn 2 , Montree 

Jaturunpinyo 3 , Krit thirapanmethee 4 , Jaturong 

Pratuangdejkul 5 



kkosit@hotmail.com; 2 Department of Pharmacognosy, 


Bangkok 10400, Thailand, E-mail: pyvnk@mahidol.ac.th; 



10400, Thailand, E-mail: pymjt@mahidol.ac.th 4 Department 



pyktr@mahidol.ac.th; 5 Department of Microbiology, Faculty 


10400, Thailand, E-mail: pyjpd@mahidol.ac.th 

Owing to the need of new antibacterial therapy to 

overcome the problem of antibiotic resistance for emergence and 

re-emergence of infections diseases, there is an urgent need to 

identify new target and to discover new lead compound which 

has potential to develop as antibacterial drug. We are interested 

in protein that plays a crucial role in bacterial cell division, named 

FtsZ. In this study, we focus our attention on protein FtsZ from 

Mycobacterium tuberculosis. Previous report shows that 

compound 2-Alkoxycarbonylaminopyridines SRI-3072 inhibits 

FtsZ polymerization and growth of M. tuberculosis H37Rv 

without any perturbation on polymerization of bovine brain 

tubulin. Thus, SRI-3072 might be developed into new antituberculosis 

drugs effective against the current multidrug-resistant 

strains of M. tuberculosis. However, there is no structural 

information available for the mode of interaction between SRI- 

3072 and M. tuberculosis FtsZ at atomic level. Recently, the 

crystal structure of FtsZ from M. tuberculosis H37Rv is deposited 

in Protein Data Bank (PDB) with accession code 2Q1Y. 

Accordingly, the interaction of SRI-3072 with catalytic core 

domain of M. tuberculosis FtsZ can be deduced by molecular 

modeling calculations including computational docking and 

molecular dynamics simulation. Our predicted structural data 

suggests the plausible binding site and binding mode of SRI- 

3072 in M. tuberculosis FtsZ. The docking scores, hydrophobic, 

hydrogen bond donor and hydrogen bond acceptor were analyzed 

in order to identify the structural features of SRI-3072 involved 

in binding interactions with active site FtsZ-residues. Trajectory 

obtained from molecular dynamics simulation were further 

analysed to validate the binding mode of SRI-3072 in M. 

tuberculosis FtsZ. Our results shed light on the structural 

information of M. tuberculosis FtsZ and its inhibitor which can


be used for the design of potent anti-FtsZ agents as well as a 

virtual tool for screening of new lead from chemical library in 

the future. 

Key words: FtsZ, mycobacterium, Antibiotics 

IN VITRO ASSAY OF ANTI-DENGUE VIRUS 

ACTIVITY FROM THAI MEDICINAL PLANT 

EXTRACTS (NO. 0734) 

Napatsanan Klawikkan 1 , Veena Nukoolkarn 2 , Suthee Yoksan 3 , 

Chanpen Wiwat 4 and Krit Thirapanmethee 5 


University, Sri-ayudhya road, Bangkok 10400, Thailand; 



Thailand, E-mail: pyvnk@mahidol.ac.th; 3 Center for Vaccine 

Development, Mahidol University; 4 Department of 



pycww@mahidol.ac.th; 5 Department of Microbiology, 


Bangkok 10400, Thailand, E-mail: pyktr@mahidol.ac.th 

Dengue fever (DF) and Dengue hemorrhagic fever 

(DHF) are becoming increasingly an important public health 

problem in the tropical and subtropical regions. Dengue has been 

recognised in over 100 countries and 2-5 billion people live in 

areas where dengue is endemic. The diseases are caused by dengue 

virus (DENV) which is belonged to Flavivirus genus of the 

Flavividae family. It exists as four related serotypes (DENV1, 2, 

3 and 4). At present, neither a licensed vaccine nor antiviral drug 

are available to control dengue disease. Thus, the aim of this 

study is to investigate the in vitro inhibitory activity of Thai 

medicinal plants toward DENV2 infection in Vero cells. For this 

purpose, ten medicinal plants were collected from Siri 

Ruckhachati Natural Park, Salaya campus, Mahidol University. 

Subsequently, these plants were extracted with dichloromethane 

and ethanol. All extracts were evaluated for their anti-dengue 

activity in Vero cell using MTT method. The results showed that 

all dichloromethane extracts had no activity against DENV2. 

While ethanol extracts of Rhizophora apiculata Blume., 

Flagellaria indica Linn. and Cladogynos orientalis Zipp. at a 

concentration 12.5 µg/ml exhibited inhibitory activity on DENV2 

with 56.14%, 45.52% and 34.85 %, respectively. In addition, 

Houttuynia cordata Thunb.exhibited inhibition activity on 

DENV2 with 35.99% at a concentration of 1.56 µg/ml. These 

results indicated that ethanol crude extracts of Rhizophora 

apiculata Blume., Flagellaria indica Linn., Cladogynos 

orientalis Zipp. and Houttuynia cordata Thunb. have an in vitro 

inhibitory activity against DEN2. 

Key words: Dengue virus type 2; Thai medicinal plants; Antidengue 

activity 

DISTRIBUTION, BIOACTIVE COMPONENTS 

AND BIOLOGICAL ACTIVITIES OF STEMONA 

SPECIES IN THAILAND (NO. 0735) 

Sumet Kongkiatpaiboon and Wandee Gritsanapan 

Department of Pharmacognosy, Faculty of Pharmacy, 

Mahidol University, Bangkok, 10400, Thailand, E-mail: 

pywgs@mahidol.ac.th 

Stemona species are monocotyledonous plants in the 

family Stemonaceae occurring as twining herbs with perennial 

tuberous roots. They are widely used in Thailand and southeast 

Asian countries for their insecticidal properties and as antitussive 

remedy. Stemona plants in Thailand comprise 9 known species, 

i.e. S. aphylla Craib, S. burkilliiPrain, S. cochinchinensis 

Gagnep., S. collinsiae Craib, S. curtisii Hook.f., S. kerrii Craib, 

S. phyllantha Gagnep.,S. pierrei Gagnep. and S. tuberosa Lour., 

and 2 unidentified species. Infact, they can be separated into 2 

main groups according to their morphological characters and 

bioactive components accumulation i.e. tuberosa and nontuberosa 

groups. Tuberosa group has large and thick tuber, 

comprising S. tuberosa and S. phyllantha. Non-tuberosa group 

is composed of curtisii group (S. curtisii, S. collinsiae, S. aphylla 

and S. burkillii) which has long and slender tuber, and kerrii 

group (S. kerrii, S. cochinchinensis and S. pierrei) which has 

short and small tuber. However, similar shape of tuberous roots 

from different species of Stemona in Thailand markets is called 

the same name as “Non Tai Yak”, making taxonomic confusion 

in some literatures. Phytochemical investigations of Stemona 

species have revealed the presence of alkaloids, stilbenoids, 

tocopherols (chromenols), etc. Stemona alkaloids constitute a 

unique chemical feature of the Stemonaceae and cannot be 

detected so far in any other plant families.Their alkaloid structures 

are characterized by a pyrrolo[1,2-a]azepine core which can be 

classified into three types based on biosynthetic consideration 

i.e. stichoneurine- (tuberostemonine-), protostemonine- and 

croomine-types of alkaloids. Tuberosa group contains 

stichoneurine- and croomine-types, demonstrating antitussive 

activity, while non-tuberosa group contains protostemonine type 

alkaloid which promotes potent insecticidal activity. Biological 

testings showed that insecticide and antitussive activities of 

Stemona are depended on Stemona alkaloids while antifungal 

activity against plant-pathogenic fungi is depended on the 

presence of stilbenoids. Moreover, preliminary tests on anticancer 

and antioxidant activities of some Stemona species have been 

reported. 

Key words : Stemona, Stemonaceae, Non Tai Yak 

(Published in Medicinal Plants 2010;2(1), 1-12) 

Grant: The Thailand Research Fund (Royal Golden Jubilee 

Ph.D. Program) and Mahidol University 

ANTI-ACNE-INDUCING BACTERIAL ACTIVITY 

OF MANGOSTEEN FRUIT RIND EXTRACT 

(NO. 0736) 

Werayut Pothitirat 1,3 , Mullika Traidej Chomnawang 2 , 

Wandee Gritsanapan 1

296 

1 Departments of Pharmacognosy, Faculty of Pharmacy, 

Mahidol University; 2 Department of Microbiology, Faculty 

of Pharmacy, Mahidol University; 3 Faculty of Pharmacy, 

Siam University, Bangkok, Thailand. E-mail: 


Objective: The aims of this study were to determine 

the most effective solvent extract of mangosteen, anti-acne 

inducing bacterial activity and the amount of α-mangostin, a 

major active component in each mangosteen fruit rind extract, 

using high-performance liquid chromatography (HPLC). 

Materials and Methods: The fruit rinds of mangosteen were 

extracted with hexane, dichloromethane, ethanol and water. The 

extracts were tested for antibacterial activity against bacteria that 

induce acne, including Propionibacterium acnes and 

Staphylococcus epidermidis . Thin-layer chromatographic 

autobiography against these bacteria was also performed for each 

extract, while the α -mangostin content was analyzed using a 

validated HPLC method. Results: The dichloromethane extract 

exhibited the strongest antibacterial effect with minimum 

inhibitory concentration values for both bacterial species at 3.91 

μg/ml, while the minimum bactericidal concentration values 

against P. acnes and S. epidermidis were 3.91 and 15.63 μg/ml, 

respectively. Thin-layer chromatographic autobiography indicated 

that α-mangostin was present in all extracts, except the water 

extract, and is a major active component against both P. acnes 

and S. epidermidis . Using HPLC, the dichloromethane extract 

yielded the highest content (46.21% w/w) of α -mangostin 

followed by the ethanol extract (18.03% w/w), the hexane extract 

(17.21% w/w) and the water extract (0.54% w/w). Conclusions: 

Dichloromethane extract exhibited the strongest anti-acneinducing 

bacterial effect and this extract yielded the highest 

amount of α -mangostin. 

Key words: Acne vulgaris, anti-acne , mangosteen 

(Published in Med Princ Pract 2010;19:281–286) 

Grant: Mahidol University 

QUANTITATIVE ANALYSIS AND TOXICITY 

DETERMINATION OF ARTIFACTS ORIGINATED 

IN A THAI TRADITIONAL MEDICINE 

PRASAPLAI (NO. 0737) 

Prasan Tangyuenyongwatana, and Wandee Gritsanapan 


Mahidol University, Bangkok, Thailand, E-mail: 


Prasaplai is a Thai traditional medicine for relieving 

dysmenorrhea and adjusting the menstrual cycle. Three fatty acid 

esters, (E)-4-(3,4-dimethoxyphenyl)but-3-en-1-yl linoleate(1), 

(E)-4-(3,4-dimethoxyphenyl)but-3-en-1-yl oleate (2) and (E)-4- 

(3,4-dimethoxyphenyl)but-3-en-1-yl palmitate (3) are formed 

dur-ing storage from the reaction of chemical components in two 

herbs, i.e., fatty acids in Nigella sativa (L.) (Ranunculaceae) and 

(E)-4-(3,4-dimethoxyphenyl)but-3-en-1-ol (compound D) in 

Zingiber cassumunar (Roxb.) (Zingiberaceae). The formations 

of these artifacts were monitored for 1 year and their amounts 

were analyzed by HPLC at certain periods of time. The results 


showed that artifact formation was satu-rated after 73 days of 

storage. The amount of each artifact in the saturation period 

ranged from 3.93 ± 0.06 to 4.30 ± 0.18% w/w for compound 1, 

1.69 ± 0.08 to 1.9 ± 0.13% w/w for compound 2 and 0.09 ± 

0.003 to 0.1 ± 0.005% w/w for compound 3. Cytotoxicity of the 

artifacts was evaluated using NCI-H187, KB, and BC cancer cell 

lines and found that the IC 50 of all artifacts in all tests were higher 

than 20 ìg/mL. For acute toxic-ity in mice, the LD 50 of each 

artifact was more than 300 mg/kg. 

Key words: Artifact formation, Prasaplai, fatty acid ester 

(Published in Pharmaceutical Biology, 2010; 48(5): 584–588) 

FREE RADICAL SCAVENGING AND ANTI- 

ACNE ACTIVITY OF MANGOSTEEN FRUIT 

RIND EXTRACTS PREPARED BY DIFFERENT 

EXTRACTION METHODS (NO. 0738) 

Werayut Pothitirat 1 , Mullika Traidej Chomnawang 2 , 

Roongtawan Supabphol 3 , and Wandee Gritsanapan 1 


Mahidol University, Bangkok,Thailand; 2 Department of 

Microbiology, Faculty of Pharmacy, Mahidol University, 

Bangkok, Thailand; 3 Department of Physiology, Faculty of 

Medicine, Srinakarintarawirot University, Bangkok, 

Thailand, E-mail: pywgs@mahidol.ac.th 

Key words: Acne vulgaris, anti-acne, Garcinia mangostana 

The ethanol extracts of mangosteen fruit rinds prepared 

by several extraction methods were examined for their contents 

of bioactive compounds, DPPH-scavenging activity, and antiacne 

producing bacteria against Propionibacterium acnes and 

Staphylococcus epidermidis. The dried powder of the fruit rind 

was extracted with 95% ethanol by maceration, percolation, 

Soxhlet extraction, ultrasonic extraction, and extraction using a 

magnetic stirrer. Soxhlet extraction promoted the maximum 

contents of crude extract (26.60% dry weight) and α-mangostin 

(13.51%, w/w of crude extract), and also gave the highest antiacne 

activity with MIC 7.81 and 15.63 μg/mL and MBC 15.53 

and 31.25 μg/mL against P. acnes and S. epidermidis, respectively. 

