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Multi-residue analytical methodology-based liquid chromatography-time-of-flight-mass spectrometry for the analysis of pharmaceutical residues in surface water and effluents from sewage treatment plants and hospitals

Al-Qaim FF, Abdullah MP, Othman MR, Latip J, Zakaria Z

J Chromatogr A, 2014 Jun 6;1345:139-53.
PMID: 24768127 DOI: 10.1016/j.chroma.2014.04.025

An analytical method that facilitated the analysis of 11 pharmaceuticals residue (caffeine, prazosin, enalapril, carbamazepine, nifedipine, levonorgestrel, simvastatin, hydrochlorothiazide, gliclazide, diclofenac-Na, and mefenamic acid) with a single pre-treatment protocol was developed. The proposed method included an isolation and concentration procedure using solid phase extraction (Oasis HLB), a separation step using high-performance liquid chromatography, and a detection procedure that applies time-of-flight mass spectrometry. The method was validated for drinking water (DW), surface water (SW), sewage treatment plant (STP) influent and effluent, and hospital (HSP) influent and effluent. The limits of quantification were as low as 0.4, 1.6, 5, 3, 2.2 and 11 ng/L in DW, SW, HSP influent and effluent, STP effluent, and STP influent, respectively. On average, good recoveries higher than 75% were obtained for most of the target analytes in all matrices. Matrix effect was evaluated for all samples matrices. The proposed method successfully determined and quantified the target compounds in raw and treated wastewater of four STPs and three hospitals in Malaysia, as well as in two SW sites. The results showed that a number of the studied compounds pose moderate to high persistency in sewage treatment effluents as well as in the recipient rivers, namely; caffeine, simvastatin, and hydrochlorothiazide. Ten out of 11 compounds were detected and quantified in 13 sampling points. Caffeine was detected with the highest level, with concentrations reaching up to 9099 ng/L in STP influent.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods*
Simple high-performance liquid chromatographic method for the determination of tocotrienols in human plasma

Yap SP, Julianto T, Wong JW, Yuen KH

J Chromatogr B Biomed Sci Appl, 1999 Dec 10;735(2):279-83.
PMID: 10670741

A simple high-performance liquid chromatographic method using fluorescence detection was developed for the determination of vitamin E especially delta-, gamma- and alpha-tocotrienols in human plasma. The method entailed direct injection of plasma sample after deproteinization using a 3:2 mixture of acetonitrile-tetrahydrofuran. The mobile phase comprised 0.5% (v/v) of distilled water in methanol. Analyses were run at a flow-rate of 1.5 ml/min with the detector operating at an excitation wavelength of 296 nm and emission wavelength of 330 nm. This method is specific and sensitive, with a quantification limit of approximately 40, 34 and 16 ng/ml for alpha-, gamma- and delta-tocotrienol, respectively. The mean absolute recovery values were about 98% while the within-day and between-day relative standard deviation and percent error values of the assay method were all less than 12.0% for alpha-, gamma- and delta-tocotrienol. The calibration curve was linear over a concentration range of 40-2500, 30-4000 and 16-1000 ng/ml for alpha-, gamma- and delta-tocotrienol, respectively. Application of the method in a bioavailability study for determination of the above compounds was also demonstrated.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods*
Simple high-performance liquid chromatographic method for the determination of ranitidine in human plasma

Wong CF, Peh KK, Yuen KH

J Chromatogr B Biomed Sci Appl, 1998 Oct 23;718(1):205-10.
PMID: 9832378

A simple high-performance liquid chromatographic method was developed for the determination of ranitidine in human plasma. Prior to analysis, ranitidine and the internal standard (metoprolol) were extracted from alkalinized plasma samples using dichloromethane. The mobile phase was 0.05 M potassium dihydrogenphosphate-acetonitrile (88:12, v/v) adjusted to pH 6.5. Analysis was run at a flow-rate of 1.3 ml/min and at a detection wavelength of 229 nm. The method is sensitive with a detection limit of 1 ng/ml at a signal-to-noise ratio of 3:1, while the quantification limit was set at 15 ng/ml. The calibration curve was linear over a concentration range of 15-2000 ng/ml. Mean recovery value of the extraction procedure was about 90%, while the within-day and between-day coefficients of variation and percent error values of the assay method were all less than 15%.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods*
Determination of flavonoids from Orthosiphon stamineus in plasma using a simple HPLC method with ultraviolet detection

