Mahidol University Annual Research Abstracts, Vol. 31

Mahidol University Annual Research Abstracts, Vol. 31 207 

EFFECT OF DIFFERENT CONCENTRATION OF 

COCONUT OIL, OLIVE OIL, SESAME OIL AND 

YEAST EXTRACT ON GROWTH OF PITYROS- 

PORUM SPECIES (NO. 548) 

Mansuang Wuthi-udomlert 1 and Omboon Luanratana 2 

1 Department of Microbiology, 2 Department of Pharmacognosy, 

Faculty of Pharmacy, Mahidol University. 

Key words : Pityrosporum spp., olive oil, coconut oil, sesame oil 

Direct examination is used to demonstrate the causative 

agent of pityriasis (tinea) versicolor and dandruff associated organism, 

Pityrosporum orbiculare (P. ovale or Malassezia furfur). In 

some particular cicumstance, this yeast requires a culture in which 

some kind of oil or supplements are substantially needed. In this 

study, local coconut oil (C), local sesame oil (S) and commercial 

olive oil (O) at 0.25% and 0.5% as well as yeast extract (Y) 2% and 

4% were incorporated into Sabouraud dextrose medium (SDA) to 

culture pure clinical isolates of Pityrosporum spp. Colonial size and 

characteristic were recorded macroscopically. After full growth of 

organisms were obtained, the length and width of yeast from Gram 

stain were measured and average, therefore, the cellular area were 

obtained. Growth enhancing ability of each medium with different 

concentration of oil were compared. The data concluded that using 

0.25% of each oil with 2% yeast extract, average width and average 

length of yeast cells obtained were statistically similar (p > 0.01). 

According to area data, local coconut oil can apparently be a substitution 

for conventional olive oil for culturing purpose of Pityrosporum 

spp. 

(Proceeding of The 41 th Kasetsart University Annual Conference 2003 

: 216-220.) 

INHIBITORY EFFECT OF LYOPHILIZED 

AQUEOUS EXTRACT OF THAI TRADITIONAL 

HERBS AGAINST PROPIONIBACTERIUM ACNES 

(NO. 549) 

Mansuang Wuthi-udomlert 1 , Sompop Prathanturarug 2 , and 

Juree Jearanaisilavong 3 

1 Department of Microbiology, 2 Department of Pharmaceutical 

Botany, Faculty of Pharmacy, 3 Department of Microbiology, Faculty 

of Medicine, Siriraj Hospital, Mahidol University. 

Key words : Propionibacterium acnes, anti-acnes, lyophilization 

Six Thai commercial herbs from local distributor were 

investigated for their efficiencies on bacteria associated in acne formation. 

Form volunteers, with various degree of acne, 42 strains of 

Propionibacterium acnes were isolated anaerobically, using modified 

blood agar medium. Lyopholization of aqueous extracts were 

obtained from Acacia concinna Wild. DC., Cardiospermum 

helicacabum L., Clerodendrum indicum L., Curcuma xanthorriza 

Roxb., Plumbagp zeylanica L. and Terminalia citrina Roxb., with 

20.36, 3.98, 1.86, 7.20, 1.95 and 27.58% yield (w/w), respectively. 

Herbal prohibition effects demonstrated as inhibitory zone diameters 

(IZD) by the use of agar diffusion method. Comparison with referenced 

drug, 10 mg per disc ampicillin, T. cirtrina exhibited statisti- 

cally similar effect against tested organisms (p > 0.01) as well as the 

effect of C. helicacabum but for different number of susceptible 

strains (42 and 4 strains, respectively). 

Active ingredient responsible to these inhibitions, local 

dermal irritation and possibility to formulate into external traditional 

drug are interesting for further study. 

(Proceeding of The 3 rd World Congress on Medicinal and Aromatic 

Plants for Human Welfare. Chiang Mai, Thailand, 2003 : 413.) 

ETHNOMEDICAL USES OF THAI ANNON- 

ACEOUS PLANTS (1) (NO. 550) 

Wongsatit Chuakul 1 , Noppamas Soonthornchareonnon 2 

1 Department of Pharmaceutical Botany, 2 Department of Pharmaceutical 

Pharmacognosy, Faculty of Pharmacy, Mahidol University. 

Key words : Ethnomedical, Thai Annonaceous Plants, Thailand. 

A survey on the utilization of medicinal plants at seventeen 

provinces; i.e. Nan. Sukhothai, Ubon Ratchathani, Si Sa Ket, 

Chaiyaphum, Yasothon, Nakhon Ratchasima, Surin, Saraburi, 

Kanchanaburi, Chanthaburi, Krabi, Surat Thani, Songkhla, 

Phattalung, Narathiwat and Yala were carried out by interviewing 

herbalists followed by collecting plant specimens and identifying 

the specimens. In addition, the plant specimens were compared with 

the authentic specimens at two herbaria: the Bangkok Herbarium 

(BK), Botany Section, Botany and Weed Science Division, Department 

of Agriculture, Ministry of Agriculture and Cooperatives and 

the Royal Forest Herbarium (BKF), National Park, Wildlife and Plant 

Conservation Department, Ministry of National Resources and Environment. 

Sixty-two medicinal plants of Thai annonnaceous plants 

were recordet from ten provinces in Thailand. Twenty-five genera 

of these ANNONACEAE; Alphonsea, Anaxagorea, Annona, 

Artabotris, Cananga, Cyathostemma, Dasymaschalon, Desmos, 

Ellipeia, Miliusa, Mitreflora, Monocarpia, Nervopetalum, Orophea, 

Polyalthia, Pseuduvaria, Rauwenhoffia, Stelechocarpus, Uvaria and 

Xylopia and all of these plants were ethomedicinal used. 

(Thai J Phytopharm 2003; 10(1): 25-32.) 