Ethanol 70% and 50% (v/v) were also compared in Soxhlet 

extraction. Ethanol 50% promoted the extract with maximum 

amounts of total phenolic compounds (26.96 g gallic acid 

equivalents/100 g extract) and total tannins (46.83 g tannic acid 

equivalents/100 g extract), and also exhibited the most effective 

DPPH-scavenging activity (EC50 12.84 μg/mL). Considering 

various factors involved in the process, Soxhlet extraction carried 

a low cost in terms of reagents and extraction time. It appears to 

be the recommended extraction method for mangosteen fruit rind. 

Ethanol 50% should be the appropriate solvent for extracting free 

radical-scavenging components, phenolic compounds, and 

tannins, while 95% ethanol is recommended for extraction of αmangostin, 

a major anti-acne component from this plant. 

Key words: Acne vulgaris, anti-acne, Garcinia mangostana 

(Published in Pharmaceutical Biology, 2010; 48(2): 182–186) 

Grant: Mahidol University


IN VITRO ANTI-PROLIFERATIVE ACTIVITY 

OF ALCOHOLIC STEM EXTRACT OF 

COSCINIUM FENESTRATUM IN HUMAN 

COLORECTAL CANCER CELLS (NO. 0739) 

Piyanuch Rojsanga 1,2 , Mugdha Sukhthankar 1 , Chutwadee 

Krisanapun 1 , Wandee Gritsanapan 2 , Darunee Buripakdi 

Lawson 3 and Seung Joon Baek 1 

1 Department of Pathobiology, College of Veterinary Medicine, 

University of Tennessee, Knoxville, USA; 2 Department of 


Bangkok, E-mail: pywgs@mahidol.ac.th 3 Faculty of 

Veterinary Science, Nakornpathom, Thailand, 

Coscinium fenestratum (Gaertn.) Colebr. is 

traditionally used for the treatment of cancer, arthritis and diabetes 

mellitus. The purpose of this study was to determine the molecular 

mechanisms by which this plant shows beneficial effects. An 80% 

ethanolic extract of C. fenestratum (80ET) was separated by its 

polarity into dichloromethane (DCM) and aqueous fractions 

(WF), and the anti-proliferative effects of 80ET, DCMand WF 

were investigated. Berberine, one of the major components of C. 

fenestratum, was used as a control. The 80ET, DCM, WF and 

berberine showed anti-proliferative activity as assessed by cell 

growth assay. Subsequently, the pro-apoptotic proteins NAG-1 

and ATF3 were increased and the cell cycle protein cyclin D1 

was decreased by the extract and its fractions. Interestingly, only 

the fraction exhibited the induction of peroxisome proliferatoractivated 

receptor ã (PPARã) binding activity, which represents 

a pro-apoptotic activity in colorectal cancer cells. The overall 

results of this study indicate that the extract from this plant has 

anti-proliferative activity through the activation of pro-apoptotic 

proteins and PPARã, and may have potential as a preventive 

regimen in the treatment of cancer. 

Key words: Coscinium fenestratum, anti-proliferative, berberine 

(Published in Experimental and therapeutic medicine 1: 181- 

186, 2010) 



PHYTOCHEMICAL AND ANTIOXIDANT 

STADIES OF LAURERA BENGUELENSIS 

GROWING IN THAILAND (NO. 0740) 

Nedeljko T. Manojlovic 1 , Perica J. Vasiljevic 2 , Wandee 

Gritsanapan 3 , Roongtawan Supabphol 4 and Ivana 

Manojlovic 2 

1Department of Pharmacy, Medical Faculty, University of 

Kragujevac, Kragujevac, Serbia, e-mail: ntm@kg.ac.rs; 

2Department of Biology, Faculty of Science, University of Nis, 

Nis, Serbia; 3Department of Pharmacognosy, Faculty of 

Pharmacy, Mahidol University, Bangkok, Thailand, 

4Department of Physiology, Faculty of Medicine, 

Srinakarinwirote University, Thailand 

The aim of this study was to investigate metabolites 

of the lichen Laurera benguelensis. A high-performance liquid 

chromatographic (HPLC) method has been developed for the 

characterization of xanthones and anthraquinones in extracts of 

this lichen. Lichexanthone, secalonic acid D, norlichexanthon, 

parietin, emodin, teloschistin and citreorosein were detected in 

the lichen samples, which were collected from two places in 

Thailand. Components of the lichen were identifed by relative 

retention time and spectral data. This is the frst time that a detailed 

phytochemical analysis of the lichen L.benguelensis was reported 

and this paper has chemotaxonomic signifcance because very 

little has been published on the secondary metabolites present in 

Laurera species. Some of the metabolites were detected for the 

frst time in the family Trypetheliaceae. The results of preliminary 

testing of benzene extract and its chloroform and methanol 

fractions showed that all samples showed a weak radical 

scavenging activity. The chloroform extract showed the highest 

antioxidant activity. 

Key words: anthraquinones, antioxidant activity, Laurera 

benguelensis, 

(Published in Biological Research 43(2) 169-176, 2010) 

MICROSCOPIC CHARACTERIZATION AS A 

TOOL FOR SEPARATION OF STEMONA 

GROUPS (NO. 0741) 

Sumet Kongkiatpaiboon 1 ,Vichien Keeratinijakal 2,3 and 

Wandee Gritsanapan 1 



pywgs@mahidol.ac.th; 2 National Center for Agricultural 

Biotechnology, Kasetsart University, Bangkok, 3 Agronomy 

Department, Faculty of Agriculture, Kasetsart University, 

Bangkok, Thailand 

Introduction: Stemona plants have been traditionally 

used as an insecticide, scabicide and pediculocide, and for the 

treatment of skin and respiratory diseases. Stemona can be 

separated into two groups according to their morphological 

characters and bioactive components i.e. stichoneurine and 

protostemonine groups. Protostemonine group contains alkaloids 

that possess potent insecticidal activity while stichoneurine group 

accumulates alkaloids with antitussive activity. In Thailand, a 

vernacular name “Non Tai Yak” refers to the roots of different 

species of Stemona, making it confusing to discern different 

species. The purposes of this study are to investigate the 

microscopic characteristics of the roots of seven species of 

Stemona growing in Thailand and to distinguish and identify these 

groups of Stemona. Methods: Cross-sectional histology of fresh 

root samples and powdered drug characteristics of 7 species of 

Stemona were studied under a microscope. Results: The roots of 

Stemona in the stichoneurine group (S. tuberosa andS. phyllantha) 

contained a non-lignified large pith while the roots of 

protostemonine group (S. burkillii, S. cochinchinensis, S. 

collinsiae, S. curtisii and S. kerrii) had a small lignified one. 

The powder of stichoneurine group contained numerous thinwalled 

parenchyma, but only few thick-walled parenchyma and 

lignified fibers and vessels were present. In contrast, thick-walled 

parenchyma and lignified fibers and vessels were frequently found 

in the powdered roots of protostemonine Stemona. These 

characteristics could be used to discern between Stemona in the 

stichoneurine and protostemonine groups. Conclusions: The

298 

microscopic characterizations can be used as a primary tool to 

categorize and separate 2 main Stemona groups. 

Key words: Non Tai Yak, protostemonine, Stemona 

(Published in Pharmacognocy Journal 2 (17) 2010, 1-4.) 



IN VITRO EVALUATION OF ANTIFUNGAL 

ACTIVITY OF ANTHRAQUINONE DERIVA- 

TIVES OF SENNA ALATA (NO. 0742) 

Mansuang Wuthi-udomlert 1 , Pavena Kupittayanant 2 and 

Wandee Gritsanapan 2* 

1 Department of Microbiology, Faculty of Pharmacy,Mahidol 

University, Bangkok 10400, Thailand; 2 Department of 


Bangkok 10400, Thailand *e-mail: pywgs@mahidol.ac.th 

Senna alata leaf powder was used to obtain five 

extracts which contain anthraquinone compounds in different 

forms i.e. anthraquinone aglycone extract,anthraquinone 

glycoside extract, anthraquinone aglycones from glycosidic 

fraction, crude ethanol extract, and anthraquinone aglycone from 

crude ethanol extract. All extracts were tested against clinical 

strain of dermatophytes: Trichophyton rubrum, T. 

mentagrophytes, Epidermophyton floccosum, and Microsporum 

gypseum by diffusion and broth dilution techniques to find out 

the active form for antifungal activity. Thin layer chromatography 

was developed to demonstrate the fingerprints of chemical 

constituents of each extract. This investigation pointed out the 

best in vitro antifungal activity of anthraquinone aglycones from 

glycosidic fraction qualitatively and quantitatively, compared to 

other extracts. 

Key words: anthraquinone, antifungal, Cassia alata 

(Published in J Health Res 2010, 24(3): 117-122) 

Grant: The Thailand Research Fund (IRPUS) 

INHIBITORY EFFECTS OF N-ACETYLCY- 

STEINE ON PROLIFERATIVE, INVASIVE 

AND MIGRATORY CAPABILITIES OF 

HUMAN BLADDER CANCER CELLS (NO. 0743) 

Athikom Supabphol 1 , Verasing Muangman 2 , Warinthorn 

Chavasiri 3 , Roongtawan Supabphol 4 , Wandee Gritsanapan 5 

1 Department of Surgery, Faculty of Medicine, 

Srinakharinwirot University, Bangkok, Thailand; 

2 Department of Surgery, Vichaiyut Hospital and Medical 

Center, Bangkok, Thailand, 3 Department of Chemistry, 

Faculty of Science, Chulalongkorn University, Bangkok, 

Thailand, 4 Department of Physiology, Faculty of Medicine, 

Srinakharinwirot University, Bangkok, Thailand; 


Mahidol University, Bangkok, ThailandE-mail: 



Purpose: One of the diseases that creates the major 

clinical impact for Thai urologists along with its burden on public 

health and economically issues for the Royal Thai government 

and patients is the bladder cancer. Our aim was to study the 

inhibitory effects of N-acetylcysteine (NAC), a cheap, safe and 

widely used over-the-counter-drug in Thailand, on the 

proliferative, invasive and migratory capabilities of human 

bladder cancer cells. Methods: Effects of NAC at concentrations 

between 0-40 mM, on the proliferation of human bladder cancer 

cells were determined by using 3-(4,5-dimethylthiazol-2-yl)-2,5diphenyl 

tetrazolium bromide (MTT) assay. Chemotaxis chamber 

was used to assess the effects of NAC on the invasive and 

migratory capabilities of the bladder cancer cells. Results: NAC 

could directly and significantly suppressed the proliferative, 

invasive and migratory capabilities of human bladder cancer cells 

in a dose-dependent fashion. The 50% inhibitory concentration 

(IC50) value for cell proliferation, invasion and migration were 

33.33 + 0.78, 22.20 + 0.85 and 12.82 + 0.20 mM, respectively. 

Conclusion: The proliferation, migration, and invasion constitute 

the cellular mechanisms of cancer cell metastasis. Our study 

significantly showed those three inhibitory effects of NAC on 

human bladder cancer cells. This could imply that NAC has high 

potential in inhibiting bladder cancer cell metastasis. The 

urologists may apply our results to use NAC as the adjuvant 

therapy by intravesical instillation before, during and/or after the 

transurethral resection of bladder tumor (TURBT) with or without 

oral or parenteral NAC supplementation in order to lessen the 

severity of the disease. 

Key words: NAC, N-acetylcysteine, bladder cancer 

(Published in J Med Assoc Thai 2009; 92 (9): 1171-1177.) 

INVESTIGATION OF THAI TRADITIONAL 

WAY FOR DETOXIFICATION OF ALOE 

(NO. 0744) 

Prasan Tangyuenyongwatana 1 and Wandee Gritsanapan 2 

1 Faculty of Oriental Medicine, Rangsit University, 

Pathumtani, Thailand 2 Department of Pharmacognosy, 

Faculty of Pharmacy, Mahidol University, Bangkok, 

Thailand. E-mail : pywgs@mahidol.ac.th 

In Thai traditional medicine, dried juice extract of Aloe 

vera (L.) Burm has been used in many formulations for mild 

laxative effect for a long time. The Thai traditional way 

recommends detoxifying the dried juice extract before combining 

with other herbal constituents. The reason for detoxification is to 

decrease the gastrointestinal tract irritation of aloe. The traditional 

method to detoxify aloe is adding small amount of water into the 

dried juice extract and heating until the dried and crispy mixture 

is performed. The objective of this study is to verify the chemical 

conversion of major ingredients in aloe dried juice extract using 

HPTLC analysis (Silica gel 60 GF254, solvent system; ethyl 

acetate : methanol : water 100 : 13.5 : 10, detection at 420 nm). 