Loon YH, Wong JW, Yap SP, Yuen KH

J Chromatogr B Analyt Technol Biomed Life Sci, 2005 Feb 25;816(1-2):161-6.
PMID: 15664346

A simple liquid chromatographic method was developed for the simultaneous determination of flavonoids from Orthosiphon stamineus Benth, namely sinensitin, eupatorin and 3'-hydroxy-5,6,7,4'-tetramethoxyflavone, in plasma. Prior to analysis, the flavonoids and the internal standard (naproxen) were extracted from plasma samples using a 1:1 mixture of ethyl acetate and chloroform. The detection and quantification limits for the three flavonoids were similar being 3 and 5 ng/ml, respectively. The within-day and between-day accuracy values, expressed as percentage of true values, for the three flavonoids were between 95 and 107%, while the corresponding precision, expressed as coefficients of variation, for the three flavonoids were less than 14%. In addition, the mean recovery values of the extraction procedure for all the flavonoids were between 92 and 114%. The calibration curves were linear over a concentration range of 5-4000 ng/ml. The present method was applied to analyse plasma samples obtained from a pilot study using rats in which the mean absolute oral bioavailability values for sinensitin, eupatorin and 3'-hydroxy-5,6,7,4'-tetramethoxyflavone was 9.4, 1.0 and 1.5%, respectively.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods
Liquid chromatographic method for the determination of plasma itraconazole and its hydroxy metabolite in pharmacokinetic/bioavailability studies

Wong JW, Nisar UR, Yuen KH

J Chromatogr B Analyt Technol Biomed Life Sci, 2003 Dec 25;798(2):355-60.
PMID: 14643517

A sensitive and selective high-performance liquid chromatographic method was developed for the determination of itraconazole and its active metabolite, hydroxyitraconazole, in human plasma. Prior to analysis, both compounds together with the internal standard were extracted from alkalinized plasma samples using a 3:2 (v/v) mixture of 2,2,4-trimethylpentane and dichloromethane. The mobile phase comprised 0.02 M potassium dihydrogen phosphate-acetonitrile (1:1, v/v) adjusted to pH 3.0. Analysis was run at flow-rate of 0.9 ml/min with excitation and emission wavelengths set at 260 and 365 nm, respectively. Itraconazole was found to adsorb on glass or plastic tubes, but could be circumvented by prior treating the tubes using 10% dichlorodimethylsilane in toluene. Moreover, rinsing the injector port with acetonitrile helped to overcome any carry-over effect. This problem was not encountered with hydroxyitraconazole. The method was sensitive with limit of quantification of 3 ng/ml for itraconazole and 6 ng/ml for hydroxyitraconazole. The calibration curve was linear over a concentration range of 2.8-720 ng/ml for itraconazole and 5.6-720 ng/ml for the hydroxy metabolite. Mean recovery value of the extraction procedure for both compounds was about 85%, while the within-day and between-day coefficient of variation and percent error values of the assay method were all less than 15%. Hence, the method is suitable for use in pharmacokinetic and bioavailability studies of itraconazole.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods*
Determination of plasma testosterone using a simple liquid chromatographic method

Ng BH, Yuen KH

J Chromatogr B Analyt Technol Biomed Life Sci, 2003 Aug 15;793(2):421-6.
PMID: 12906917