ETHNOMEDICAL USES OF THAI ZINGIBERA- 

CEOUS PLANTS (1) (NO. 551) 

Wongsatit Chuakul, Ampol Boonpleng 

Department of Pharmaceutical Botany, Faculty of Pharmacy, 

Mahidol University 

Key words : Ethnomedical, Thai Zingiberaceous Plants, Thailand 

A survey on the utilization of medicinal plants at twentytwo 

provinces; i.e. Nan, Chiang Mai, Lampang, Sukhothai, 

Phetchabun, Chaiyaphum, Maha Sarakham, Ubon Ratchathani, Si 

Sa Ket, Yasothon, Surin, Buri Ram, Saraburi, Chanthaburi, 

Kanchanaburi, Phetchaburi, Surat Thani, Nakhon Si Thammarat, 

Phattalung, Trang, Krabi and Pattani were carried out by interviewing 

herbalists followed by collecting plant specimens and identify-

208 

ing the specimens. In addition, the plant specimens were compared 

with the authentic specimens at two herbaria: the Bangkok Herbarium 

(BK), Botany Section, Botany and Weed Science Division, Department 

of Agriculture, Minisry of Agriculture and Cooperatives and 

the Royal Forest Herbarium (BKF), National Park, Wildlife and Plant 

Conservation Department, Ministry of National Resources and Environment. 

Fifty-eight species of medicinal plants belonging to 

ZINGIBERACEAE family were recorded as medicinal plants. Twelth 

genera of these ZINGIBERACEAE; each each of 11 Curcuma and 

Zingiber, each of 8 Alpinia and Kaempferia, 6 Etlingera, 4 

Boesenbergia, each of 3 Amomum and Globba, 2 Elletariopsis and 

each of 1 Gagnepainia and Hedychium; and all of these plants were 

ethnomedicinal used. 

(Thai J Phytopharm 2003; 10(1): 33-9.) 

ETHNOMEDICAL USES OF THAI ORCHIDACEOUS 

PLANTS (NO. 552) 

Wongsatit Chuakul 

Department of Pharmaceutical Botany, Faculty of Pharmacy, 

Mahidol University. 

Key words : Ethnomedical, Thai Orchidaceous Plants, Thailand 

A survey on the utilization of medicinal plants at 

Sukhothai, Phitsanulok, Maha Sarakham, Ubon Ratchathani, 

Chaiyaphum, Yasothon, Surin, Kanchanaburi, Krabi, and Yala, were 

carried out by interviewing herbalists followed by collecting plant 

specimens and identifying the specimens. In addition, the plant specimens 

were compared with the authentic specimens at two herbaria: 

the Bangkok Herbarium, Botany Section, Botany and Weed Science 

Division, Department of Agriculture, Ministry of Agriculture and 

Cooperatives and the Forest Herbarium, Royal Forest Department, 

Ministry of Agriculture and Cooperatives. Forty-six medicinal plants 

of Thai Orchidaceous plants were recordet from ten provinces in 

Thailand. Twenty-six genera of these ORCHIDACEAE; Acampe, 

Apostasi, Bulbophyllum, Calanthe, Cirrhopetalum, Cleisostoma, 

Coelogyne, Cymbidium, Dendrobium, Doritis, Eria, Eulophia, 

Gastrochilus, Geodorum, Grammatophyllum, Habenaria, Luisia, 

Neuwiedia, Nervillia, Pecteilis, Robiquetia, Spathoglottis, Sunipia, 

Thrixspermum, Trias, and Vanilla and all of these plants were 

ethnomedicinal used. 

(Mahidol University Journal of Pharmaceutical Science 2002; 29 

(3-4), 41-5.) 

HIGH-FREQUENCY SHOOT MULTIPLICATION 

IN CURCUMA LONGA L. USING THIDIAZURON 

(NO. 553) 

Sompop Prathanturarug 1 , Noppamas Soonthornchareonnon 2 , 

Wongsatit Chuakul 1 , Yuvaluk Phaidee 1 , Promjit Saralamp 1 

1 Department of Pharmaceutical Botany and 2 Department of Pharmacognosy, 

Faculty of Pharmacy, Mahidol University, E-mial 

:pyspr@mahidol.ac.th 

Key words : Curcuma longa, In vitro propagation, Thidiazuron, Turmeric 

Faculty of Pharmacy 

The effects of plant growth regulators, explant tpes, and 

culture regimens were investigated on in vitro shoot proliferation 

from terminal bud explants of Curcuma longa. Each bud was longitudinally 

divided into four equal pieces,each 1 cm in length, and 

used as explants. These were then cultured on MS medium supplemented 

with 18.17 mM tidiazuron for 4 weeks prior to transfer to 

MS medium without growth regulator for 8 weeks. Under these 

conditions, a shoot induction rate of 18.22+0.62 shoots/explant was 

obtained after 12 weeks of cultures. Spontaneous rooting was 

achieved. The regenerated plants were transferred to soil under 

greenhoust conditions and subsequently grown successfully in the 

field. 

(Acknowledgements: This work was supported by the National Research 

Council of Thailand. Plant Cell Reports 21 (2003): 1054- 

1059.) 

IDENTIFICATION OF PUERARIA CANDOLLEI 

GRAHAM EXBENTH. VARIETIES USING RAPD 

ANALYSIS (NO. 554) 

Sompop Prathanturarug 1 , Noppamas Soonthornchareonnon 2 , 

Wongsatit Chuakul 1 , Promjit Saralamp 1 , Takayuki Tohge 3 , 

Hiroshi Sudo 3 , Mami Yamazaki 3 , Tsutomu Ishikawa 4 , Kazuki 

Saito 3 


Faculty of Pharmacy, Mahidol University, Thailand, 

3 Department of Molecular Biology and Biotechnology and 4 Department 

of Medicinal Organic Chemistry, Graduate school of 

Pharmaceutical Sciences, Chiba University, Japan, E-mail : 

pyspr@mahidol.ac.th 

Key words : Pueraria candollei, RAPD analysis, Metabolite profiling 

Kwao Keur is used in Thai traditional medicine for rejuvenation. 

Its tuberous root contains phytoestrogen, such as genistein, 

genistin, daidzein, daidzin, coumestrol, microestrol, and 

deoxymiroestrol. Pueraria candollei var. candollei and P. candollei 

var. mirifica was used as the source of Kwao Keur. Using botanical 

characters is difficult to separate one from another varieties. The 

aim of present investigation is to distinguish between P. candollei 

var. candollei and P. candollei var. mirifica using the random amplified 

polymorphic DNA (RAPD) analysis. Genetic variation among 

sixteen plants of P.candollei var. candollei and P. candollei var. 

mirifica collected from three areas of Thailand was analyzed using 

eleven oligonucleotide primers by RAPD method. The number of 

polymorphic/monomorphic bands among the pairwise combinations 

of the plants and the total number of bands were determined to categorize 

all the plants in different groups of similarity and dissimilarity 

at the genetic level. The procedure described also allowed the 

identification and discrimination of the varieties of Pueraria 

candollei. Furthermore, secondary metabolites of the tuberous roots 

of both varieties were profiled using LC-MS spectroscopy. Puerarin 

was identified as a major component of the tuberous root extracts. 