Our study found that the amount of a major compound aloin (Rf 

0.34) in the dried juice extract was decreased from to after 

heating while the amount of its aglycone aloe-emodin (Rf 0.70) 

was increased from 50 µg/mL to 473.2 µg/mL. This result explains 

that traditional herbalists have known by their experiences in 

detoxifying aloe by heating the dried juice extract with small


amount of water. By this way, the hydrolysis and oxidation 

reactions can occur and some aloin is conversed to aloe-emodin 

resulted in decreasing anthraquinone glycoside intensity, which 

can cause irritation of the gastrointestinal tract and increase the 

chance for gripping action [2]. This study gives the scientific 

support for detoxification of selected herbal ingredients in Thai 

traditional medicines. The further study of others glycosides and 

aglycones in the dried juice extract of aloe is in our progress. 

Key words: Aloe vera, detoxify, anthraquinone 

(Presented at 58 th International Congress and Annual Meeting 

of the Society for Medicinal Plant and Natural Product 

Research, 29 August-2 September 2010, Berlin. Published in 

Planta Medica 2010: 76: 1216) 

ANTHRAQUINONE CONTENT AND TOXICITY 

TEST OF CASSIA FISTULA POD EXTRACTS 

(NO. 0745) 

Gritsanapan W, Sakulpanich A, Thongpraditchote S 

Faculty of Pharmacy, Mahidol University, 447 SriAyudthaya 

Road, Ratchatewi Bangkok, Thailand, E-mail: 


Cassia fistula Linn. (Caesalpiniaceae) can be easily 

found in all parts of Thailand as an ornamental plant and its ripe 

pods are almost treated as a waste. In Thai traditional medicine, 

the ripe pods of C. fistula have long been used as a laxative drug. 

They contain several anthraquinones such as rhein, aloe-emodin 

and sennosides, These anthraquinones promote laxative effect of 

which their glycosides contents indicate the laxative potency. In 

this study, the ripe pods of C. fistula were collected from 10 

different provinces of Thailand. The pod pulp was separated and 

extracted with distilled water by decoction. The water extracts 

were determined for the contents of total anthraquinones and total 

anthraquinone glycosides by a UVvis spectrophotometric method 

at 515 nm and the acute toxicity of the extracts was investigated 

in mice and rats. Extract ratio (crude drug: 1g crude extract) of 

all the extracts are 12:1. The contents of total anthraquinones 

and total anthraquinone glycosides in the extracts calculated as a 

major anthraquinone rhein were 1.451.85% w/w (average 1.63% 

w/w) and 0.380.71% w/w (average 0.53% w/w), respectively 

while in the fresh pod pulp contained 0.891.03% w/w (average 

0.94% w/w) and 0.220.39% w/w (average 0.31% w/w), 

respectively. After tested mice and rats were administered with 

the extract at dose level 5 g/kg, no mortality or any sign of toxicity 

were found within 14 days. The body weight gain of all animals 

was not significantly different between the treated groups and 

the respective control group. This indicats that C. fistula pod 

extract was grouped in a slightly toxic (LD50 >5g/kg). In Thai 

traditional medicine, the fresh pod pulp 48g has been used as a 

dose for laxative drug. This dose amount was equal to 37.6752 

mg of total anthraquinones and 12.4248 mg of total anthraquinone 

glycosides. From our preliminary study, it suggests that the 

decoction extract of C. fistula pod might be used as an alternative 

source of natural laxative drugs and should be further developed 

as a modern pharmaceutical laxative preparation. 

Key words: C. fistula, anthraquinone, laxative 





HEMOSTATIC PROPERTY OF SIAM WEED 

EXTRACTS (NO. 0746) 

Hataichanok Pandith 1 , Suchitra Thongpraditchote 2 , Yuvadee 

Wongkrajang 2 , Kanjana Muangklum 3 , Wandee Gritsanapan 1* 


Mahidol University; 2 Department of Physiology, Faculty of 

Pharmacy, Mahidol University, Bangkok, Thailand, 

3 Department of Pathology, Faculty of Medicine Siriraj 

Hospital, Mahidol University, Bangkok 10700, Thailand. Email 

: pywgs@mahidol.ac.th 

Chromolaena odorata (L.) King and Robinson or Siam 

weed is a perennial scandent or semi-woody shrub of the family 

Asteraceae. It has been used as a medicinal herb for variety of 

ailments in Tropical Africa and Asia. In Thailand, the fresh leaves 

are widely used in rural areas as a haemostatic agent to stop 

bleeding. Many studies both in vitro and in vivo have 

demonstrated that C. odorata extracts can enhance hemostasis. 

This study was investigated to find the appropriate extraction 

method and solvent which yield the extract with the highest 

hemostatic activity. The lyophilized aqueous fresh mature leaf 

extract and the extracts from maceration of fresh and dried mature 

leaves by various solvents (50, 70, and 95% v/v ethanol) were 

tested for hemostatic activity in vivo. The bleeding time was 

studied in 4 weeks old male Wistar rats. The incision was made 

at a foot pad of each rat following by applying the 20 µl of each 

C. odorata extract to the wound. Bleeding time was recorded 

immediately after making the wound until the blood stop. It was 

found that 70% ethanolic extract of dried mature leaves could 

reduce the bleeding time more than any others. This result showed 

that maceration of dried samples with 70% ethanol should be the 

appropriate extraction method which yielded the extract with the 

highest hemostatic activity. Consequently, the mechanisms of 

hemostatic activity of C. odorata were investigated using blood 

coagulation and platelet aggregation in vitro tests. All of aqueous 

and alcoholic extracts of fresh and dried mature leaves were 

investigated for blood coagulation test by coagulogram study 

including APTT (activated thromboplastin time) and PT 

(prothrombin time) tests. Only 70% ethanolic extract of dried 

leaves was investigated for platelet aggregation. Sheep plasma 

was used in both tests. It was found that all extracts neither clotted 

plasma nor increased the platelet aggregation. This result showed 

that sheep plasma might not be suitable for investigation of 

hemostatic mechanisms. 

Key words: Chromolaena odorata, platelet aggregation, 

hemostasis 

(Presented at The First UKM-MU Joint Scientific Conference 

2010, October 12 13, 2010, Selangor, Malaysia.) 

Grant: Office of The Higher Education Commission

300 

CONTENT OF BIOLOGICALLY ACTIVE 

COMPONENTS IN VEGETATIVE EDIBLE PARTS 

OF SENNA SIAMEA (NO. 0747) 

Wandee Gritsanapan 1 , Aurapa Sakulpanich 1,2 , Tanit 

Padumanonda 1,3 


Mahidol University, Bangkok, Thailand, 2 Department of 

Pharmacognosy, Faculty of Pharmaceutical Sciences, 

Huachiew Chalermprakeit University, Bangpli, 

Samutprakarn, Thailand, 3 Faculty of Pharmaceutical 

Sciences, KhonKaen University, KhonKaen, Thailand. Email: 


Senna siamea (Lam.) Irwin & Barneby of the family 

Fabaceae is a well-known medicinal plant in Thailand and Asian 

countries. The fresh young leaves and/or young flowers of this 

plant have been used as vegetables in a popular Thai food called 

“khi lek curry” which promotes mild laxative and sleeping aid 

effects. The active constituent for sedative action is barakol while 

glycosides of anthraquinones such as rhein and aloe-emodin are 

laxative components. This work determined and compared the 

contents of barakol and total anthraquinone glycosides in young 

leaves, young flowers and mature leaves of this plant using 

validated TLC-densitometric and UV-vis spectrophotometric 

methods, respectively. Barakol content in the young leaves, young 

flowers and mature leaves were 1.67, 1.43 and 0.78% w/w of 

dried powder, respectively while the content of total 

anthraquinone glycosides, calculated as rhein, were 104.6, 12.4 

and 8.9 mg% dry weight, respectively. Therefore, young leaves 

contain biologically active components > young flowers > mature 

leaves. This can explain a reason of using young leaves and young 

flowers (not mature leaves) of S. siamea as vegetables for cooking 

as food and medicinal purposes. 

Key words: Senna siamea, anthraquinone, khi lek 

(Presented at The First UKM-MU Joint Scientific Conference 

2010, October 12 13, 2010, Selangor, Malaysia.) 

ASSESSMENT OF THE PHENOLIC CONTENT 

ANDFREE RADICAL SCAVENGING CAPACITY 

OF EXTRACTS OBTAINED FROM THE 

PERICARP OF GARCINIA MANGOSTENA L. 

Pothitirat W 1,2 , Gritsanapan W 2 

(NO. 0748) 

1 Faculty of Pharmacy, Siam University, Bangkok, Thailand; 

2 Department of Phamacognosy, Faculty of Pharmacy, 

Mahidol University, Bangkok, Thailand. E-mail: 


The pericarp of Garcinia mangostana . is popularly 

used as a food supplement due to its high amount of phenolic 

compounds i. e., xanthones, flavonoids and tannins which 

promote high free radical scavenging activity. This work was 

designed to examine the amount of phenolic constituents and 

the free radical scavenging capacity of extracts obtained from 

the peicarp of G. mangostana collected from 15 different locations 


in Thailand. The 95% ethanolic extract were prepared by soxhlet 

extraction. The average content of total phenolic compounds, total 

tannins and total flavonoids per 100 grams of extract in all 

samples were found to be 22.33± 3.25 g of gallic acid equivalent, 

36.38 ± 8.46 g of tannic acid equivalent, and 4.13 ±1.10 g of 

quercetin equivalents evaluated using Folin – Ciocalteu 

procedure, protein precipitation method, and aluminium 

colorimetric method, respectively. The average EC50 of all 

extracts was found to be 17.59± 5.69μg/ml determined by the 

DPPH scavenging method. The regression analysis between the 

amount of total phenolics and total tannins in the extract exhibited 

a high positive relationship (r=0.9302), while the correlation 

coefficient between antiradical activity (1/ EC50) and the content 

of total phenolic and total tannins of all ethanolic extract were 

both higher than 0.8, indicating a significant positive relationship 

between these parameters. Therefore, phenolic compounds and 

tannins seem to play an important role as free radical scavengers 

in the pericarp extract of G. mangostana. 

Key words: Garcinia mangostan, DPPH, free radical scavenging 

capacity 

(Presented at 58th International Congress and Annual Meeting 




Grant: Mahidol University 

DEVELOPMENT OF WHITENING SKIN 

LOTION FROM SELECTED MEDICINAL 

PLANTS AND ITS ANTITYROSINASE 

ACTIVITY (NO. 0749) 

Pothitirat W 1 , Pluemlamai J 2 , Satniyom S 2 , Leelamanitiya W 2 , 

Gritsanapan W 2 





In Thailand, herbal cosmetics are becoming more 

popular especially for nourishing and ±. In this study, Artocarpus 

lakoocha Roxb. (AL), Glycyrrhiza glabra Linn. (GG) and 

Bombyx mori Linn. (BM) were extracted and formulated as a 

herbal whitening skin lotion. Antityrosinase activity of each 

extract and of the lotion preparation were tested using the 

dopamine method [1]. Additionally, the stability study; tyrosinase 

inhibitory activity, TLC, colour and odour of the formulation kept 

at different temparatures (4-8, 25-28 and 45°C) for 4 weeks were 

studied. The percentage of tyrosinase inhibition of AL, GG and 

BM at concentration of 1 mg/ml were found to be 87.88+ - 0.31 

(IC50=50 μg/ml), 74.24 ± 0.62 (IC50 = 140μg/ml) and 5.19 ± 

0.61 (IC50>1000μg/ml), respectively, while the activity of a 

reference standard kojic acid at the same concentration was 83.30 

± 0.51% (IC50=130μg/ml). The whitening skin lotion containing 

1.03% of AL, 0.42% of GG and 0.20% w/w of BM, exhibited 

antityrosinase activity at 98.60 ± 0.45%. During 4 weeks of 

storage at 4 C, the antityrosinase activity, TLC, colour and odour 

of the lotion were not changed. Thus, the preparation containing 

AL, GG and BM which promoted high antityrasinase activity 

was appropriate as a herbal whitening skin lotion and should be 

stored at 4°C for good stability.