A simple and sensitive high-performance liquid chromatographic (HPLC) method using ultraviolet detection was developed for the determination of testosterone in human plasma. Testosterone and the internal standard, griseofulvin, were extracted from 0.50 ml plasma sample using a mixture of dichloromethane-2,2,4-trimethylpentane (3:2, v/v). The mobile phase, consisted of 0.02 M sodium dihydrogenphosphate-acetonitrile-methanol (51:47:2, v/v) adjusted to pH 3.1 and delivered to a C(18) analytical column (150 x 4.6 mm I.D., 4 microm particles) at a flow-rate of 1 ml/min while the detection wavelength was set at 240 nm with a sensitivity range of 0.005 a.u.f.s. The method has a quantification limit of 1.6 ng/ml. Recoveries of testosterone were all greater than 92% over the linear concentration range of 1.6-400 ng/ml while that of griseofulvin was approximately 95%. The within- and between-day RSD values were all less than 8% while the accuracy values ranged from 96.0 to 106.0% over the concentration range studied. The method was applied to the analysis of early morning plasma testosterone levels of 12 healthy human male volunteers. The levels were found to range from 3.1 to 8.4 ng/ml, within the normal range reported in the literature.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods*
Evaluation and application of a simple and rapid method for the analysis of aflatoxins in commercial foods from Malaysia and the Philippines

Ali N, Hashim NH, Yoshizawa T

Food Addit Contam, 1999 Jul;16(7):273-80.
PMID: 10656052

For application to the analysis of aflatoxins (AF) in commercial peanut and corn products, the ISOLUTE multimode column (IMC, solid phase multifunctional column) method was validated by comparing with the modified Florisil column (MFC) method. Twenty-two peanut and eight corn products from Malaysia and the Philippines were analysed for AFB1, AFB2, AFG1 and AFG2 firstly by the MFC method and then by the IMC method. For peanut products, 14 out of 22 samples were positive by the two methods in the range of 1-378 micrograms/kg of AF, and correlation coefficients (r) for AFB1 and AFB2 were 0.987 and 0.997, respectively. For corn and corn products, all the samples were positive in the range of 1-130 micrograms/kg, and r values were 0.992 and 0.805 for AFB1 and AFB2 respectively. Thus, the results were significantly (p < 0.01) in close agreement, particularly for lower range of 1-50 micrograms/kg of AF concentrations in all the samples. For the occurrence of AF, 11 (65%) of peanut products from Malaysia were contaminated with AF at a mean level of 50 micrograms/kg (maximum 180 micrograms/kg) and two (40% products from the Philippines were contaminated with as high as 375 micrograms/kg and 177 micrograms/kg of AF, respectively. All the corn products from the Philippines were contaminated with AF at a mean level of 44 micrograms/kg (maximum 130 micrograms/kg). Contamination of commercial foods with high levels of AF is a very important issue to both the countries since these foods are very popular among children.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods
Evaluation of a method to determine the natural occurrence of aflatoxins in commercial traditional herbal medicines from Malaysia and Indonesia

Ali N, Hashim NH, Saad B, Safan K, Nakajima M, Yoshizawa T

Food Chem Toxicol, 2005 Dec;43(12):1763-72.
PMID: 16019122

Traditional herbal medicines, popularly known as 'jamu' and 'makjun' in Malaysia and Indonesia, are consumed regularly to promote health. In consideration of their frequent and prolonged consumption, the natural occurrence of aflatoxins (AF) in these products was determined using immunoaffinity column clean-up and high-performance liquid chromatography with pre-column derivatization. The evaluated method, which entails dilution of sample extracts with Tween 20-phosphate buffered saline (1:9, v/v) and a chromatographic system using isocratic mobile phase composed of water-methanol-acetonitrile (70:20:10, v/v/v), was effective in separating AFB1, AFG1 and AFG2 from interference at their retention times. Results were confirmed using post-column derivatization with photochemical reactor. For 23 commercial samples analyzed, mean levels (incidence) of AFB(1), AFB(2) and AFG1 in positive samples were 0.26 (70%), 0.07 (61%) and 0.10 (30%) microg/kg, respectively; one sample was positive for AFG2 at a level of 0.03 (4%) microg/kg. In contrast to the high levels of AF in crude herbal drugs and medicinal plants reported previously by other researchers, the low contamination levels reported in this study may be attributed to the higher selectivity to AF of the method applied. Based on the AFB1 levels and the daily consumption of positive samples, a mean probable daily intake of 0.022 ng/kg body weight was calculated.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods
Improved high performance liquid chromatographic analysis of omeprazole in human plasma