(This work was supported by the National Research Council of Thailand 

and NRCT-JSPS Core University Program. Presented at the 

sixth JSPS-NRCT Joint Seminar: Recent advances in Natural Medicine 

Research, December 2-6, 2003, Bangkok, Thailand.)


IN VITRO PROPAGATION OF MALLOTUS 

REPANDUS (WILLD.) MUELL. ARG. (NO. 555) 

Songsri Kaewsuwan 1,2 , Noppamas Soonthornchareonnon 2 , 

Sompop Prathanturarug 1 


Faculty of Pharmacy, Mahidol University, E-mail : 

pyspr@mahidol.ac.th 

Key words : Mallotus repandus, in vitro propagation, medicinal 

plant, anti-inflamatory drug 

An in vitro propagation protocol has been developed for 

Mallotus repandus (Willd.) Muell. Arg. (Euphorbiaceae), a potential 

medicinal plant for anti-inflammatory drug development. Nodal 

segments and shoot tips from 2 month-old appearing shoots were 

aseptically cultured on solid Murashige and Skoog (MS) basal medium 

supplemented with three cytokinins alone or in combination 

with 1-naphthylacetic acid (NAA) to induce shoot formation for 8 

weeks. Of the three cytokinins tested (6-benzylaminopurine (BA), 

isopentenylaminopurine (2iP), and kinetin (Kin), BA induced shoot 

development most efficiently. The best shoot regeneration rate (3.36 

shoots/response explant) was obtained when the explants were cultured 

on MS medium supplemented with 4,44 μM BA. Addition of 

0.54 μM NAA to the media suppressed shoot induction rate, but 

induced callus formation. The regenerated shoots were cut and cultured 

on MS medium containing 10.74-53.71 μM NAA for rooting. 

The best root induction (73.08%) was revealed using MS medium 

supplemented with 32.23 μM NAA. Regenerated plants were successfully 

transferred to soil and grown under greenhouse conditions. 

The stem of the regenerants will be chemicall analysed to compared 

with their mother plants. 

(This work was supported by the National Research Council of Thailand. 

Presented at the 3rd World Congress on Medicinal and Aromatic 

Plants for Human Welfare, February 3-7, 2003, Chiangmai, 

Thailand.) 

MANAGENESE COMPLEXES OF CURCUMIN 

AND ITS DERIVATIVES : EVALUATION FOR 

THE RADICAL SCAVENGING ABILITY AND 

NEUROPROTECTIVE ACTIVITY (NO. 556) 

Opa Vajragupta 1 , Preecha Boonchong 1 , Hiroshi Watanabe 2 , 

Michihisa Tohda 2 , Naparat Kummasud 1 , and Yaowared 

Sumanont 1 

1 Faculty fo Pharmacy, Mahidol University, Bangkok, Thailand 

and 2 Institute of Natural Medicine, Toyama Medical and Pharmaceutical 

University, Toyama, Japan 

Key words : Managanese complex, Superoxide dismutase mimic, 

2-Vesselocclusion, MPTP, Dopamine 

In this study, three managanese complexes of curcumin 

(Cp) and related compounds, diacetylcurcumin (AcyICp) and ethylenediamine 

derivative (CpED), were synthesized and evaluated in 

vitro for antilipid peroxidation and superoxide dismutase activity. 

The manganese complexes exhibited a great capacity to protect brain 

lipids against peroxidation with IC 50 of 6.3-26.3 μM. All manga- 

nese complexes showed much greater SOD activity than their corresponding 

antioxidant ligands as well as trolox with IC 50 values of 

8.9 - 29.9 μM. AcylCp and curcumin manganese complexes 

(AcylCpCpx and CpCpx) also gave the highest inhibitory activity to 

H 2 O 2 -induced cell damage (oxidative stress) at 0.1 μg/ml (0.2 μM) 

in NG108-15 cells, which were more potent than curcumin and related 

compounds. The neuropharmacological tests in mice supported 

the idea that the SOD mimicking complexes were able to penetrate 

to the brain as well as their role in the modulation of brain neurotransmitters 

under the aberrant conditions. The complexes significantly 

improved the learning and memory impairment induced 

by transient ischemic/reperfusion. AcylCpCpx, CpCpx, and 

CpEDCpx showed significant protection at 6.25, 25 and 50 mg/mg 

(i.p.), respectively, whereas managanese acetate and curcumin had 

no effect at doses of 50 mg/kg. In addition, treatment of AcylCpCpx 

and curcumin significantly attenuated MPTP-induced striatal dopamine 

depletion in mice, which was in accordance with the increase in 

the density of dopaminergic neurons when compared with MPTPtreated 

mice. These results support the important role of manganese 

in importing SOD activity and consequently, the enhancement of 

radical scavenging activity. AcylCpCpx and CpCpx seem to be the 

most promising neuroprotective agents for vascular dementia. 

(Free Rodical Biology & Medicine, 35(12), 1632-44, 2003.) 

MANGANESE-BASED COMPLEXES OF RADICAL 

SCAVENGERS AS NEUROPROTECTIVE AGENTS 

(NO. 557) 

Opa Vajragupta 1 , Preecha Boonchong 1 , Yaowared Sumanont 1 , 

Hiroshi Watanabe 2 , Yuvadee Wongkrajanga and Naparat 

Kammasuda 

1 Faculty of Pharmacy, Mahidol University, 447 Sri-Ayudhya 

Road, Bangkok, Thailand, 2 Institute of Natural Medicine, 

Toyama Medical and Pharmaceutical University, 2630 Sugitani, 

Toyama 930-0194, Japan. 