Key words: Antityrosinase activity, Artocarpus lakoocha, 

Glycyrrhiza glabra 



Research, 29 August-2 September 2010, Berlin, Published in 


IN SILICO STRATEGY FOR THE IDENTIFI- 

CATION OF CYCLOOXYGENASE INHIBITORS 

FROM THE THAI MEDICINAL MIXTURE 

PRASAPLAI (NO. 0750) 

Waltenberger B 1 , Schuster D 2 , Paramapojn S 3 , Gritsanapan 

W 3 , Wolber G 2 , Rollinger J 1 , Stuppner H 1 

1 Institute of pharmacy, Pharmacognocy and center for 

Molecular Biosciences, University of Innsbruck, Austria; 

2 Computer-Aided Molecular Design Group, Institute of 

Pharmacy, Pharmaceutical Chemistry and center for 

Molecular Biosciences, University of Innsbruck, Austria; 

3 Department of Phamacognosy, Faculty of Pharmacy, 

Mahidol University, Bangkok, Thailand 

Prasaplai is a medicinal plant mixture that is used in 

Thailand to treat primary dysmenorrhea which is characterized 

by painful uterine contractility and caused by a significant 

increase of prostaglandin release Cyclooxygenase (COX) 

represents a key enzyme in the formation of prostaglandins. 

Former studies revealed that extracts of Prasaplai inhibited COX- 

1 and COX-2 . The major aim of this study was to predict which 

compounds of the complex mixture of natural products might be 

responsible for the COX inhibitory activity. Therefore, a 

comprehensive literature survey for known constituents of 

Prasplai was performed. A multiconformational 3D database was 

generated comprising 683 molecules. Virtual parallel screening 

using a set of six validated pharmacophore models for COX 

inhibitors was performed resulting in a hit list of 166 compounds. 

46 Prasaplai compounds with already determined COX activity 

were used for the external validation of this set of COX 

pharmacophore models. 57% of these components were predicted 

correctly by the pharmacophore models, i.e. virtual hits with 

inhibitory activity on COX as well as inactive non-hits. The 

findings of this theoretical study confirm that the virtual approach 

provides a helpful tool for the fast identification of novel COX 

inhibitors 

Key words: virtual approach, COX inhibitor, Prasaplai 

(Presented at 58th International Congress and Annual Meeting 




Grant: The Thailand Research Fund (Royal Golden Jubilee Ph.D. 

Program) 

ANTIBACTERIAL ACTIVITY AGAINST 

STAPHYLOCOCCUS EPIDERMIS AND 

INHIBITION OF GLUCOSE-STIMULATED 

OXYGEN CONSUMPTION BY GARCINIA 

MANGOSTANA EXTRACT (NO. 0751) 

Nantapong N 1 , Kamkhunthod M 1 , Tengking S 1 , Gritsanapan 

W 2 , Chudapongse N 1 

1 Institute of Science, School of Biology, Nakhon Ratchasima, 

Thailand; 2 Faculty of Pharmacy, Department of Pharmacognosy, 

Bangkok, Thailand 

Staphylococcus epidermidis, a skin flora, is one of 

the major opportunistic pathogen which has been recognized as 

a leading cause of nosocomial infections. Due to multi-drug 

resistance of S. epidermidis. the search for medicinal plantderived 

antibacterial agents against this pathogen has accelerated 

in recent years. It has been previously reported that mangosteen, 

Garcinia mangostana, exhibits an antimicrobial action against 

certain bacteria, fungi and viruses. Our current study is to 

investigate the antibacterial activity of G. mangostana extract 

against S. epidermidis. G. mangostana used in this study was 

collected from Chumporn province located in the southern region 

of Thailand. The crude extract was prepared by extracting the 

pericarps with ethanol, and the extract was then used to evaluate 

the antibacterial activity against S. epidermidis. The result showed 

that the ethonolic extract of G. mangostana exhibited the 

antimicrobial effect against S. epidermidis with MIC of 50 µg/ 

ml. In addition, we had observed that the extract inhibited the 

glucose-induced increase in oxygen consumption of intact S. 

epidermidis cells. This result suggested that the extract interfered 

with the bacterial glucose metabolism; and this action may play 

a role in the antibacterial activity of G. mangostana extract against 

S. epidermidis. 

Key words: Staphylococcus epidermidis, Garcinia mangostana, 

oxygen consumption 



Research, 29 August-2 September 2010, Berlin, Published in 


CHEMICAL QUALITY EVALUATION OF 

TURMERIC CAPSULES PREPARED IN 

HOSPITALS (NO. 0752) 

Werayut Pothitirat 1 , Ekphol Thanuvattana 1 , Sutatip Ouitin , 

Chanai Noysang 2 and Wandee Gritsanapan 3 


2 Thai Traditional Medicine college, Rajamangala University 

of Technology Thanyaburi, Pathumthani, Thailand.; 

3 Department of Pharmacognosy, 3 Faculty of Pharmacy, 

Mahidol University, Bangkok, Thailand 

Turmeric (Curcuma longa Linn. ) of Zingiberaceae 

family is usually called “Khamin Chan” in Thai language. It is 

well-known for a Thai traditional medicine and herbal cosmetic. 

Carminative and antifungal properties of this plant arise from

302 

volatile oil while antioxidant effect comes from yellow 

curcuminoids. at present, turmeric capsules can be produced and 

dispensed to patients by the pharmacy sector of the hospitals. A 

concerned problem is the quality of turmeric raw material and 

finish capsules have not been controlled for the contents of volatile 

oil and total curcuminoids. Therefore, the aim of this study was 

to determine the amount of volatile oil and total curcuminoids in 

turmeric capsules prepared at 10 different hospitals in Thailand 

using method recommended in Thai Herbal Pharmacopoeia 

(THP). In addition, thin layer chromatography (TLC)-fingerprint 

of volatile oil and turmeric extract from all sample were also 

performed. The average content of volatile oil in all samples was 

found to be 7.59 ± 1.51% v/w, while the average content of total 

curcuminoids was 5.58 ± 0.59% w/w. The limits of volatile oil 

and total curcuminoids in all turmeric capsules were in the range 

of 5 to 10% v/w and 4.60 to 6.66% w/w, respectively. Only one 

of ten samples contained volatile oil and total curcuminoids less 

than the amounts recommended by THP. Ninety percents of all 

samples contained the active ingredients within the limits 

recommended by THP. TLC-fingerprints of each sample showed 

similar pattern of major band of ar –turmerone in the volatile 

oil. These results indicate that most of the turmeric capsules 

prepared in the hospitals in Thailand have a high standard in 

term of chemical quality. These data will be useful as a basic 

information for patient , s confidence in using turmeric capsules 

prepared and dispensed by hospitals in Thailand . 

Key words: curcuminoids, Turmeric, Curcuma longa 

(Presented at the 27 th Annual Research Conference in 

Pharmaceutical Sciences, 2010, Bangkok, Thailand, December 

3, 2010.) 

IN VIVO HEMOSTATIC AND ANTI-INFLAM- 

MATORY ACTIVITIES OF CHROMOLAENA 

ODORATA LEAF EXTRACTS (NO. 0753) 

Hataichanok Pandith 1 , Suchitra Thongpraditchote 2 , Yuvadee 

Wongkrajang 2 , and Wandee Gritsanapan 1* 


Mahidol University; 2 Department of Physiology, Faculty of 

Pharmacy, Mahidol University, Bangkok, Thailand. *E-mail: 


Introduction: Chromolaena odorata (L.) King and 

Robinson or Siam weed is a semi-woody shrub of the family 

Asteraceae. In Thailand, the fresh leaves are used in rural areas 

to stop bleeding. This study was investigated to find the 

appropriate extraction method and solvent which yield the extract 

with the highest hemostatic activity and also investigated its in 

vivo anti-inflammatory activity. Objectives: To find the 

appropriate extraction method and solvent which yield the C. 

odorata leaf extract with the highest hemostatic activity and to 

investigate the in vivo anti-inflammatory activity of C. odorata 

extract. Methods: The lyophilized aqueous fresh mature leaf 

extract and the extracts from maceration of fresh and dried mature 

leaves by various solvents (50, 70, and 95% v/v ethanol) were 

studied. Wistar rats were studied for both bleeding time and antiinflammatory 

effect using ethylphenylpropiolate (EPP)-induced 

ear edema model. Each extract was applied after making the 

incision at a foot pad of each rat. Bleeding time was recorded 


and compared with control. Only 70% ethanolic extract was 

studied for anti-inflammatory activity. The percentage of swelling 

and inhibition will be calculated and compared with the standard 

and control group. Results: The 70% ethanolic extract of dried 

mature leaves of C. odorata showed the highest hemostatic 

activity but this extract and the standard (Indomethacin) could 

slightly inhibit inflammatory effect induced by EPP. Discussion 

and Conclusion: The 70% ethanolic extract is the appropriate 

extract with has the highest hemostatic activity and Wistar rats 

might not be suitable for in vivo investigation of antiinflammatory 

activity in which mice should be used to instead. 

Key words: C. odorata, Siam weed, hemostasis, 

(Presented at Commission on Higher Education Congress III- 

University Staff Development Consortium (CHE-USDC Congress 

III), 9-11 September 2010, Pattaya, Thailand) 

Grant: Office of The Higher Education Commission 

ACUTE AND CHRONIC ORAL TOXICITY OF 

STANDARDIZED WATER EXTRACT FROM 

THE FRUIT OF PHYLLANTHUS EMBLICA 

LINN. (NO. 0754) 

Jaijoy K 1 , Soonthornchareonnon N 2 , Lertprasertsuke N 3 , 

Panthong A 1 *, Sireeratawong S 4 * 

1 Department of Pharmacology, Faculty of Medicine, Chiang 

Mai University, Chiang Mai 50200, Thailand; 2 Department 


Bangkok 10400, Thailand; 3 Department of Pathology, Faculty 

of Medicine, Chiang Mai University, Chiang Mai 50200, 

Thailand; 4 Division of Pharmacology, Department of 

Preclinical Science, Faculty of Medicine, Thammasat 

University, Pathumthani 12120, Thailand. 

Summary: Phyllanthus emblica Linn. (Euphorbiaceae) 

is an herbal plant commonly used in Asian traditional medicine 

systems for treatment of many disorders. In the present study, we 

investigated for the first time acute and chronic toxicity of the 

standardized water extract of P. emblica fruit. The water extract 

of P. emblica was prepared according to the Thai Herbal 

Pharmacopoeia and standardized to 20% gallic acid. For studying 

acute toxicity study, single oral dose of 5000 mg water extract / 

kg body weight was administered to Sprague Dawley rats (five 

females, five males). The results showed no toxicity in terms of 

general behavior change, mortality, or change in gross appearance 

of internal organs (LD50 > 5,000 mg/kg). Chronic toxicity was 

studied by daily oral dose (ten females, ten males) of 300, 600 

and 1,200 mg/kg for 270 days. The results showed slightly 

significant differences in the body and organ weights between 

the control and treatment groups. In addition, the rats were 

analyzed for final body and organ weights, necropsy, and 

hematological, blood chemical and histopathological parameters. 

Hematological analysis and clinical blood chemistry revealed 

slightly changes, but were within the normal limits. No gross or 

histopathology findings were observed in the treatment groups. 

Altogether, the standardized water extract from the fruit of P. 

emblica did not produce acute and chronic toxicity in its 

traditional uses. Industrial relevance: Traditional medicine is still 

stronghold of more than 50% of the world population, especially 

in developing countries. Nowadays, a number of medicinal plants


(such as Phyllanthus emblica Linn.) are commercially available 

on local markets as a tonic, food and/or dietary supplement 

product commonly used for clinical management of several 

conditions. Despite the widespread use of this plant, there is still 

little literature on the scientific evaluation of their toxicity. Results 

of the current study provide pivotal evidences for ascertaining 

the safety of the stardardized water extract of P. emblica fruit 

that could be used as food supplements, dietary supplements, 

etc. in folkloric medicine. 

Key words: Phyllanthus emblica Linn.; Acute toxicity; Chronic 

toxicity 

(International Journal of Applied Research in Natural Products 

Vol. 3 (1), pp. 48-58, March-April 2010) 

ANTITUMOUR EFFECTS OF PHYLLANTHUS 

EMBLICA L.: INDUCTION OF CANCER CELL 

APOPTOSIS AND INHIBITION OF IN VIVO 

TUMOUR PROMOTION AND IN VITRO 

INVASION OF HUMAN CANCER CELLS 

(NO. 0755) 

C. Ngamkitidechakul, 1 * K. Jaijoy, 2 P. Hansakul, 3 N. 

Soonthornchareonnon 4 and S. Sireeratawong 5 

1 Department of Biochemistry, Faculty of Medicine, 

ChiangMai University, Chiang Mai 50200, Thailand; 

2 Department of Pharmacology, Faculty of Medicine, 

ChiangMai University, Chiang Mai 50200, Thailand; 

3 Division of Biochemistry, Department of Preclinical Science, 

Faculty of Medicine, Thammasat University, Pathumthani 

12120, Thailand; 4 Department of Pharmacognosy, Faculty of 

Pharmacy, Mahidol University, Bangkok 10400, Thailand; 

5 Division of Pharmacology, Department of Preclinical Science, 

Faculty of Medicine, Thammasat University, Pathumthani 

12120, Thailand 

Phyllanthus emblica Linn. (PE) is a medicinal fruit 

used in many Asian traditional medicine systems for the treatment 

of various diseases including cancer. The present study tested 

the potential anticancer effects of aqueous extract of PE in four 

ways: (1) against cancer cell lines, (2) in vitro apoptosis, (3) mouse 

skin tumourigenesis and (4) in vitro invasiveness. The PE extract 

at 50–100 ìg/mL signifi cantly inhibited cell growth of six human 

cancer cell lines, A549 (lung), HepG2 (liver), HeLa (cervical), 

MDA-MB-231 (breast), SK-OV3 (ovarian) and SW620 

(colorectal). However, the extract was not toxic against MRC5 

(normal lung fi broblast). Apoptosis in HeLa cells was also 

observed as PE extract caused DNA fragmentation and increased 

activity of caspase-3/7 and caspase-8, but not caspase-9, and upregulation 

of the Fas protein indicating a death receptor-mediated 

mechanism of apoptosis. Treatment of PE extract on mouse skin 

resulted in over 50% reduction of tumour numbers and volumes 

in animals treated with DMBA/TPA. Lastly, 25 and 50 ìg/mL of 

PE extract inhibited invasiveness of MDA-MB-231 cells in the 

in vitro Matrigel invasion assay. These results suggest P. emblica 

exhibits anticancer activity against selected cancer cells, and 

warrants further study as a possible chemopreventive and 

antiinvasive agent. 