Yuen KH, Choy WP, Tan HY, Wong JW, Yap SP

J Pharm Biomed Anal, 2001 Feb;24(4):715-9.
PMID: 11272330

A simple high-performance liquid chromatographic method was developed for the determination of omeprazole in human plasma. Omeprazole and the internal standard, chloramphenicol, were extracted from alkalinized plasma samples using dichloromethane. The mobile phase was 0.05 M Na2HPO4-ACN (65:35, v/v) adjusted to pH 6.5. Analysis was run at a flow rate of 1.0 ml/min at a detection wavelength of 302 nm. The method was specific and sensitive with a detection limit of 2.5 ng/ml at a signal-to-noise ratio of 4:1. The limit of quantification was set at 5 ng/ml. The calibration curve was linear over a concentration range of 5-1280 ng/ml. Mean recovery value of the extraction procedure was about 96%, while the within and between day coefficient of variation and percent error values of the assay method were all less than 14%.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods*
Porous graphitic carbon shows promise for the rapid screening partial DPD deficiency in lymphocyte dihydropyrimidine dehydrogenase in Chinese, Indian and Malay in Singapore by using semi-automated HPLC-radioassay

Liem LK, Choong LH, Woo KT

Clin Biochem, 2002 May;35(3):181-7.
PMID: 12074825

OBJECTIVE: Dihydropyrimidine dehydrogenase (DPD) catalyzes the degradation of thymine, uracil, and the chemotherapeutic drug 5-Fluorouracil. In general reverse-phase high pressure liquid chromatography is the standard method for separating 5-[2-(14)C]Fluorouracil and 5-[2-(14)C]Fluoro-5,6-dihydrouracil. However, the use of 100% aqueous solution (as HPLC mobile phase) may collapse the C-18 bonded phase and result in a retention time shift. The aim of this study is to develop a rapid, reproducible, sensitive method for screening partial DPD deficiency in healthy volunteers.
DESIGN AND METHODS: The activity of DPD was measured using 5-[2- (14)C]Fluorouracil (5-[2-(14)C]FUra) followed by separation of substrate and product 5-[2-(14)C]FUraH(2) with a 15 x 4.6 mm I.D., 5 microm particle size (d(p)) porous graphitic carbon (PGC) column (Hypercarb(R)) and HPLC with online detection of the radioactivity. This was standardized using the protein concentration of the cytosol (NanoOrange(R) Protein Quantitation).
RESULTS: Complete baseline separation of 5-[2-(14)C]Fluorouracil (5-[2-(14)C]FUra) and 5-[2-(14)C]Fluoro-5,6-dihydrouracil (5-[2-(14)C]FUraH(2)) was achieved using a porous graphitic carbon (PGC) column. The detection limit for 5-[2-(14)C]FUraH(2) was 0.4 pmol.
CONCLUSIONS: By using linear gradient separation (0.1% Trifluoroacetic acid [TFA] in water to 100% Methanol) protocols in concert with PGC columns (Hypercarb(R)), we have demonstrated that a PGC column has a distinct advantage over C-18 reverse phase columns in terms of column stability (pH 1-14). This method provides an improvement on the specific assay for DPD enzyme activity. It is rapid, reproducible and sensitive and can be used for routine screening for healthy and cancer patients for partial and profound DPD deficiency before treatment with 5- FUra.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods*
Determination of biogenic amines in chicken, beef, and mutton by dansyl chloride microwave derivatization in Malaysia

Yue CS, Lim AK, Chia ML, Wong PY, Chin JSR, Wong WH

J Food Sci, 2023 Feb;88(2):650-665.
PMID: 36624628 DOI: 10.1111/1750-3841.16404

In this study, an improved dansyl-chloride derivatization technique using a microwave synthesizer was used for the qualitative and quantitative analyses of biogenic amine in the fresh meat samples. The derivatization technique was optimized in terms of temperature, reaction time, and spinning speed. The derivatization method together with a validated reversed-phase HPLC-DAD method was used for the determination of biogenic amines in chicken, beef, and mutton sold in the wet market. The results of the analyses showed that tryptamine, putrescine, and histamine were generally detected in all the three types of meat. Higher levels of histamine were found in chicken and beef. However, low levels of histamine were observed in mutton. Tyramine was either detected low or moderate in all the three types of meat. The biogenic amines of the fresh meat sold in the wet market is generally higher than the reported values. The mechanisms of biogenic amines-dansyl-chloride formation were investigated and proposed. PRACTICAL APPLICATION: The biogenic amine derivatization method was improved. The improved derivatization method can be potentially used for various food products beside meats for routine biogenic amine analyses due to its fast analysis time and simplicity. High levels of biogenic amines were generally found in the meat sold in the wet markets. However, proper handling of the raw meat can reduce the risk of infection.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods
LC-PDA-MS/MS-Based Characterization of Key Phytochemicals in Eurycoma Longifolia Roots