Key words : Managese complex, Superoxide dismutase mimic, Radical 

scavengers 

Managese was incorporated in the structure of the selected 

antioxidants to mimic the superoxide dismutase (SOD) and to increase 

radical scavenging ability. Five manganese complexes (1-5) 

showed potent SOD activity in vitro with IC 50 of 1.18-1.84 mM and 

action against lipid peroxidation in vitro with IC 50 of 1.97-8.00 mM 

greater than their ligands and trolox. The manganese complexes 

were initially tested in vivo at 50 mg/kg for antagonistic activity on 

methamphetamine (MAP)-induced hypermotility resulting from 

dopamine release in the mice brain. Only manganese complexes of 

kojic acid (1) and 7-hydroxyflavone (3) exhibited the significant 

suppressions on MAP-induced hypermotility and did not significantly 

decrease the locomotor activity in normal condition. Manganese 

complex 3 also showed protective effects against learning and 

memory impairment in transient cerebral ischemic mice. These results 

supported the brain delivery and the role of manganese in SOD 

activity as well as in the modulation of brain neurotransmitters in 

the aberrant condition. Manganese complex 3 from 7-hydroxyflavone 

was the promising candidate for radical implicated neurodegenerative 

diseases. 

(Bioorganic & Medicinal Chemistry 2003, 11, 2329-2337.)

210 

CYTOPROTECTIVE AND CYTOTOXIC EFFECTS 

OF CURCUMIN: DUAL ACTION ON H 2 O 2 -INDUCED 

OXIDATIVE CELL DAMAGE IN NG 108-15 CELLS 

(NO. 558) 

Promote Mahakunakorn 1 , Michihisa Tohda 1 , Yukihisa 

Murakami, A Kinzo Matsumoto 1 , Hiroshi Watanabe 1 , Opa 

Vajaragupta 2 

1 Institute of Natural Medicine, Yoyama Medical and 

Parmaceutical University, Yoyama, Japan. 2 Faculty of Pharmacy, 

Mahidol University, Bangkok, Thailand. 

Key words : Manganese complex, Superoxide dismutase mimic, 

Radical scavengers 

The ability of curcumin, a natural antioxidant isolate from 

Curcuma longa, to inhibit hydrogen peroxide (H 2 -O 2 )-induced cell 

damage in NG 108-15 cells was examined. When added simultaneously 

with 500 mM H 2 -O 2 , curcumin (25-100 mM) effectively 

protected cells from oxidative damage. However, when the cells 

were pretreated with curcumin (25-100 mM) for 1.5 h before H 2 O 2 

exposure, curcumin was unable to inhibit H 2 O 2 -induced cell damage. 

Instead, it caused a significant concentration-dependent decrease 

in cell viability after H 2 O 2 exposure. This dual action of 

curcumin suggests that pretreatment with curcumin by itself did not 

have any significant effect on the viability of the NG 108-15 cells, 

but it sensitized them to oxidative damage induced by H 2 O 2 under 

our experimental conditions. It appears that these events may not 

relate to the antioxidant and free radical scavenging activities of 

curcumin. 

(Biol. Pharm Bull 26(5), 725-28, 2003.) 

SIMULTANEOUS DETERMINATION OF 

PARACETAMOL AND CHLORPHENIRAMINE 

MALEATE BY MICELLAR ELECTROKINETIC 

CHROMATOGRAPHY (NO. 559) 

Leena Suntornsuk 1 , Ongart Pipitharome 1 , Prapin Wilairat 2 

1 Department of Pharmaceutical Chemistry, Faculty of Pharmacy, 

2 Department of Chemistry, Faculty of Science, Mahidol University, 

Bangkok 10400, Thailand (pyll@mahidol.ac.th). 

Key words : micellar electrokinetic chromatography (MEKC), 

paracetamol (PARA), chlorpheniramine maleate (CPM), cold remedy 

ingredients 

A micella electrokinetic chromatography (MEKC) method 

was established for determination of paracetamol (PARA) and 

chlorpheniramine maleate (CPM) in cold tablets. Separation of both 

drugs as well as other seven cold remedy ingredients was achieved 

in 25.5 min using a sodium dihydrogenphosphate-sodium tetraborate 

buffer (10 mM, pH 9.0) containing sodium dodecyl sulfate (SDS) 

(50 mM) and acetonitrile (26% v/v). The effective capillary length 

of 50 cm, the separating voltage of 15 kV and the temperature of 

30 o C was optimized. Detection was by a diode array detector at 214 

nm. Method linerity was excellent (r 2 > 0.999) over the concentration 

tested (10-250 μg/mL) with good precision and accuracy. Recoveries 

were good (> 99%) with limits of detection of 0.4 and 0.5 

μg/mL and limits of quantitation of 2 (%RSD = 3.1%) and 4 (%RSD 

= 2.4%) μg/mL, for PARA and CPM, respectively. The developed 

method was applied for determination of ingredients in cold tablets 

and was found to be simple, rapid and efficient. 

(J Pharm. Biomed. Anal 2003, 33:441-449.) 

METHOD DEVELOPMENT FOR SEPARATION 

OF ACTIVE INGREDIENTS IN COLD MEDICINES 

BY MICELLAR ELECTROKINETIC CHROMATO- 

GRAPHY (NO. 560) 

Leena Suntornsuk 1 , Ongart Pipitharome 1 , Prapin Wilairat 2 


Mahidol University, 447 Sri-Ayudhaya Rd., Rajathevee; 2 Department 

of Chemistry, Faculty of Science, Mahidol University, Rama 

VI Rd., Rajathevee, Bangkok 10400, Thailand (pylll@ 

mahidol.ac.th) 

Key words : cold medicine ingredients, micellar electrokinetic chromatography 

(MEKC) 

Separation of nine commonly used active ingredients in 

cold medicines, were demonstrated by micellar electrokinetic chromatography. 

The ingredients included paracetamol, chlorpheniramine 

maleate, diphenhydramine hydrochloride, triprolidine hydrochloride, 

phenlpropanolamine hydrochloride, dextromethorphan 

hydrobromide, loratadine, aspirin and caffeine. Effects of buffer 

concentrations, pH, organic modifiers and capillary length were investigated. 

The optimum conditions were achieved in 10 mM sodium 

dihydrogenphosphate-sodium tetraborate buffer, pH 9.0, containing 

50 mM sodium dodecyl sulfate and 28% v/v acetonitrile using 

the effective length of 50 cm, the separating voltage of +15 kV 

and the capillary temperature of 30 o C. Separation of all peaks was 

obtained within 28.4 min with a resolution of 1.2. 

(Sci. Pharm. 2003, 71: 235-250.) 