Key words: Phyllanthus emblica; Emblica offi cinalis; 

antitumour; apoptosis; tumourigenesis; caspase; invasion. 

(Phytother. Res. 24: 1405–1413 (2010) 

CHARACTERISATION AND IMMUNO- 

STIMULATING ACTIVITY OF POLYSAC- 

CHARIDES FROM THAI MEDICINAL PLANTS 

(NO. 0756) 

J. Burana-Osot 1 *, K. Pattanapanyasat 2 , N. Soonthornchareonnon 

3 , K. Sukapirom 2 and T. Toida 4 


Pharmacy, Silpakorn University, Nakorn-Pathom 73000, 

Thailand; 2 Office for Research and Development, Faculty of 

Medicine, Siriraj Hospital, Mahidol University, Bangkok 

10700, Thailand; 3 Department of Pharmacognosy, Faculty of 

Pharmacy, Mahidol University, Bangkok 10400, Thailand; 

4 Graduate School of Pharmaceutical Sciences, Chiba 

University, Chiba 263-8522, Japan 

Water-soluble polysaccharides were isolated from the 

tubers of Butea superba Roxb. and Pueraria candollei Wall. Ex 

Benth. var. mirifica (Shaw et Suvat.) C. Niyomdham, the leaves 

of Centella asiatica (L.) Urb, Ocimum basilicum L., Psidium 

guajava and Andrographis paniculata (Burn. f.) Nees, the stems 

of Cymbopogon citratus (Stapf ExG), and the fruits of Psidium 

guajava and Scaphium scaphigerum. The immunological impacts 

of the polysaccharides on T-lymphocyte proliferation in vitro was 

investigated by flow cytometric (immunofluorescence) analysis 

using staphylococcal enterotoxin B (SEB) as a positive control. 

It was found that the polysaccharides enhanced T-lymphocyte 

proliferation, ranging from 4.5 to 27.0% at a concentration of 

100 mgmL_1, while the activity of SEB was 13.3%. The 

medicinal plants showing the highest immuno-stimulating activity 

were the tubers of Butea superba Roxb. The water-extracted tubers 

contained 60.0% (w/w) carbohydrates with 6.6% (w/w) uronic 

acid. The major constituent monosaccharides of the tubers were 

28.2 mol% galactose, 10.5mol% arabinose and 36.4 mol% 

glucose. 

Key words: Thai medicinal plant; polysaccharide compositions; 

immunostimulating activity; flow cytometry 

(Natural Product Research Vol. 24, No. 15, 20 September 2010, 

1403–1412) 

EFFECT OF THAI PLANT EXTRACTS ON 

P-GLYCOPROTEIN FUNCTION AND VIABILITY 

IN P ACLITAXEL-RESISTANT HEPG2 CELLS 

(NO. 0757) 

Masashi Kawmi 1 , Ryoko Yumoto 1 , Junya Nagai 1 , Varaporn 

Buraphacheep JunyaprasertJ 2 , Noppamas Soonthorncharareonnon 

2 , Denpong Patanasethanont 3 , Bung-orn 

Sripanidkulchai 3 and Mikihisa TAKANO 1 ,* 

1 Department of Pharmaceutics and Therapeutics, Graduate 

School of Biomedical Sciences, Hiroshima University, 

Hiroshima, Japan; 2 Faculty of Pharmacy, Mahidol University, 

Bangkok, Thailand; 3 Faculty of Pharmaceutical Sciences, 

Khon Kaen University, Khon Kaen, Thailand 

The effects of ethanol extracts from Thai plants on Pglycoprotein 

(P-gp) function and cell viability were examined 

using paclitaxel-resistant HepG2 (PR-HepG2) cells. KP018 from

304 

Ellipeiopsis cherrevensis and AT80 from Ancistrocladus tectorius 

increased both rhodamine 123, a typical P-gp substrate, and 

[ 3 H]paclitaxel uptake in PR-HepG2 cells. However, some extracts 

such as MT80 from Microcos tomentosa increased rhodamine 

123, but not [ 3 H]paclitaxel, uptake, while MM80 from 

Micromelum minutum increased only [3H]paclitaxel uptake. 

Thus, the effects of extracts of Thai plants on rhodamine 123 

uptake were not necessarily the same as those on [ 3 H]paclitaxel 

uptake. Purified compounds such as bergapten did not affect the 

uptake of either substrate. KP018, AT80, and MM80 increased 

[ 3 H]paclitaxel uptake and decreased the cell viability in a 

concentration-dependent manner. Among these extracts, KP018 

showed the most potent cytotoxicity. The cytotoxic potency of 

KP018 on PR-HepG2 cells was similar to that on wild-type HepG2 

cells, and was not potentiated by verapamil. At concentrations 

resulting in no cytotoxicity, AT80 and MM80 potentiated 

paclitaxel-induced cytotoxicity in PR-HepG2 cells. These results 

indicate that K018 may be a useful source to search for a new 

anticancer drug, while AT80 and MM80 may be useful as 

modulators of P-gpmediated multidrug resistance in cancer cells. 

Key words: Thai plant extracts; P-glycoprotein (MDR1/ABCB1); 

paclitaxel; rhodamine 123; multidrug resistance; paclitaxelresistant 

HepG2 cells 

(Drug Metab. Pharmacokinet. 25 (2): 155–162 (2010). 

DETERMINATION OF GALACTURONIC 

ACID FROM POMELO PECTIN IN TERM 

OF GALACTOSE BY HPAEC WITH 

FLUORESCENCE DETECTION (NO. 0758) 

Jankana Burana-osot 1 , Noppamas Soonthornchareonnon 2 , 

Amornrut Chaidedgumjorn 1 , 

Saori Hosoyama 3 , Toshihiko Toida 3 


Pharmacy, Silpakorn University, Nakorn-pathom 73000, 

Thailand; 2 Department of Pharmacognosy, Faculty of 

Pharmacy, Mahidol University, Bangkok 10400, Thailand; 3 

Graduate School of Pharmaceutical Sciences, Chiba 

University, Chiba 263-8522, Japan. 

Galacturonic acid (GalA) in pomelo pectin was 

converted to galactose (Gal) by saponification and reduction 

reaction of carboxy group. The characterization of the product 

was investigated by 1 H NMR spectroscopy. The reduced pectin 

was then hydrolyzed with trifluoroacetic acid (TFA) and analyzed 

by high-performance anion-exchange chromatography (HPAEC) 

with fluorescence detector. The content of GalA was calculated 

from an increased Gal after reaction. The GalA content obtained 

from this method was higher than that obtained from HPAECpulsed 

amperometric detection (PAD), which affords to analyze 

GalA directly after hydrolysis. The established method was 

validated and the results showed good linearity, high precision 

and high sensitivity. This method could be further applied for 

the analysis of GalA content in pomelo pectin. 

Key words: Galacturonic acid, Galactose, Pectin, HPAECfluorescence 

detector 

(Carbohydrate Polymers 81 (2010) 461–465) 


GROWTH AND ISOFLAVONOID ACCUMU- 

LATION OF PUERARIA CANDOLLEI VAR. 

CANDOLLEI AND P. CANDOLLEI VAR. 

MIRIFICA CELL SUSPENSION CULTURES 

(NO. 0759) 

Panitch Boonsnongcheep 1 , Sirintra Korsangruang 1 , 

Noppamas Soonthornchareonnon 2 , Yupyn Chintapakorn 3 , 

Promchit Saralamp 1 , Sompop Prathanturarug 1 

1 Department of Pharmaceutical Botany, Faculty of Pharmacy, 

Mahidol University, 447 Sri-ayuthaya Road, Bangkok 10400, 


Pharmacy, Mahidol University, 447 Sri-ayuthaya Road, 

Bangkok 10400, Thailand; 3 Department of Botany, Faculty 

of Science, Chulalongkorn University, Phayathai Road, 

Bangkok 10330, Thailand. 

We established cell suspension cultures derived from 

leaf, stem, and root calli of Pueraria candollei var. candollei 

and P. candollei var. mirifica using liquid Murashige and Skoog 

(MS) medium supplemented with 0.56 lM 6-benzyladenine (BA) 

and 4.52 lM 2,4-dichlorophenoxyacetic acid (2,4-D). Growth of 

the cell suspension cultures progressed to the stationary phase 

within 15–24 days. Methanolic extracts of cell suspension cultures 

of both varieties of P. candollei were analyzed using a validated 

HPLC protocol. All cell lines derived from leaf, stem, and root 

explants produced four major isoflavonoids: daidzein, daidzin, 

genistein, and genistin; these isoflavonoids were detected only 

in the roots of intact plants. Furthermore, the isoflavonoid contents 

of the cell suspension cultures were higher than those of intact 

plants. Thus, cell suspension culture of both varieties of P. 

candollei may be an effective tool for isoflavonoid production. 

Key words: Analytical method validation, Cell suspension 

culture, HPLC, Isoflavonoids, Pueraria candollei. 

(Plant Cell Tiss Organ Cult (2010) 101:119–126) 

IN VITRO AND IN VIVO ANTIPLASMODIAL 

ACTIVITY AND CYTOTOXICITY OF WATER 

EXTRACTS OF PHYLLANTHUS EMBLICA, 

TERMINALIA CHEBULA, AND TERMINALIA 

BELLERICA (NO. 0760) 

Khosit Pinmai 1 , Wanwarang Hiriote 1 , Noppamas 

Soonthornchareonnon 3 , Krisada Jongsakul 4 Seewaboon 

Sireeratawong 2 , Siripen Tor-Udom 1 

1 Division of Microbiology, Department of Preclinical Science, 

Faculty of Medicine, Thammasat University, Pathumthani, 

Thailand; 2 Division of Pharmacology, Department of 

Preclinical Science, and Research Unit of Pharmacology and 

Toxicology of Herbal Medicine, Research Center, Faculty of 

Medicine, Thammasat University, Pathumthani, Thailand; 


Mahidol University, Bangkok, Thailand; 4 Armed Forces 

Research Institute of Medical Sciences, US Army Medical 

Component, Bangkok, Thailand 

Objective: To evaluate the in vitro and in vivo 

antiplasmodial activity and the cytotoxicity of Phyllanthus


emblica Linn, Terminalia chebula Retz, and Terminalia bellerica 

(Gaertn) Roxb extracts. Material and Method: Standard 

phytochemical screening tests were used to detect metabolites in 

the plant extract. The water extracts of medicinal plants were 

tested for their antiplasmodial activity in vitro by assessing their 

ability to inhibit the uptake of [ 3 H] hypoxanthine into the 

Plasmodium falciparum K1 multidrug-resistant strain. 

Cytotoxicity of all extracts was determined on Vero cell line. The 

in vivo antiplasmodial activity in Plasmodium berghei infected 

mice was evaluated by the standard 4-day suppressive test. 

Results: Phytochemical screening of the water extracts of three 

plants revealed the presence of flavonoids, hydrolysable tannins, 

saponin and terpenes. All plant extracts showed antimalarial 

activity (IC 50 values ranging from 14.33 + 0.25-15.41+ 0.61 μg/ 

ml). The water extract of Terminalia bellerica (Gaertn) Roxb. 

had the highest in vitro antiplasmodial activity followed by 

Phyllanthus emblica Linn. and Terminalia chebula Retz. The 

cytotoxic activity was exhibited by all plant extracts on Vero cells 

with IC 50 values of 157.86 to 238.70 mg/ml. All of the plant 

extracts showed selectivity with the selectivity index (SI) ranged 

from 11 to 17. A standard 4-day suppressive test on P. berghei 

infected mice was used to evaluate the in vivo antiplasmodial 

activity of the extracts at 250 mg/kg/day. The results revealed 

that in vivo antiplasmodial activity with good suppression activity 

ranged from 53.40% to 69.46%. Conclusion: All of the plant 

extracts exhibited interesting in vitro and in vivo antiplasmodial 

activity with good selectivity. 