Chua LS, Segaran A, Wong HJ

J Chromatogr Sci, 2021 Jun 21;59(7):659-669.
PMID: 33876232 DOI: 10.1093/chromsci/bmab041

The objective of the study was to fractionate the crude extract of Eurycoma longifolia (E. longifolia) roots and identify the intense peaks using HPLC-PDA-MS/MS, UPLC-MS/MS and H-NMR. Column chromatography was used to fractionate the crude extract into individual fractions using six solvent systems ranged from ethyl acetate, methanol and water in increasing polarity. Two fractions with nearly pure and intense peaks were selected for compound identification. Chromenone (coumarin) and chromone derivatives were putatively identified, besides several previously reported quassinoid glycosides (eurycomanone derived glycoside, 2,3-dehydro-4α-hydroxylongilactone glucoside, eurycomanol glycoside and eurycomanol trimer) in the fraction 11 of 100% methanol. A newly reported compound, namely hydroxyl glyyunanprosapogenin D (838 g/mol) was proposed to be the compound detected in the fraction 11 of 50% ethyl acetate and 50% methanol. This is also the first study to report the identification of chromenones and chromones in E. longifolia extract.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods*
Validation of LC for the determination of alpha-mangostin in mangosteen peel extract: a tool for quality assessment of Garcinia mangostana L

Yodhnu S, Sirikatitham A, Wattanapiromsakul C

J Chromatogr Sci, 2009 Mar;47(3):185-9.
PMID: 19298703

Mangosteen, Garcinia mangostana L., is known as the "Queen of fruits" and can be cultivated in the tropical rainforest such as Malaysia, Indonesia, and Thailand. Compounds isolated from the fruit peel of mangosteen contain abundant xanthones (especially alpha-mangostin). It has been used as traditional medicine such as anti-inflammatory and antibacterial and is popularly applied to cosmetic and pharmaceutical products. However, there is little information for quality and quantity determination of alpha-mangostin in mangosteen. Thus, the aim of this study was to set up a validated and stability-indicated isocratic reverse-phase high-performance liquid chromatographic (HPLC) method for quality control and quantity determination of a-mangostin from mangosteen peel extract. The assay was fully validated and shown to be linear (r(2) > 0.999), sensitive (LOD = 0.02 microg/mL and LOQ = 0.08 microg/mL), accurate (intra-day was between 98.1-100.8%, inter-day was between 90.0-101.3%), precise (intra-day variation < or = 1.8%, inter-day variation < or = 4.3%), specific, and with good recovery. Total analysis was approximately 8 min. The finalized method is also a stability-indicating assay. The present method should be useful for analytical research and for routine quality control analysis of alpha-mangostin in mangosteen peel extract and products of mangosteen.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods*
Method development and validation of a high-performance liquid chromatographic method for tramadol in human plasma using liquid-liquid extraction

Gan SH, Ismail R, Wan Adnan WA, Wan Z

J Chromatogr B Analyt Technol Biomed Life Sci, 2002 May 25;772(1):123-9.
PMID: 12016023

An HPLC system using a simple liquid-liquid extraction and HPLC with UV detection has been validated to determine tramadol concentration in human plasma. The method developed was selective and linear for concentrations ranging from 10 to 2000 ng/ml with average recovery of 98.63%. The limit of quantitation (LOQ) was 10 ng/ml and the percentage recovery of the internal standard phenacetin was 76.51%. The intra-day accuracy ranged from 87.55 to 105.99% and the inter-day accuracy, 93.44 to 98.43% for tramadol. Good precision (5.32 and 6.67% for intra- and inter-day, respectively) was obtained at LOQ. The method has been applied to determine tramadol concentrations in human plasma samples for a pharmacokinetic study.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods*
Ionic liquid-impregnated agarose film two-phase micro-electrodriven membrane extraction (IL-AF-μ-EME) for the analysis of antidepressants in water samples