QUANTITATIVE ANALYSIS OF AGLYCONE 

QUERCETIN IN MULBERRY LEAVE (MORUS 

ALBA I.) BY CAPILLARY ZONE ELECTRO- 

PHORESIS (NO. 561) 

Leena Suntornsuk 1 , Saowapak Kasemsook 2 , Surapote Wongyai 2 


Mahidol University, Bangkok, Thailand; 2 Faculty of Pharmacy, 

Rangsit University, Bangkok, Thailand. (pylll@mahidol.ac.th) 

Key words : Aglycone queretin, capillary electrophoresis, rutin 

A capillary zone electrophoresis method was established 

for analysis of aglycone quercetin in mulberry leaves (Morus alba 

L.). Influence of background electrolyte concentrations and pHs, 

surfactant concentrations, organic solvents, oven temperature and 

voltage on the separation of aglycone quercetin, other flavonoids 

with related structures and a phenolic acid, were systematically investigated. 

The optimum condition providing baseline separation 

of all compounds within 16.5 min was obtained in 150 mM boric 

acid (pH 10.0) using a fused silica capillary with an effective length 

of 42.5 cm (50 μm, inner diamete), oven temperature 0f 32 o C and


applied voltage of 15 kV. Linearity of the method was excellent (r 2 

> 0.999) over the concentration tested (40-160 μg/mL). The relative 

standard deviations (%RSDs) from injection, intraday and inter-day 

precision using rutin as an internal standard were less than 2.5%. 

Recoveries were good (= 100.0%, %RSD = 0.8%) with a limit of 

detection and limit of quantitation of 0.86 and 3.16 μg/mL (%RSD 

= 1.8%), respectively. The aglycone quercetin found in the mulberry 

leaves was 0.194 g/100 g (%RSD = 0.9%) on dry weight. 

(Electrophoresis 2003, 24: 1236-1241. 3 rd International Symposium 

on Separation in the BioScience 5-8 May 2003, Moscow, Russia.) 

FEATHER DEGRADATION BY BACILLUS SP. 

FS 46 IN SUBMERGED CULTIVATION (NO. 562) 

Worapot Suntornsuk 1 , Leena Suntornsuk 2 

1 Department of Microbiology, Faculty of Science, King Mongkut’s 

University of Technology Thonburi, Bangkok 10140, Thailand; 


Mahidol University, Bangkok 10400, Thailand. 

Key words : Feather; Biodegradations; Bacillus sp.; Keratinase; 

Animal feed 

Cultivation conditions affecting feather degradation by 

Bacillus sp. FK 46 were investigated. The results showed that feather 

was almost completely degraded under the following conditions : 

1% whole chicken feather as a substrate at the initial medium pH of 

9 with 5% bacterial inoculum, at a temperature of 37 o C and a shaking 

speed of 250 rev/min. Glucose, methanol, Tween 80 and Triton 

X-100, however, had no effect on feather degradation. After feather 

was degraded, its residue and fermented broth would become a protein 

feed for animals. 

(Bioresouce Tech. 2002, 86 : 239-243.) 

ANTI-VENOM POTENTIAL OF BUTANOLIC- 

EXTRACT OF ECLIPTA PROSTRATA AGAINST 

MALAYAN PIT VIPER VENOM (NO. 563) 

Pimolpan Pithayanukul 1 , Sasitorn Laovachirasuwan 1 , Rapepol 

Bavovada 2 , Narumol Pakmanee 3 , Rutt Suttisri 2 

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

2 Faculty of Pahrmaceutical Sciences, Chulalongkorn University, 

Bangkok, Thailand. 3 Queen Saovabha Memorial Institute, Thai 

Red Cross Society, Bangkok, Thailand. 

Key words : Eclipta prostrata; Malayan Pti viper; Anti-snake venom 

The butanolic and purified butanolic extracts of Eclipta 

prostrata were evaluated for their anti-venom potential. Inhibition 

of letha, hemorrhagic, proteolytic, and phospholipase A 2 activites of 

Calloselasma rhodostoma (Malayan pit viper, MPV) venom by these 

extracts were determined. Demethylwedelolactone was indentified 

as their major constituent. The butanolic extract, at 2.5 mg/mouse, 

was able to completely neutralize teh lethal activity of 2LD 50 of 

MPV venom, but increasing the dose diminished the effect. The 

purified butanolic extract, at 1.5-4.5 mg/mouse,was able to neutralize 

the lethality o the venom at around 50-58%. Both extracts par- 

tially inhibited the hemorrhagic activity but displayed very low antiphospholipase 

A 2 activity and did not inhibit proteolytic activity of 

MPV vemon. 

(Journal of Ethnopharmacology 2004, 90(2-3): 347-352.) 

VARIATION OF ANTHRAQUINONE CONTENT 

IN SENNA SIAMEA (LAM.) IRWIN & BARNEBY 

LEAVES (NO. 564) 

Wandee Gritsanapan and Somsak Nualkaew 

Department of Pharmacognosy, Faculty of Pharmacy, Mahidol 

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

Key words : Senna siamea, Cassia siamea, Anthraquinone content 

Senna siamea (Lam.) Irwin & Barneby or Cassia siamea 

Lamk. which is a native plant of South East Asia is widely cultivated 

in all parts of Thailand. The young leaves and flowers of this 

plant were used to be recommended as a bitter tonic, mild laxative 

drug and sleeping aid. A sleeping aid action is due to CNS depressant 

activity of a major component, barakol while a laxative effect is 

due to anthraquinone components. Since glycosides of anthraquinones 

in plants are reported to be the active form of laxative action, 

plants containing higher yield of anthraquinone glycosides should 

have a better laxative potency. Thus, quantitative analysis of anthraquinone 

content in the leaves of S. siamea collected from different 

geographical areas of Thailand in all seasons were studied. By 

UV-vis spectrophotometric method, the content of total anthraquinones 

in the leaves collected from northern, northeastern, central and 

southern parts of Thailand in summer, rainy season and winter were 

found to be 0.07 - 0.20% while the content of total anthraquinone 

glycosides were 0.06-0.12% dry weight. The sample containing highest 

total anthraquinone content (0.20%) was the northern sample collected 

in summer while the highest total anthraquinone glycosides 

content (0.12%) was the central-winter sample. It was shown that 

total anthraquinone glycoside content in the leaves of S. siamea growing 

in all parts of Thailand was high in winter and summer. So, 

collection of the leaves of this plant for laxative drug should be done 

in winter and summer. However, the variation of both total anthraquinones 

and total anthraquinone glycosides in the leaves collected 

from several areas and seasons were not significantly different 

(p < 0.05). Due to anthraquinone glycoside content, S. siamea 

leaf is not a good source for anthraquinone laxative drug. 