Key words: Phyllanthus emblica, Terminalia chebula, 

Terminalia bellerica, Antiplasmodial activity, Cytotoxicity 

(J Med Assoc Thai 2010; 93 (Suppl. 7) ) 

PARTIAL DEPOLYMERIZATION OF PECTIN 

BY A PHOTOCHEMICAL REACTION (NO. 0761) 


Saori Hosoyama 3 , Robert J. Linhardt 4 , Toshihiko Toida 3 


Pharmacy, Silpakorn University, Nakorn-pathom 73000, 


Pharmacy, Mahidol University, Bangkok 10400, Thailand; 3 

Graduate School of Pharmaceutical Sciences, Chiba 

University, Chiba 260-8675, Japan; 4 Department of 

Chemistry and Chemical Biology, Center for Biotechnology 

and Interdisciplinary Studies, Rensselaer Polytechnic 

Institute, Troy, NY 12180, USA. 

Complex heterogeneous polysaccharides that comprise 

pectin were partially depolymerized by a photochemical reaction 

using ultraviolet light in the presence of titanium dioxide catalyst. 

In a period of 6 h at pH 7, this UV/TiO2 process decreased the 

average molecular weight of pectin from 400 kDa to 200 kDa. 

The characterization of the partially depolymerized pectin, which 

was fractionated by size-exclusion chromatography, was 

performed by 1H NMR spectroscopy, and the spectra obtained 

showed that the resulting oligosaccharides and polysaccharides 

maintained the intact core structure of pectin. The 

monosaccharide content and depolymerization profile were 

determined by high-performance anion-exchange 

chromatography coupled with pulsed amperometric detection. 

This controlled photochemical depolymerization technique might 

be useful for preparation of pectin oligosaccharides as an 

ingredient in food and pharmaceutical products. 

Key words: Pectin, Photochemical depolymerization, NMR 

spectroscopy, HPAEC–PAD 

(Carbohydrate Research 345 (2010) 1205–1210) 

PHOTOLYTIC DEPOLYMERIZATION : NEW 

APPROACH FOR PREPARATION OF 

OLIGOSACCHARIDES (NO. 0762) 


and Toshihiko Toida 3 

1 Department of Pharmaceutical Chemistry. Faculty of 

Pharmacy. Silpakorn University. Nakorn-pathom 73000. 

Thailand ; 2 Department of Pharmacognosv, Faculty of 

Pharmacy. Mahidol University, Bangkok 10400. Thailand ; 

3 Graduate School of Pharmaceutical Sciences. Chiba 

University, Chiba 260-7865. Japan 

Introduction : Oligosaccharides currently produced for 

commercial markets include, fructo-oligosaccharides, galactooligosaccharides 

and soy oligo-saccharides mainly for the 

prebiotic markets in Japan and Europe. Plant polysaccharides 

can be hydrolyzed in several ways, yielding different sets of 

oligomeric degradation products depending on the hydrolysis 

condition. Generally, oligosaccharides were produced by chemical 

hydrolysis and enzymatic degradation. Acid hydrolysis results in 

random cleavage along the polysaccharide chains and produces 

oligosaccharides fragments with unmodified hexuronic acid. The 

disadvantage of this method is the decomposition of sugars which 

decreasing the yield, formation of other products and causes a 

lot of environmental pollution with the strong chemicals used. 

The enzymatic method requires different types of enzyme 

activities and may cause the contamination by microorganisms. 

Photo-catalysis reaction is the acceleration of a photochemical 

reaction by the presence of a catalyst process. The most widely 

used photo-catalyst is titanium dioxide (T102) because of its 

desirable properties, such as chemical stability, high catalytic 

activity under UV-light, low cost and most importantly its low 

biologically toxicity. In this study, a new technological approach 

“photolytic-depolymerization” was developed in order to take full 

advantage of preparation of oligosaccharides degraded from the 

two occurs naturally polysacchalides; alginates and pectin. Results 

and Discussion : Alginate MW avg dramatically decreased after 3 

h of photolysis in the presence of Ti0 2 and more slowly decreased 

after 6 h of photolysis. The MW avg of the alginate samples obtained 

at 3 h and 6 h of photolysis were 108 and 70 kDa, respectively, 

corresponding to a 45.5% and 64.6% decrease in the MW avg of 

the starting alginate polysaccharide. The MW avg of alginate 

samples obtained at pH 4, 7 and 1 0 for 3 h of the reaction time 

was not different. The 1 H-NMR spectra of degraded alginate 

sample obtained from photochemical reaction at pH 4, 7 and 10 

and that obtained from photochemical reaction after 1, 3 and 6 h 

were observed. Each signal was slightly changed by an altered 

distribution of alginate chain lengths. The signals for degraded 

alginate samples could be observed at the same chemical shifts 

of the intact alginate sample, indicating that the chemical 

structures were indistinguishable. The MW avg of pectin in all

306 

solvent decreased and at pH 10 showed the best efficiency for 

degradation with corresponding to 71 .2 % decrease in the MW avg 

of the starting pectin. Pectin MW avg dramatically decreased after 

6 h of photolysis and more slowly decreased after I 0 h of 

photolysis with different results at different pH. At pH 4 and 7, 

the signals derived from galacturonate could be observed in their 

1 H-NMR spectra indicating that the glycosidic linkages were 

mainly hydrolyzed, but the sugar ring units were maintained to 

form the lower-molecular pectin. Degradation in pH 10 caused 

de-esterification of galacturonate as no signal of -COOCH3 was 

observed. The optimal conditions for photochemical reaction to 

obtain degraded sodium alginate and pectin were established as 

exposure to the UV light in the presence of Ti0 2 for 3 h at pH 7 

and 6h at pH 7, respectively. The change in molecular weight of 

alginates and pectin monitored by HPSEC-RI demonstrated that 

the alginate and pectin had been depolymerized by Ti0 2 -catalyzed 

photochemical reaction affording polysaccharides and 

oligosaccharides of lower molecular weight. The 1 H-NMR spectra 

illustrated that photochemical degradation occurs through the 

random breakage of the glycosidic bonds in the alginate and pectin 

polysaccharide. Conclusion : A photochemical reaction has been 

developed for partially de-polymerization of alginate and pectin. 

The T102 used as a catalyst could be easily removed, resulting in 

the convenient production of purified degraded products. This 

process is simpler, and more environmentally benign than 

conventional methods, such as acid hydrolysis. Moreover, it 

appeared as very efficient to cleave glycosidic linkages keeping 

the basic structure. This new depolymerization method might 

represent a convenient and reliable method for a routine 

production of alginate and pectin oligosaccharides and it could 

be applied for the preparation of oligosaccharide derived from 

the other polysaccharides in the near future. Materials and 

Methods : A sodium alginate sample prepared from Lessonia 

nigrescens having a MW avg of ~198 kDa was kindly provided by 

KIMICA Corporation, JAPAN. Commercial citrus pectin (DM = 

6 %) having a M r of ~ 400 kDa, titanium dioxide (anatase type, 

particle size average, 50 ìm) and monosaccharide standards were 

purchased from Wako Pure Chemical Co. (Osaka, Japan). The 

pectin sample was further purified from low-molecular-weight 

contaminants by dialysis against Milli-Q water in a Spectrapor ® 

dialysis tube (molecular weight cut off = 500 Da) for 3 days in 

the cool room arid then freezed dried prior to use. All other 

reagents were of analytical reagent grade. The photochemical 

reaction experiment device (Sen Lights Corporation, Osaka, 

Japan) consists of a VG1500 reaction tank with 5 inlets, a UV 

light source (low-pressure mercury lamp HL400B-8, 400 Watts), 

a power source (HB400P- 1) and a lamp jacket-quartz glass JW- 

2Q. The apparatus is connected with water circulating system to 

cool the lamp. Depolymerizatlon of alginate and pectin by 

photochemical UV/titanium dioxide process Sodium alginate was 

dissolved in water with 1 mg of titanium dioxide (Ti0 2 ) particles 

and closed loosely by a screw cap. The sample tube (borosilicate 

glass) was then placed in the photochemical reaction tank and 

exposed to UV light. The alginate was degraded at room 

temperature and the light exposure time was range from 1 up to 

12 hr. A mechanical stirrer was used in addition to magnetic stirrer 

to ensure the dissolution of air in the solution. Pectin sample was 

also degraded by the photochemical UV/titanium dioxide process 

as described above. Alginate sample solutions and pectin sample 

solutions with different pH values were prepared to study the 

influence of pH on the degradation. After the reaction, all samples 

were centrifuged at 1,500 x g for 5 mm at 20 ° C and the supernatant 

was filtered through 0.45 ìm membrane filter to iliminate excess 


Ti0 2 , dialyzed and lyophilized. The change in molecular weights 

of degraded alginate and pectin samples were evaluated by high 

performance size exclusion chromatography coupling with 

refractive index detector. The characterization of decomposed 

compounds was elucidated by 1 H-NMR spectroscopy. 

Key words: Photocatalysis; Polysaccharides; Alginates; Pectin. 

(The9 th NRCT-JSPS Joint Seminar Natural Medicine Research 

for the Next Decade: New Challenges and Future Collaboration) 

ANTI-INFLAMMATORY AND ANALGESIC 

ACTIVITIES OF THE WATER EXTRACT 

FROM THE FRUIT OF PHYLLANTHUS 

EMBLICA LINN. (NO. 0763) 

Jaijoy K 1 , Soonthornchareonnon N 2 , Panthong A 1 *, 

Sireeratawong S 3 * 

1 Department of Pharmacology, Faculty of Medicine, Chiang 

Mai University, Chiang Mai 50200, Thailand; 2 Department 


Bangkok 10400, Thailand; 3 Division of Pharmacology, 

Department of Preclinical Science, Faculty of Medicine, 

Thammasat University, Pathumthani 12120, Thailand. 

The fresh or dry fruit of Phyllanthus emblica Linn. is 

used in traditional medicines for the treatment of diarrhea, 

jaundice and inflammatory disorder. In the present study, we 

investigated the anti-inflammatory and analgesic activities of the 

standardized water extract from the fruit of Phyllanthus emblica 

was prepared according to the Thai Herbal Pharmacopoeia (THP). 

P. emblica water extract was evaluated for its anti-inflammatory 

activity in rats using ethyl phenylpropiolate (EPP)-induced and 

arachidonic acid (AA)-induced ear edema, carrageenan-induced 

paw edema as well as cotton pellet-induced granuloma models, 

and its analgesic activity in mice using formalin test. The extract 

at 1 mg/ear exhibited anti-inflammatory effect on EPP-induced 

ear edema, but not on AA-induced ear edema. Oral administration 

of P. emblica at the doses of 150, 300 and 600 mg/kg caused 

dose dependent inhibition of carrageenan-induced rat paw edema. 

P. emblica at 600 mg/kg did reduce neither transudative and 

proliferative phases nor body weight gain and thymus weight in 

cotton pellet induced granuloma formation. The extract at the 

doses of 150, 300 and 600 mg/kg elicited a significant analgesic 

activity in a dose-dependent manner on both the early and late 

phase of formalin test. The anti-inflammatory and analgesic 

mechanism of activity of the standardized water extract of P. 

emblica seems to be similar to NSAIDs rather than to steroidal 

drugs. Inhibitory effect on the synthesis and/or release of 

inflammatory or pain mediators may be the main mechanisms of 

action of P. emblica water extract. Industrial relevance: Medicinal 

plants have long been recognized as an important source of 

therapeutically effective treatment for inflammatory diseases. 

Many patients are turning to herbal medicine as their primary, 

complementary or alternative therapies because of the adverse 

effects of the pharmaceutical drugs. P. emblica fruit has been 

used in traditional management of some painful and inflammatory 

conditions and this has been supported by the present study. This 

work will be useful to the industry to produce standardized herbal 

formulation more effective in the treatment of pain and


inflammation with less toxic and less costly than the current 

synthetic drugs. 

Key words: Phyllanthus emblica Linn.; Analgesic activity; Antiinflammatory 

activity 

(International Journal of Applied Research in Natural Products 

Vol. 3 (2), pp. 28-35, June-July 2010) 

THAI TRADITIONAL MEDICINE AND 

SUCCESSFUL CASE REPORTS OF 

INTEGRATED EAST-WEST APPROACHES 

FOR CHRONIC DISEASES (NO.0764) 

Omboon Vallisuta 1 and Suraphan Sirithamwanich 2 


Mahidol University, Bangkok, Thailand. pyoln@mahidol. 

ac.th; 2 Ban Mor Pan-Thai Clinic, District Muang, Rajburi, 

Thailand. 

The most well known Asian Medicines are Traditional 

Chinese medicine and Ayuraveda developed thousands of years 

ago. Both have their own unique theory on human health and 

how to diagnose and treatment, however they share the same idea 

of holistic health and use natural medicine in combination. 

