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African Journal of Pharmacy and Pharmacology Vol. 6(26), pp. 1890-1900, 15 July, 2012 Available online at http://www.academicjournals.org/AJPP DOI: 10.5897/AJPP12.128 ISSN 1996-0816 ©2012 <strong>Academic</strong> <strong>Journals</strong> Full Length Research Paper New solid phase extraction reversed phase high performance liquid chromatography ultraviolet (RP-HPLC-UV) method for simultaneous determination of tenofovir and emtricitabine in Chinese population Rui Tan 1 , Bo Yan 2 , Jingchuan Shang 2 , Junqing Yang 1 *, Wenxiang Huang 3 , Xiaoni Zhong 3 , Ailong Huang 3 and Mingfeng Wang 1 1 Department of Pharmacology, Chongqing Medical University, Chongqing, China. 2 Department of Pharmaceutical Analysis, Chongqing Medical University, Chongqing, China. 3 Institute for Viral Hepatitis, Key Laboratory of Molecular Biology on Infectious Diseases, Ministry of Education, Chongqing Medical University, Chongqing, China. Accepted 23 May, 2012 The objective of this study is to develop and validate a method of reversed phase high performance liquid chromatography (RP-HPLC) assay with ultraviolet (UV) detector after solid phase extraction (SPE) for simultaneous measure tenofovir and emtricitabine in Chinese population. Detection of analytes was performed with a Phecda C18 (250 � 4.6 mm, 5 μm), reversed-phase analytical column and a security guard column C18 (4 � 3.0 mm, 5 μm). An obviously extrusive separation was successively followed with the SPE method using BOND ELUT-C18 Varian columns and a 30 min gradient elution consisting of potassium phosphate monobasic buffer with 0.08% of triethanolamine (pH 3.52) and methanol with UV detector at 270 nm. The results of this study showed that the method used in this study was simple, accurate and sensitive with a wide linear range from 10 to 5000 ng/ml for both tenofovir and emtricitabine when 300 μl aliquots was analyzed. The intra- and inter-day precision and accuracy for both analytes were lower than ±20% at the limit of quantification (LOQ) and ±15% at the other quality control (QC) levels. The absolute recoveries for tenofovir and emtricitabine were 80.6 and 86.5%, respectively. This method is suitable for routinely monitor plasma concentrations of tenofovir and emtricitabine in Chinese population. Key words: High performance liquid chromatography ultraviolet (HPLC-UV), solid phase extraction (SPE), pre-exposure chemoprophylaxis (PrEP), tenofovir, emtricitabine. INTRODUCTION With a global estimate of 2.7 million, new human immunodeficiency virus (HIV) infections occurring worldwide each year and even more undiagnosed cases waiting to be discovered, discovering novel methods to help stem the spread of this virus is critical. While behavior change programs have contributed to dramatic reductions in the number of annual infections in the U.S. *Corresponding author. E-mail: cqjqyang2004@yahoo.com.cn. Tel: +86 23 6848 5071. Fax: +86 23 6848 5161. and many other nations, far too many individuals remain at high risk. With an effective vaccine years away, there is a growing body of evidence that antiretroviral drugs may be able to play an important role in reducing the risk of HIV infection (http:// www. cdc. gov, 2008). Preexposure chemoprophylaxis (PrEP) is being explored as a novel strategy to interrupt the spread of HIV in at-risk patient populations such as men who have sex with men (MSM), injecting drug users (IDU), and female sex workers (Clauson et al., 2009). The Centers for Disease Control and Prevention (CDC) is sponsoring three clinical trials of PrEP, and is participating in a University of
Figure 1. Chemical structure of tenofovir disoproxil fumarate (TDF). Figure 2. Chemical structure of emtricitabine. Washington-sponsored trial in Kenya and Uganda. Similar PrEP trials are also being conducted by Family Health International (FHI), the National Institutes of Health (NIH), FHI and the Microbicide Trials Network (MTN) (http:// www. cdc. gov, 2008). Recently, a project named feasibility study of PrEP to decrease the infection of HIV for High Risk Group in Western China has been initiated by Chongqing Medical University to study the safety, compliance and resistance of tenofovir and emtricitabine for PrEP. Some researchers believe that an antiretroviral drug taken as a daily oral preventative is one of the most important new prevention approaches of HIV infection and several sources of data suggest that the use of antiretroviral drugs in this manner may be effective. Animal