(The research was partially granted by Plant Genetic Conservation 

Project, The Royal Chitralada Project, Dusit Palace. Proceeding 

Book of a Seminar on Thai Natural Resources : Nature of Life, 

Bangkok, 10-12 May 2003.) 

ANTIOXIDANT ACTIVITY OF AZADIRACHTA 

INDICA (NO. 565) 

Pongtip Sithisarn 1 , Rungtawan Supabphol 2 , and Wandee 

Gritsanapan 1 

1 Department of Pharmacognosy, Faculty of Pharmacy, Mahidol 

University; 2 Department of Physiology, Faculty of Medicine, 

Srinakarintarawirot University. E-mail : pywgs@mahidol.ac.th

212 

Key words : Antioxidant activity, Azadirachta indica, Siamese neem 

tree 

Siamese neem tree (Azadirachta indica A. Juss var. 

siamensis Valeton, Meliaceae) is a tropical plant found in several 

countries such as Southern Laos, Western Cambodia and also in 

Thailand. There are many reports concerning its medicinal properties 

such as antipyretic, antidiarrhea, insecticidal and antioxidant 

activities. Antoixidant activity of the bark extract of this plant compared 

to standard antioxidant was reported to be high (TEAC = 

0.9130). It was also mentioned about cancer chemopreventive components 

of Siamese neem tree flowers and antioxidant activity of the 

seed extract. Thus, antioxidant activity of several parts of this plant 

are interesting to be investigated for more details. Several parts of 

Siamese need tree i.e. leaf, flower, ripe fruit rind, ripe fruit flesh, raw 

fruit, seed and stem bark were studied for antioxidant activity using 

DPPH assay. It was found that aqueous leaf extract,ethanol flower 

extract and ethanol stem bark extract showed quite high activities 

(EC 50 = 26.48, 27.91 and 30.55 μg/ml, respectively) but when using 

ABTS-metmyoglogin assay, the activities at 1 mg/ml concentration 

of the extracts were shown to be 0.9592, 0.9881 and 1.0640 mmol/ 

L, respectively. The leaf samples are easy for collection in all seasons 

while the flower and bark samples can be collected in some 

seasons only. So, active components from the leaves of Siamese 

neem tree would be further separated and identified. 

(The research was partially granted by National Center for Genetic 

Engineering and Biotechnology (BIOTEC). Proceeding Book of 

Seminar on Thai Natural Resources : Nature of Life, Bangkok, 10- 

12 May 2003.) 

COMPARISON OF ANTIOXIDANT ACTIVITY OF 

TWO VARIETIES OF AZADIRACHTA INDICA 

(NO. 566) 


Gritsanapan 1 


Univesity, 2 Department of Physiology, Faculty of Medicine, 

Srinakarintarawirot University. E-mail : pywgs@mahidol.ac.th 

Key words : Siamese neem tree, neem, antioxidant activity 

Three parts of neem and Siamese need tree i.e. leaf, flower 

and stem bark were extracted and tested for antioxidant activity using 

DPPH scavenging assay and total antioxidant activity determination. 

It was found that EC 50 of antioxidant activities tested by 

DPPH scavenging assay from leaf aqueous extract, flower ethanol 

extract and stem bark ethanol extract of Siamese neem tree were 

26.48, 27.91 and 30.55 μg/mL, respectively while the results from 

neem were 104.73, 89.04 and 11.05 μg/mL, respectively. The total 

antioxidant activities of leaf aqueous extract, flower ethanol extract 

and stem bark ethanol extract of Siamese neem tree tested by total 

antioxidant activity determination were 0.9592, 0.9881 and 1.0640 

mmol/L, respectively while the results from neem were 0.8122, 

0.6793 and 1.2318 mmol/L, respectively. 


Engineering and Biotechnology (BIOTEC). Proceeding Book of the 

3 rd Indochina Conference on Pharmaceutical Science III Bangkok, 

20-23 May 2003.) 


PHENOLIC COMPONENTS FROM MELODORUM 

SIAMENSE (NO. 567) 

Junya Intaranongpai 1 and Wandee Gritsanapan 2 

1 Pharmaceutical Chemistry and Phytochemistry Graduate 

Programme, Faculty of Pharmacy, Mahidol University, Bangkok, 

Thailand; 2 Department of Pharmacognosy, Faculty of Pharmacy, 

Mahidol University, Bangkok, Thailand. E-mail : pywgs@ 

mahidol.ac.th 

Key words : Melodorum siamense, Annonaceae, Nom maeo 

Melodorum siamense Scheff. (Rauwenhoffia siamensis 

Scheff., Annonanceae) which is called in Thai as Nom maeo (central 

is a climber usually found in the central part of Thailand (1-5). Its 

flowers have been traditionally used for cardiotonic agent while the 

wood is used as antipyretics (6). This plant is botanically related to 

M. fruiticosum Lour. which its leaves and branches were reported to 

contain butenolides with cytotoxic activity in several tumor cell lines 

(4). There is no report about chemical constituent and biological 

activity of M. siamense before. Thus, the aim of this study is to 

investigate the chemical constituents of the wood and stem bark of 

this plant. 

By column chromatographic technique, two phenolic 

compounds which are 1-(3,4,5-trimethoxy benzene)-1-methoxy-1ethylene-2-benzoate 

ester and 1,2-bezenedi carboxylic acid bis(5methylheptyl)ester 

were isolated from the n-hexane extract of the 

wood and ethyl acetate extract of the stem bark, respectively. The 

structure elucidations of the compounds were done based on spectroscopic 

and reported data. The compounds should be further investigated 

for cytotoxic activity to compare with reported activity of 

M. fruiticosum. 

(Proceeding Book of the 3 rd Indochina Conference on Pharmaceutical 

Science III Bangkok, 20-23 May 2003.) 