Another traditional Asian Medicine not as widely known is Thai 

Traditional Medicine (TTM) which has been developed in this 

area now called Thailand (formerly Siam) since Buddha’s time 

i.e. 543 years B.C. or 2553 years ago. Some medicinal plants 

recorded in the Buddhist’s teaching book (Mahavejsandorn 

Chadok) are still used in present time. It is clear that TTM in 

Thailand has its root as long as Buddhism and the ancient TTM 

doctors have developed theory and herbal formula from their 

knowledge and experiences, their intuition combining with Thai 

flora made TTM another unique traditional medicine. There are 

four principles for diagnosis i.e. Dhatu Samuthan (the 4 

elements), Aryu Samuthan (the age), Ritu Samuthan (the season) 

and Kala Samuthan (the time). There are five principles for disease 

development i.e. Mahabhutarupa, the Five Ailments, Tridosha, 

Prated Samuthan and Prana Chakra system. The above theories 

and Kumpi Suppakhun (Materia Medica) made it possible for 

the formulation of multi-herbal formula to cure patients with 

chronic diseases i.e. cancer, gout, rheumatoid, hypertension, 

diabetes etc. In conclusion, the advancement of science in western 

medicine should be appropriately selected for integration with 

traditional medicine. The western medicine and method of 

treatment is prime importance for the acute and sudden symptoms 

which will be life threatening but for chronic diseases eastern 

paradigm of holistic healing is better and safer. 

Key words: Thai Traditional Medicine, principles for diagnosis, 

principles for disease development 

(The Fifth National Symposium on Ethnobotany and The Fourth 

Asia-Pacific Forum on Ethnobotany, Beijing, September 10- 

13, 2010.) 

THE APPLICATION OF PHARMACOGNOSY 

FOR PHARMACISTS (NO. 0765) 

Omboon Vallisuta 


Mahidol University. pyoln@mahidol.ac.th 

The American Society of Pharmacognosy defines 

pharmacognosy as “the study of the physical, chemical, 

biochemical and biological properties of drugs, drug substances 

or potential drugs or drug substances of natural origin as well as 

the search for new drugs from natural sources.” Pharmacognosy 

is interdisciplinary, drawing on a broad spectrum of biological 

and socio-scientific subjects, including botany, ethnobotany, 

medical anthropology, marine biology, microbiology, herbal 

medicine, chemistry, biotechnology, phytochemistry, 

pharmacology, pharmaceutics, clinical pharmacy and pharmacy 

practice. The pharmacists are expected from the public to be able 

to provide information concerning drug whether it is from 

chemical synthesis or from natural source. At present, there is an 

increasing popularity in using drugs from natural sources as well 

as traditional medicine. The 9th congress of Italian Society of 

Pharmacognosy reported the market value of herbal products in 

1998 as follow: Germany $2.5 billion, France $1.6 billion and 

Italy $600 billion. In the US, where the use of herbal products 

has never been as prevalent as in continental Europe, the market 

for all herb sales reached a peak in 1998 of $700 billion. The 

application of Pharmacognosy by pharmacists could be 

categorized into two aspects i.e. for western and eastern/ 

traditional types of medicine. In the western system a pharmacist 

can give practical recommendations to : the farmer to grow and 

collect quality herbs, the manufacturer to produce consistent 

effective herbal product, the customer to select the suitable herbal 

products, the patients to select appropriate herbal medicine and 

to recognize the interaction between herbs and chemical drugs. 

The use of single herb or extract falls into the western medical 

system. These can be accomplished through researchers. However, 

it is clear that western medicine cannot cure chronic diseases 

such as cancer, diabetics, gout, etc since western medicine is 

based on reductionism paradigm relies on mechanistic model of 

medical science which require single identifiable active chemical 

ingredient. This concept works well in the area of antimicrobial 

and operation but cannot be applied to complex etiology of 

those in chronic diseases. This is the area that eastern/traditional 

system of medicine proven to be helpful with many patients who 

seek alternatives. The nature of holistic health paradigm in 

traditional medicine and thousands of year in clinical practice 

had made it more suitable for the treatment of chronic diseases 

which involve many functions of human body. Recently, it has 

been accepted that herbs have synergy and polyvalence activities 

such as antioxidant, anti-inflammatory, antimicrobial, apoptosis, 

antiplatelet, hepatoprotective, immunomodulator, 

antiangiogenesis, etc which can better cope with the multiple 

physiological malfunction of patients suffering from chronic 

diseases. It is the new challenge for pharmacognocist to develop 

appropriate ways in providing the quality control of these herbal 

formulations and to prove their efficiency/effectiveness of these 

products using the limiting available methods of scientific studies. 

Key words: Pharmacognosy, pharmacist, chronic diseasess 

(Invited Lecture in 13 th Jordanian Pharmaceutical Conference, 

Amman, Jordan, 22-24 April 2010.)

308 

ANTI-PROLIFERATIVE AND ANTIOXIDATIVE 

ACTIVITIES OF THAI NONI/YOR (MORINDA 

CITRIFOLIA LINN.) LEAF EXTRACT (NO. 0766) 

Wasina Thani 1 , Omboon Vallisuta 1 , Pongpan Siripong 2 and 

Nongluck Ruangwises 3 . 

1 Department of Pharmacognosy , Faculty of Pharmacy, 

Mahidol University; 2 National Cancer Institute, Medical 

Department, Ministry of Public Health, Bangkok, Thailand; 


Pharmacy, Mahidol University 

In Thai Traditional medicine, the leaves of Thai noni/ 

Yor, (Morinda citrifolia Linn.) have been used for the treatment 

of several symptoms including cough, vomiting and pain. 

However, scientific evidence of the benefits of noni leaf is limited. 

In this study, Thai noni leaves were extracted by several methods 

and were evaluated against human cancer cell lines: KB (human 

epidermoid carcinoma), HeLa (human cervical carcinoma), MCF- 

7 (human breast carcinoma) and HepG 2 (human hepatocellular 

carcinoma) cell lines as well as vero (African green monkey 

kidney cell) cell line employing the MTT colorimetric method, 

comparing to damnacanthal, rutin, and scopoletin. The 

dichloromethane extract of fresh leaf showed better inhibitory 

effect against KB and HeLa cells with IC 50 values of 21.67 and 

68.50 μg/ml, respectively. In a similar way, the dichloromethane 

extract of dried leaves revealed cytotoxicity against KB cell with 

IC 50 values of 39.00 μg/ml, whereas lyophilized, decoction, 

ethanolic and methanolic extracts as well as the single 

compounds; rutin and scopoletin, showed reduced antiproliferative 

effect in all cancer cell lines (IC 50 103 to over 600 

μg/ml). Interestingly, damnacanthal showed potent cytotoxicity 

against either all cancer cell lines or vero cell lines. The results 

suggested that Thai noni extracts were safer than the pure 

compounds due to their higher safety ratios, which is a good 

indication for cancer treatment. The leaves have beneficiaries as 

a functional food for chemoprevention against epidermoid- and 

cervical cancers. 

Key words: Morinda citrifolia, anti-proliferative, anti-oxidative 

(Southeast Asian J Trop Med Public Health, 2010: 41(2); 482- 

489.) 

YES! TO ORGANIC MEDICINE (NO. 0767) 

Omboon Vallisuta 


Mahidol University, Bangkok Thailand. pyoln@mahidol.ac.th 

The failure of conventional medicine based on the 

obsolete mechanistic model has prompted the search for better 

sciences for life, and the revival of an ancient paradigm of holistic 

theory of health and organic medicine.Fritjof Capra had foreseen 

the problems created by reductionism 30 years ago in his famous 

book, The Turning Point, where he warned of the wrong 

mechanistic Cartesian paradigm being used for medicine, 

although physicists themselves have gone beyond it. 

Unfortunately, this is still true to-day. Capra pointed out that all 

scientific theories are approximations to the true nature of reality; 

and that each theory is valid for a certain range of phenomena. 

New theories would have to be found to replace the old one, or, 

rather, to extend it by improving the approximation. This article 

provides evidences for better approach in treatment as practised 

by a Thai traditional doctor. The Eastern paradigm of human 

physiology is different from the western medicine and involves 

energy and network of energy lines which govern the functions 

of human body. Various herbs have been used since ancient 

time and have been scientifically proved as antioxidant, 

antimicrobial, antiproliferative, immunomodulator, antiinflammatory, 

enzyme inhibitor, antimutagenic etc and most of 

all, herbs give nourishment to normal cells. The traditional 

doctor’s view about the pathogenesis of cancer is that there is 

weakness in the body’s energy system (the Tridosha) i.e. Pitta or 

homeostasis, Vata or metabolism and Kapha or anabolism which 

allows the accumulation of toxic substances or free radicals. As 

a result a cyst or tumour develops and later turns to cancer. 

Example of these herbs are Allium sativum, Aloe vera, Asparagus 

racemosa, Curcuma longa, Melia azedarach, Nigella sativa, 

Ocimum sanctum, Paederia foetida, Phyllanthus emblica, 

Plumbago zeylanica, Picrorhiza kurroa, Tinospora cordifolia, 

Terminalia chebula etc. It is essential that doctors from the two 

systems must intercommunicate with respect and with the same 

aim i.e. to heal the patients, not to get rid of the disease. Another 

urgent matter is that many medicinal plants in the world are being 

lost by the lack of knowledge in their usefulness, and therefore 

no conservation is attempted. 

Key words: Fritjof Capra, Thai Traditional Medicine, cancer 

ANTIOXIDANT ACTIVITY AND PHENOLIC 

CONTENT OF ACANTHOPANAX TRIFOLIATUS 

AND TODDALIA ASIATICA (NO. 0768) 

Pongtip Sithisarn 1 , Siripen Jarikasem 2 



Mahidol University, 447 Sri-Ayudhaya Rd., Rajathevi, 

Bangkok 10400 Thailand; pypst@mahidol.ac.th ; 

2 Pharmaceutical and Natural Products Department, Thailand 

Institute of Scientific and Technological Research, 35 M.3, 

Lieab Klong 5 Rd., Klong 5,Klong Luang, Pathum 

Thani,12120, Thailand 

Decoction and 75% ethanolic extracts from the leaves 

of Acanthopanax trifoliatus (AT) and Toddalia asiatica (TA) were 

spectrophotometrically examined for antioxidant activity and 

phenolic contents. Phytochemical characteristics of the decoction 

extracts were also investigated by thin layer chromatography 

(TLC). Leaf decoction extract of AT significantly exhibited the 

highest in vitro antioxidant activity determined by 1,1-diphenyl- 

2-picrylhydrazyl (DPPH) scavenging assay and thiobarbituric acid 

reactive substances (TBARS) method. TA decoction and 75% 

ethanolic extracts significantly contained the highest amount of 

total phenolic and total flavonoid contents, respectively. Analysis 

of decoction extracts of AT and TA by TLC showed specific 

fingerprints composed of chlorogenic acid and rutin. 

Key words : Acanthopanax trifoliatus, Toddalia asiatica, 

antioxidant


ADAPTOGENIC-RELATED ACTIVITY AND 

PHENOLIC CONTENT OF SELECTED GINSENG- 

LINE HERBS IN THAILAND (NO. 0769) 

Pongtip Sithisarn 1 , Siripen Jarikasem 2 , Krittiya 

Thisayakorn 2 , Winai Supatanakul 3 , 

Vullapa Arunpairojana 2 


Mahidol University, 447 Sri-Ayudhaya Rd., Rajathevi, 

Bangkok 10400 Thailand; pypst@mahidol.ac.th ; 2 

Pharmaceutical and Natural Products Department, Thailand 



Thani,12120, Thailand; 3 Agricultural Technology 

Department, Thailand Institute of Scientific and 

Technological Research, 35 M.3, Lieab Klong 5 Rd., Klong 

5,Klong Luang, Pathum Thani,12120, Thailand 

Eleven plants traditionally as adaptogens were 

collected from the north and northeastern parts of Thailand and 

evaluated for adaptogenic-related properties including antioxidant 

and anti-anxiety activities as well as total phenolic and total 

flavonoid contents. Plant samples were extracted by various 

methods then were tested for in vitro antioxidant activity and 

were investigated for phenolic and flavonoid contents by 

spectrophotometric techniques. Thirty tested extracts showed 

antioxidant activity with EC50 ranged from 14.50 ± 1.04 to 

783.68 ± 19.94 and 11.18 ± 2.60 to 745.24 ± 24.54 ?g/mL using 

DPPH scavenging assay and thiobarbituric acid reactive 

substances (TBARS) method, respectively. Their total phenolic 

and total flavonoid contents are in the range of 1.93 ± 0.04 to 

31.74 ± 1.08 g% chlorogenic acid equivalent (g% CAE) and 0.38 

± 0.01 to 12.39 ± 1.40 g% rutin equivalent (g% RE), respectively. 

It was found that the leaf decoction of Acanthopanax trifoliatus 

(ATD) exhibited strong antioxidant activity with high amount of 

phenolic and flavonoid contents. ATD was further tested for in 

vivo anti-anxiety activity using light-dark task and hole-board 

test. Animals receiving ATD at the concentrations of 500 to 1000 

mg/kg significantly (P < 0.05) increased the number of entries 

(80%) and time spent (90%) in light chamber in light-dark task. 

For the hole-board test, animal group receiving 1000 mg/kg ATD 

significantly increased the number of head-dip (37%). The results 

indicated that plant samples, especially the leaves of 

Acanthopanax trifoliatus possess antioxidant and anti-anxiety 

activities partly supports their ethnomedical uses as adaptogenic 

agents. 