Mohamad Hanapi NS, Sanagi MM, Ismail AK, Wan Ibrahim WA, Saim N, Wan Ibrahim WN

J Chromatogr B Analyt Technol Biomed Life Sci, 2017 Mar 01;1046:73-80.
PMID: 28142101 DOI: 10.1016/j.jchromb.2017.01.028

The aim of this study was to investigate and apply supported ionic liquid membrane (SILM) in two-phase micro-electrodriven membrane extraction combined with high performance liquid chromatography-ultraviolet detection (HPLC-UV) for pre-concentration and determination of three selected antidepressant drugs in water samples. A thin agarose film impregnated with 1-hexyl-3-methylimidazolium hexafluorophosphate, [C6MIM] [PF6], was prepared and used as supported ionic liquid membrane between aqueous sample solution and acceptor phase for extraction of imipramine, amitriptyline and chlorpromazine. Under the optimized extraction conditions, the method provided good linearity in the range of 1.0-1000μgL(-1), good coefficients of determination (r(2)=0.9974-0.9992) and low limits of detection (0.1-0.4μgL(-1)). The method showed high enrichment factors in the range of 110-150 and high relative recoveries in the range of 88.2-111.4% and 90.9-107.0%, for river water and tap water samples, respectively with RSDs of ≤7.6 (n=3). This method was successfully applied to the determination of the drugs in river and tap water samples. It is envisaged that the SILM improved the perm-selectivity by providing a pathway for targeted analytes which resulted in rapid extraction with high degree of selectivity and high enrichment factor.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods
Agarose-chitosan-C18 film micro-solid phase extraction combined with high performance liquid chromatography for the determination of phenanthrene and pyrene in chrysanthemum tea samples

Ng NT, Sanagi MM, Wan Ibrahim WN, Wan Ibrahim WA

Food Chem, 2017 May 01;222:28-34.
PMID: 28041555 DOI: 10.1016/j.foodchem.2016.11.147

Agarose-chitosan-immobilized octadecylsilyl-silica (C18) film micro-solid phase extraction (μSPE) was developed and applied for the determination of phenanthrene (PHE) and pyrene (PYR) in chrysanthemum tea samples using high performance liquid chromatography-ultraviolet detection (HPLC-UV). The film of blended agarose and chitosan allows good dispersion of C18, prevents the leaching of C18 during application and enhances the film mechanical stability. Important μSPE parameters were optimized including amount of sorbent loading, extraction time, desorption solvent and desorption time. The matrix match calibration curves showed good linearity (r⩾0.994) over a concentration range of 1-500ppb. Under the optimized conditions, the proposed method showed good limits of detection (0.549-0.673ppb), good analyte recoveries (100.8-105.99%) and good reproducibilities (RSDs⩽13.53%, n=3) with preconcentration factors of 4 and 72 for PHE and PYR, respectively.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods*
Fulltext Assessing product adulteration of Eurycoma longifolia (Tongkat Ali) herbal medicinal product using DNA barcoding and HPLC analysis

Abubakar BM, Salleh FM, Shamsir Omar MS, Wagiran A

Pharm Biol, 2018 Dec;56(1):368-377.
PMID: 30058427 DOI: 10.1080/13880209.2018.1479869