ANTIOXIDATIVE ACTIVITY OF EXTRACTS 

FROM NEEM AND SIAMESE NEEM LEAF, 

FLOWER AND STEM BARK (NO. 568) 


Gritsanapan 1 


University; 2 Department of Physiology, Faculty of Medicine, 

Srinakarintarawirot University. E-mail : pywgs@mahidol.ac.th 

Key words : antioxidative activity, Neem, Siamense neem 

Neem (Azadirachta indica A. Juss var. indica) and 

Siamese neem tree (Azadirachta indica A. Juss var. siamensis) are 

plants in Meliaceae family. Siamese neem tree is usually found in 

Thailand, Southern Lao and Western Cambodia while neem is always 

found in India, Pakistan, Indonesia, Africa and also Thailand. 

There are many reports indicating their biological activities such as 

antipyretic, antimalarial, cancer chemopreventive, insecticidal and 

also antioxidant activities. Because of their close taxonomic relationship, 

so the comparison of their antioxidant activities is interesting 

for investigation.


Three parts of neem and Siamese neem tree i.e. leaf, flower 

and stem bark were extracted and tested for antioxidant activities 

using DPPH assay. It was found that EC 50 of antioxidant activities 

from leaf aqueous extract, flower ethanol extract and stem bark ethanol 

extract of Siamese neem tree were 26.48, 27.91 and 30.55 μg/ 

mL, respectively while the results from neem were 104.73, 89.04 

and 11.05 μg/mL, respectively. 

The results show that all extracts from Siamese neem tree 

have quite high antioxidant activities while neem extracts show lower 

activities except the stem bark ethanol extract. From these results, 

antioxidant activity of both plants should be confirmed by other 

methods. Separation, purification and identification of active components 

should be further investigated. 


Engineering and Biotechnology (BIOTEC). Presented at RGJ Seminar 

Series XXIV : Pharmaceutics and Pharmaceutical Technology, 

Bangkok, 15-16 October, 2003.) 

COMPARISON OF ANTIOXIDANT ACTIVITIES 

OF THE LEAVES AND FLOWERS OF SIAMESE 

NEEM TREE (NO. 569) 

Pongtip Sithisarn 1 , Wandee Gritsanapan 1 , and Rungtawan 

Supabphol 2 


University, 2 Department of Physiology, Faculty of Medicine, 

Srinakarintarawirot University. 

Key words : antioxidant, antioxidant activity, Siamense Neem Tree, 

Azadirachta indica 

Siamese neem tree (Azadirachta A. Juss var. siamensis 

Valeton) is the plant that young leaves and flowers have been popularly 

used as vegetables of bitter tonic. The leaf aqueous and flower 

ethanol extracts of Siamese neem tree were tested for free radical 

scavenging activity using DPPH scavenging assay, total antioxidant 

activity determination and confirmed for antioxidant activity on lipid 

peroxidation in bronchogenic cancer cell culture using TBARS 

method. The results of DPPH scavenging assay showed that leaf 

aqueous and flower ethanol extracts exhibited antioxidant activities 

with EC 50 of 26.48 + 3.63 and 27.91 + 2.73 μg/ml, respectively while 

their antioxidant activities at 1 mg/ml were 0.9592 and 0.9881 mmol/ 

l when tested by total antioxidant activity determination. On bronchogenic 

cancer cell culture, treatment of both extracts at concentration 

100 μg/ml could decrease MDA level significantly from 148.72% 

to 100.97% and from 148.72 to 105.53%, respectively. The results 

show that both leaves and flowers exhibit high antioxidant activity. 

So, consuming these parts of Siamese neem tree as vegetables should 

be beneficial for the body and confirm traditional use of this plant as 

bitter tonic. 


Engineering and Biotechnology (BIOTEC). Presented at The 6 th 

JSPS-NRCT Joint Seminar “Resent Advances in Natural Medicine 

Research” Bangkok, 2-4 December 2003.) 

ACUTE TOXICITY OF A THAI TRADITIONAL 

PREPARATION : PRASAPLAI (NO. 570) 

Somsak Nualkaew 1 , Wandee Gritsanapan 1 , and Chongkol 

Tiangda 2 


University, 2 Department of Pharmacology, Faculty of Pharmacy, 

Mahidol University. E-mail : pywgs@mahidol.ac.th 

Key words : Prasaplai, traditional medicine, acute toxicity 

Administration of the water suspension of the Prasaplai 

preparation, a Thai traditional medicine, up to 2.5 g/kg and the water 

extract of up to 20 g/kg (calculated as powdered drug) which was 

the maximum dose for rats, did not produce any signs and symptoms 

of toxicity in rats during the first 24 hrs. and no rats died at these 

dose levels. After observation for a further 14 days, no toxic symtom 

nor dead rats were observed. 

(The research was partially granted by The Royal Golden Jubilee 

Ph.D. Program, The Thailand Research Fund. Presented at RGJ 

Seminar Series XXIV : Pharmaceutics and Pharmaceutical Technology, 

Bangkok, 15-16 Octover, 2003.) 

ANTISPASMODIC EFFECT OF A THAI 

TRADITIONAL PREPARATION : PRASAPLAI 

(NO. 571) 

Somsak Nualkaew 1 , Wandee Gritsanapan 1 and Chongkol 

Tiangda 2 


University, 2 Department of Pharmacology, Faculty of Pharmacy, 

Mahidol University. E-mail : pywgs@mahidol.ac.th 

Key words : Antispasmodic effect, Prasaplai, Dysmenorrhea 

The aqueous extract of Prasaplai, a Thai traditional medicine 

was studied on the isolated rat uterus in the estrous stage. The 

uterine contraction was induced by different agonists. The results 

showed that the aqueous extract of Prasaplai could inhibit the contraction 

of uterine muscle induced by submaximal doses of acetylcholine 

(2.04 x 10 -4 mg/ml), oxytocin (1.54 x 10 -4 mg/ml) and prostaglandin 

E 2 (PGE 2 ) (6.00 x 10 -4 mg/ml). The inhibition was concentration-dependent 

and reversible by tissue washing. The IC 50 of 

Prasplai extract expressed as milligram of powdered preparation per 

ml of perfusion solution for acetylcholine, oxytocin and PGE 2 were 

11.70 mg/ml 10.00 mg/ml and 5.75 mg/ml, respectively. These data 

suggest that the Prasaplai has an antispasmodic effect against uterine 

contraction with various mechanisms and corroborate the traditional 

use in the treatment of dysmenorrhea. 

(The research was partially granted by The Royal Golden Jubilee 

Ph.D. Program, The Thailand Research Fund. Presented at The 6 th 

JSPS-NRCT Joint Seminar “Resent Advances in Natural Medicine 

Research” Bangkok, 2-4 December 2003.)