Key words : adaptogen, antioxidant, anti-anxiety 

ANTIOXIDANT AND BRINE SHRIMP 

LETHALITY ACTIVITIES OF CALLICARPA 

CANDICANS (NO. 0770) 

Sarinthip Muensaen 1,3 , Siripen Jarikasem 3 , Pongtip 

Sithisarn 2 , Nongluck Ruangwiset 1 


Pharmacy, Mahidol University, Bangkok, Thailand 10400; 


Mahidol University, Bangkok, Thailand 10400; 

pypst@mahidol.ac.th; 3 Pharmaceutical and Natural Products 

Department, Thailand Institute of Scientific and 

Technological Research, Pathum Thani, Thailand, 12120 

Leaf extracts from Callicarpa candicans were tested 

for 2 biological activities; free radical scavenging activity and 

cytotoxic effects. From DPPH scavenging assay, 95% ethanolic 

and aqueous extracts showed strong inhibitory effect to the 

radicals with EC 50 values of 29.48 ± 0.41 and 22.26 ± 1.17 μg/ 

ml, respectively while the essential oil from the leaves showed 

very low effect (EC 50 values of 3197.14 ± 358.10 μg/ml). The 

result from in vitro cytotoxicity test showed that essential oil 

fraction exhibited LC 50 value lessen than 250 µg/ml which 

considered as significantly active and had the potential for further 

investigation while the ethanolic and aqueous extracts showed 

less toxicity effect. The results suggested that 95% ethanolic and 

aqueous extracts should be considered as potential sources of 

some active phytochemicals for good antioxidant activity with 

low toxic effect. Moreover, the essential oil showed significant 

cytotoxic activity which could lead to the discovery of new 

cytotoxic compounds for further development of pesticide or 

piscicidal agents. 

Key words : Callicarpa candicans, DPPH, brine shrimp lethality 

ANTI-CHOLINESTERASE ACTIVITY OF 

THAI MEDICINAL PLANTS CONTAINING 

ALKALOID (NO. 0771) 

Janthima Khaiman 1 , Chalinee Thongtip 1 , Piyanuch 

Rojsanga 1 , Pongtip Sithisarn 2 


Pharmacy, Mahidol University, Bangkok, Thailand 10400 ; 


Mahidol University, Bangkok, Thailand 10400; 

pypst@mahidol.ac.th 

Acetylcholinesterase inhibitor (AChEI) has been used 

as a drug for the symptomatic treatment of Alzheimer’s disease. 

A variety of plants has been reported to show AChE inhibitory 

activity and their alkaloids constituent have shown to be the 

bioactive compounds. In this study, the inhibitory effect of 10 

Thai medicinal plants containing alkaloids on the AChE activity 

was investigated. The plants methanolic extracts and standard 

alkaloids were tested for AChE inhibitory activity using Ellman’s 

TLC and micro-plate assays. The methanolic extracts from stems 

of Coscinium fenestratum and leaves of Camellia sinensis at 

concentration of 0.1 mg/ml inhibited more than 80% of AChE 

activity. At the 1 mg/ml, the roots of Rauvolfia serpentine and 

stems of Tinospora crispa showed 69–70% inhibitory activity. 

The results from TLC analysis corresponded to the results from 

the mcro-plate assay. In addition, the majority of active 

components in the plant extracts were alkaloids. These results 

suggest that the effective extracts should be further investigated 

to obtain new molecules for the treatment of neurodegenerative 

disorders. 

Key words : anti-cholinesterase, alkaloid, Ellman’s method

310 

CHROMATOGRAPHIC AND PHYSICAL 

CHARACTERISTIC OF ALOE VERA LEAF 

EXTRACTS AND COMMERCIAL PRODUCTS 

(NO. 0772) 

Pongtip Sithisarn 1 , Ruxjinda Wattanalai 2 


Mahidol University, Bangkok, Thailand 10400 ; 2 Faculty of 

Pharmacy, Siam University, Bangkok, Thailand 10160 

Aloe (Aloe vera (L.) Burm.f.) is a herb in 

Asphodelaceae Family. Gel from the leaves of aloe mainly 

composed of polysaccharides including pectin, cellulose, 

glucomannan and acemannan (aloeverose). This gel was reported 

to promote various biological activities such as wound healing, 

anti-inflammatory, anti-sunburn, antiulcer and immunomodulator. 

This experiment was set up in order to compare the fingerprint 

characteristics of the leaf gel extracts from Aloe vera collected 

from different locations and extraction methods using 

chromatographic and spectroscopic methods. From TLC analysis 

using Anisaldehyde-H 2 SO 4 reagent, all Aloe vera extracts showed 

dark green bands at Rf 0.40 corresponded to acemannan. 

Extraction by homogenization and centrifugation promoted more 

purified samples than fresh squeezing method. Of all Aloe vera 

cosmetic products, only gel sample which had labeled amount of 

87.399% aloe gel showed TLC band of acemannan. Aloe vera 

samples from Bangkok which the leaves were plain and had large 

size (> 7 cm. wide) showed high intensity of acemannan bands 

on TLC suggested the good source of Aloe vera gel. IR spectra of 

Aloe vera extracts and gel product showed identical pattern to 

the reported data. RI values of all aloe extracts, products and 

acemannan are in the range of 1.3340 to 1.3544 which 

corresponded to the average RI value of most commercial 

products. From the results, analysis by TLC, IR and RI of Aloe 

vera extracts and products showed specific chromatographic and 

physical characteristics. This analysis can be developed for 

standardization of Aloe vera gel raw materials and finished 

products using acemannan as a marker. 

Key words : Aloe vera, TLC fingerprint, acemannan 

DEVELOPMENT OF CENTELLA TEA BAG 

(NO. 0773) 

Kittiya Theangjit 1 , Rungthiwa Dasananda 1 , Pongtip 

Sithisarn 1* , Wandee Gritsanapan 1 , Piyanuch Rojsanga 2 

1* Department of Pharmacognosy, Faculty of Pharmacy, 

Mahidol University, Bangkok, 10400, Thailand; 

pypst@mahidol.ac.th ; 2 Department of Pharmaceutical 


Bangkok, 10400, Thailand 

Centella (Centella asiatica (L.) Urban) is an 

indigenous herb in Apiaceae family which has been popularly 

consumed as vegetable and drink in Thailand Development of 

tea bag from centella leaves was conducted in order to find the 

suitable conditions for tea bag production including drying, 

powdering and packaging which promoted centella tea with good 

physical characteristics and high amount of asiaticoside, an active 


compound, analyzed by TLC-densitometric method. The result 

showed that centella teas obtained from the raw materials from 

various drying methods showed no different physical 

characteristic. The tea from centella leaves from 16 hours drying 

in hot air oven contained the highest amount of asiaticoside. 

Centella tea from the coarse powder significantly gave higher 

amount of asiaticoside than the tea from fine powder (0.846 and 

0.640 mg per cup, respectively). Moreover, the coarse powder 

also showed better physical stabilities than the fine one. 

Aluminum foil was found to be the most suitable packaging which 

promoted the best physical stabilities over the plastic or paper 

materials. 

Key words : Centella asiatica, tea bag, TLC-densitometry 

DEVELOPMENT OF INSTANT DRINK FROM 

CENTELLA LEAVES (NO. 0774) 

Nunticha Wongvichitsilp 1 , Nutprawee Whangsuseung 1 , 

Pongtip Sithisarn 1* , Wandee Gritsanapan 1 , Piyanuch 

Rojsanga 2 

1* Department of Pharmacognosy, Faculty of Pharmacy, 

Mahidol University, Bangkok, 10400, Thailand; 

pypst@mahidol.ac.th ; 2 Department of Pharmaceutical 


Bangkok, 10400, Thailand 

Centella (Centella asiatica (L.) Urban) is a Thai herb 

that has been popularly consumed as food and drink for a long 

time. An active substance in centella which promotes good 

beneficial to human body is asiaticoside. This research was set 

up in order to study the suitable extraction method which 

promoted centella extract with high extractive yield and high 

amount of asiaticoside analyzed by TLC-densitometric method. 

The development of instant drinks from centella leaves with good 

physical characteristics by various methods was also conducted. 

The results revealed that decoction of dried centella leaf powder 

with spray drying method promoted the high extractive yield and 

asiaticoside content. It was found that the suitable amounts of 

diluent, which was lactose, were 15 and 20% w/v. Moreover, 

combination of the fresh squeezing extract which was dark green 

and extract from decoction of dried powder at the ratio of 1:1 

promote the instant drink with good physical appearance. Four 

selected formulas of centella instant drink were evaluated for 

sensory satisfaction including color, odor, flavor and appearance 

in 30 samples using 5–Point Hedonic Scale method. It was found 

that the formula of 20% lactose with no combination of fresh 

squeezing extract gave the most satisfactory centella instant drink. 

Key words : Centella asiatica, instant drink, TLC-densitometry 

PHYTOCHEMICAL STUDY ON CALLICARPA 

CANDICANS LEAVES (NO. 0775) 

Siripen Jarikasem 1 , Sarinthip Meunsaen 1 , Pongtip Sithisarn 2 

1 Pharmaceutical and Natural Products Department, Thailand 



Thani,12120, Thailand; 2 Department of Pharmacognosy,


Faculty of Pharmacy, Mahidol University, Bangkok, 10400, 

Thailand; pypst@mahidol.ac.th 

Carllicarpa candicans (Burm. F.) Hochr. is a shrub 

belongs to Lamiaceae family. This plant has been traditionally 

used as fish poison due to its strong piscicidal component named 

callicarpone. Moreover, C. candicans possesses numerous 

medicinal properties including anti-inflammatory, emmenagogue, 

antiasthma and abdominal pain relieving. Very few biological 

activities and phytochemicals have been reported from this plant 

species. Our recent study on free radical scavenging activity of 

leaf ethanolic and aqueous extracts of C. candicans showed strong 

DPPH scavenging effect with ED 50 values of 29.48 ± 0.41 μg/ml 

and 22.26 ± 1.17 μg/ml, respectively. In this study, the analysis 

of phenolic and flavonoid components in the leaf extracts by TLC 

and HPLC was carried out. TLC chromatogram with specific 

spray reagents including ferric chloride and NP/PEG showed the 

presence of phenolic acids and flavonoids. Reversed phase C-18 

HPLC analysis of the extracts with PDA detector showed specific 

fingerprints. By comparison of retention times and absorbance 

spectra with reference substances, the peaks corresponding to 

chlorogenic acid and flavonoid components were observed. The 

presence of phenolic and flavonoid components in C. candicans 

leaves obtained from this study confirmed free radical scavenging 

activity as well as medicinal properties of this plant species. 

Further studies on the isolation and structural characterization 

of bioactive compounds using spectroscopic techniques including 

1 H- and 13 C-NMR, IR and MS are ongoing. 

Key words : Callicarpa candicans, TLC, HPLC 

STANDARDIZATION OF ZANTHOXYLUM 

LIMONELLA (DENNST.) ALSTON EXTRACT 

(NO. 0776) 

Nadkanjana Srirattananont, Weena Jiratchariyakul 

Faculty of Pharmacy, Mahidol University, Bangkok 10400, 

Thaialnd. E-mail : pywju@mahidol.ac.th 

Standardization covers all measures to attain the 

quality and reproducibility of herbal products (1). The 

standardized plant extracts which are used as drug material in 

herbal products can be proved by qualitative and quantitative 

analyses, which require a marker compound. Marker compound 

is defined as the compound isolated from the used plant and 

applied for assessing the quality of the drug material (2). Here 

we report about Zanthoxylum limonella pericarp ethanolic extract, 

which will be applied as drug material in oral products. Z. 

limonella (Dennst.) Alston (Rutaceae) (3) has been traditionally 

used as remedies for stomachache, toothache, intestinal worms, 

rheumatism, scabies, snakebites, fever, cholera (4). It possessed 

weak antifungal and weak anti-inflammatory properties (5). The 

phytochemical work and the quantitative HPLC analysis of the 

pericarp extract has been performed. Compounds 1,2 and 3 were 

isolated from the pericarp and identified using modern NMR 

spectroscopy. Compound 1 was identified as β-sitosterol, 2 as a 

mixture of β-sitosteryl glucoside and stigmasteryl glucoside and 

3 as lupeol. Compound 3 was selected as a marker compound 

because it could be isolated sufficiently without difficulty. The 

analytical method was validated and conformed to the USP 26 

requirement. From our study, the content of 3 in the pericarp 

extract was 0.195% w/w. 

Key words : standardization, Zanthoxylum limonella, Marker 

compounds 

(The 9 th NRCT-JSPS Joint Seminar on Natural Medicine in 

Pharmaceutical Scieces, “Natural Medicine Research for the 

Next Decade : New Challenges and Future Collaboration” 

December 8-9, 2010. Faculty of Pharmaceutical Sciences, 

Chulalongkorn University.)

Faculty of Pharmacy - Mahidol University

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