studies have demonstrated that pre-exposure administration of tenofovir disoproxil fumarate (TDF) plus emtricitabine provided significant protection to monkeys exposed repeatedly to an HIV-like virus. These data, combined with the drug’s favorable resistance, safety profiles and long intracellular half times make TDF and TDF plus emtricitabine ideal candidates for HIV prevention trials (http:// www. cdc. gov, 2008). Tenofovir and emtricitabine are nucleoside/tide reverse transcriptase inhibitors (NRTI), commonly used to treat persons infected with HIV-1. VIREAD ® is a trademark for TDF of Gilead Sciences, Inc., which is a fumaric acid salt Tan et al. 1891 of the bis-isopropoxycarbonyloxymethyl ester derivative of tenofovir. It has a molecular formula of C19H30N5O10P•C4H4O4 and a molecular weight of 635.52. It’s structural formula is shown in Figure 1 (http://www.accessdata.fda.gov, 2008). It is an oral prodrug of tenofovir, an acyclic nucleoside phosphonate (nucleoside) analog of adenosine 5 , -monophosphate, which has potent activity against retroviruses and hepadnaviruses. Upon oral administration, it is converted in vivo to tenofovir by esterase hydrolysis, which is taken up by cells. And then tenofovir is activated to tenofovir disphosphate through phosphorylation reaction, the active form of tenofovir. Tenofovir disphosphate competes with the natural substate deoxyadenosine 5’triphosphate for incorporation into DNA during HIV transcription, which blocks the action of HIV reverse transcription, prevents further DNA chain elongation, and cause termination of viral DNA replication (Kwarney et al., 2004). Emtricitabine differs from other cytidine analogs in that it has a fluorine in the 5-position. It has a molecular formula of C8H10FN3O3S and a molecular weight of 247.24. Its structural formula is shown in Figure 2. It is phosphorylated by cellular enzymes to form emtricitabine 5’-triphosphate, which inhibits the activity of the HIV-1 reverse transcriptase (RT) by being competing with the natural substrate deoxycytidine 5’- triphosphate and by
Page 1 and 2: African Journal of Pharmacy and Pha
Page 3 and 4: Editors Sharmilah Pamela Seetulsing
Page 5 and 6: Electronic submission of manuscript
Page 7 and 8: Fees and Charges: Authors are requi
Page 9 and 10: Table of Contents: Volume 6 Number
Page 13 and 14: Minimum inhibitory concentration (M
Page 15 and 16: Kim et al. 1887 Table 4. Antimicrob
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Page 21 and 22: ng/ml by 4.9 ml blank plasma with 5
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A B Figure 3. Bax positive cells of
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African Journal of Pharmacy and Pha
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Table 1. Summary of specific reacti
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Figure 3. Effects of the sap of Jat
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Figure 1. A schematic diagram showi
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Chen et al. 1953 Figure 4. Concentr
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Figure 6. Representative photomicro
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shock. The features of PHC can expl
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Administration of single or repeate
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Table 1. Effects of green tea on SO
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Ozardali I, Bitiren M, Ali ZK, Zeri
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EXPERIMENTAL PROCEDURE Materials an
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Scavenging rate (%) Figure 1 Scaven
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A D Figure 5 Figure Effects 5. of E
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Figure 1. Level-spacing distributio
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Number of cases Pattern Shi et al.
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sodium was then dissolved in 1000 m
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Concentration (ppm) Concentration (
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various contagious diseases such as
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Table 3. Antibacterial MIC (µg/ml)
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Table 5. Contd. 1/5 14.7 ± 0.6 39.
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UPCOMING CONFERENCES 6th European C
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