214 

PHYTOCHEMISTRY AND ANTI-MALARIAL, 

ACTIVITY OF EUPATORIUM ODORATUM L. 

(NO. 572) 

Rungnapa Ongkana 1 , Peerapan Tan-ariya 2 , Weena 

Jiratchariyakul 1 and Aimon Somanabandhu 1 


University, 2 Department of Microbiology, Faculty of Science, 

Mahidol University. E-mail : pyasm@mahidol.ac.th 

Key words : Eupatorium odoratum, antimalarial constituents 

Eupatorium odoratum L. is a common scadent shrub, native 

to central and tropical South America as well as to Southeast 

Asia. In a preliminary study, Eupatorium odoratum L., a plant traditionally 

used in Thailand to treat fever and malaria, was found to 

have in vitro antiplasmodial property against Plasmodium falciparum. 

the present study is designed to search for active compounds and to 

confirm the anti-malarial activity of this plant. The leaves of Eupatorium 

odoratum L. were collected from roadsides in Ratchaburi. 

The chloroform extract was found to be active against P. falciparum 

with IC 50 value of 9 μg/ml. Column fractionation of chloroform 

extract gave six fractions, three of which were found to be active. 

The compounds isolated from active fraction(s) were identified using 

spectroscopic techniques and were further tested for their antimalarial 

activity. 

(The research was partially supported by a research grant from 

Mahidol University. Presented at Joint International Tropical Medicine 

Meeting 2002 “The 8 th Chamlong-Tranakchit Harinasuta Lecture” 

Montien Riverside Hotel, Bangkok, Thailand, 20-22 November 

2002.) 

PHYTOCHEMICAL AND BIOLOGICAL ACTIVITY 

OF ANOMIANTHUS DULCIS STEM BARK. (NO. 573) 

Lersak Ubonopas 1 , Noppamas Soonthornchareonnon 1 , Khanit 

Suwanborirux 2 , Prapai Wongsinkongman 3 , Wongsatit Chuakul 4 


University, Bangkok 10400, Thailand; 2 Department of Pharmacognosy, 

Faculty of Pharmaceutical Sciences, Chulalongkorn University, 

Bangkok 10330, Thailand; 3 Department of Medical Sciences, 

Institute of Medicinal Plant Research, Ministry of Public 

Health, Nondhaburi, 11000 Thailand; 4 Department of Pharmaceutical 

Botany, Faculty of Pharmacy, Mahidol University, 

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

Key words : cytotoxicity, Anomianthus dulcis, 10-methoxyliriodenine 

Anomianthus dulcis (Dun.) J. Sincl. is an annonaceous 

plant. It has been used as a folklore medicine in Thailand for fever 

treatment. Alcoholic extract (F001) of the stem bark was partitioned 

between dichloromethane and water (1:1), providing the water crude 

extract (F002) and dichloromethane crude extract (F003). F003 was 

further partitioned between hexane and 90% methanol (1:1), providing 

the methanol crude extract (F006) and hexane crude extract 

(F007), respectively. F002 was further partitioned with ethyl acetate 

to afford the ethyl acetate crude extract (F005) and residue 

(F004), and the fractions were concentrated under vacuum. The bio- 


logical activity of fractions were screened for cytotoxicity by using 

brine shrimp lethality test (BST) and human tumor cell line panel. 

F006 actively exhibited in BST (LD 50 0.33 μg/ml) and selectively 

cytotoxic to breast cancer cell line (MCF-7) and vincristine resistant 

cancer cell line (HCT-8) at the concentration 20 μg/ml. Bioassayguided 

fractionation led to teh isolation of two known compounds 

which were oxoaporphine alkaloids namely 9-methoxyliriodenine 

(1) by spectral data, as well as comparison with literature data. Both 

compounds were the first discovery in this plant and this study, was 

the first report of cytotoxic activity of the extract from A. dulcis 

stem bark. 

(The research was partially granted by Mahidol University. The paper 

was presented at The Sixth JSPS-NRCT Join Seminar : Recent 

Advances in Natural Medicine Research, Faculty of Pharmaceutical 

Sciences, Chulalongkorn University, December 2-4, 2003.) 

STUDIES OF PHYTOCHEMISTRY AND ACARI- 

CIDAL ACTIVITY FOR HOUSE DUST MITE 

FROM TRIGONOSTEMON REIDIOIDES CRAIB 

ROOTS (NO. 574) 

Monnun Sakayarojkul 1 , Noppamas Soonthorncharoennon 1 , 

Masahiko Isaka 2 , Vanna Mahakittikun 3 and Wongsatit Chuakul 4 


University, Bangkok 10400, Thailand. 2 National Center for Genetic 

Engineering and Biotechnology (BIOTEC), Nation Science 

and Technology Development Agency (NSTDA), Pathumthani 

12120, Thailand. 3 Department of Parasitology, Faculty of Medicine 

Siriraj Hospital, Mahidol University, Bangkok 10700, Thailand. 

4 Department of Pharmaceutical Botany, Faculty of Pharmacy, 

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

pynsk@mahidol.ac.th 

Key words : acaricidal activity, Trigonostemon reidioides, 2demethyl 

rediocide A 

Acaricidal activity of T. reidioides root extracts was investigated 

on Dermatophagoides pteronyssinus (D. p), the most com 

mon house dust mite in Thailand. 

Benzyl benzoate was used as posi 

tive acaricidal compound which 

showed LC 50 = 7.20x10 -3 mg/cm 2 

after 3 days of incubaion time. 

Bioassay-guided fractionation of 

the hexane crude extract (LC 50 = 

0.60 mg/cm 2 ) led to the isolation 

of fraction II which composed of a 

novel compound, 2-demethyl 

rediocide A, together with the 

known compound, rediocide A, 

and two unidentified compounds. 

Fraction II (LC 50 = 3.27x10 -3 mg/ 

cm 2 ) showed almost 2-fold more active than benzyl benzoate. The 

structural elucidations were based on analysis of the spectroscopic 

and literature data. 

(The research was partially granted by Mahidol University. The paper 

was presented at The Sixth JSPS-NRCT Join Seminar : Recent 

Advances in Natural Medicine Research, Faculty of Pharmaceutical 

Sciences, Chulalongkorn University, December 2-4, 2003.)

Mahidol University Annual Research Abstracts, Vol. 31

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