CONTEXT: Eurycoma longifolia Jack (Simaroubaceae) commonly known as Tongkat Ali is one of the most important plants in Malaysia. The plant extracts (particularly roots) are widely used for the treatment of cough and fever besides having antimalarial, antidiabetic, anticancer and aphrodisiac activities.
OBJECTIVES: This study assesses the extent of adulteration of E. longifolia herbal medicinal products (HMPs) using DNA barcoding validated by HPLC analysis.
MATERIALS AND METHODS: Chloroplastic rbcL and nuclear ITS2 barcode regions were used in the present study. The sequences generated from E. longifolia HMPs were compared to sequences in the GenBank using MEGABLAST to verify their taxonomic identity. These results were verified by neighbor-joining tree analysis in which branches of unknown specimen are compared to the reference sequences established from this study and other retrieved from the GenBank. The HMPs were also analysed using HPLC analysis for the presence of eurycomanone bioactive marker.
RESULTS: Identification using DNA barcoding revealed that 37% of the tested HMPs were authentic while 27% were adulterated with the ITS2 barcode region proven to be the ideal marker. The validation of the authenticity using HPLC analysis showed a situation in which a species which was identified as authentic was found not to contain the expected chemical compound.
DISCUSSION AND CONCLUSIONS: DNA barcoding should be used as the first screening step for testing of HMPs raw materials. However, integration of DNA barcoding with HPLC analysis will help to provide detailed knowledge about the safety and efficacy of the HMPs.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods
An RP-HPLC-UV method with SPE for cefotaxime in all-in-one total parenteral nutritional admixtures: application to stability studies

Iqbal MS, Bahari MB, Darwis Y, Iqbal MZ, Hayat A, Venkatesh G

J AOAC Int, 2013 6 19;96(2):290-4.
PMID: 23767352

A simple and selective RP-HPLC-UV method with SPE was developed and validated for the quantification of cefotaxime in all-in-one total parenteral nutrition (AIO-TPN) admixtures. Chromatographic separation was achieved on a 5 pm particle size C18 DB column (250 x 4.6 mm id) using the mobile phase ammonium acetate (25 mM, pH 4.0)-50% acetonitrile in methanol (80 + 20, v/v). The flow rate was 0.9 mL/min and the detection wavelength was 254 nm. The analyte was extracted from AIO-TPN admixtures by means of an SPE method. The cefotaxime calibration curve was linear over a concentration range of 100-1400 microg/mL with a correlation coefficient of > or = 0.9994. The intraday accuracy and precision for cefotaxime were < or = -3.15 and < or = 3.08%, respectively, whereas the interday accuracy and precision were < or = -2.48 and < or = 2.25%, respectively. The method was successfully applied to stability studies of cefotaxime in the presence of micronutrients together with low and high concentrations of macronutrients in AIO-TPN admixtures. Cefotaxime was degraded by 13.00 and 26.05% at room temperature (25 +/- 2 degrees C) after 72 h in low and high macronutrient concentration formulations of AIO-TPN admixtures, respectively. The values of cefotaxime degradation rates for low and high macronutrient concentration formulations of AIO-TPN admixtures were -0.164 and -0.353, respectively. These results indicated that there was a higher rate of degradation in the AIO-TPN admixture formulations containing high concentrations of macronutrients.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods*
High performance liquid chromatographic method optimized by Box-Behnken design model to determine caffeine in pharmaceutical preparations and urine samples

Najmul Hejaz Azmi S, Aqib Nasir Al Rawahi W, Ibrahim Al Yahyai A, Ali Al Qasimi A, Saif Al Fuliti K, Said Al Qalhati O, et al.

J Chromatogr B Analyt Technol Biomed Life Sci, 2024 Feb 15;1234:124035.
PMID: 38309045 DOI: 10.1016/j.jchromb.2024.124035

A UV-HPLC method optimized by Box-Behnken design model was developed to determine caffeine in pharmaceutical preparations and urine samples. The chromatographic conditions followed were mobile phase: methanol/water/ citrate buffer (pH 4.6) (40:25:35, v/v/v),AcclaimTMDionex C18 column (ODS 100A˚, 5 µm; 4.6 × 250 mm),flow rate (0.9 mL min-1), column temperature (30 °C) and UV-detection wavelength (204 nm). The chromatographic variables: pH (A), % methanol fraction (B), flow rate(C) and column temperature (D) were optimized at 50 μg mL-1caffeine using BBD model. The chromatogram resulted in the asymmetry factor (1.23), theoretical plate 13,786 and retention time (5.79 min). The proposed HPLC method's greenness point was assessed byAnalytical Eco-scale and found to be 78 (as per guidelines, ranked as excellent). The linearity was ranged from2.0 to 70 µg mL-1 with coefficient of correlation (r = 0.999) and detection limit of 0.19 µg mL-1. The proposedmethod was developed successfully and applied for the assay of active caffeine in pharmaceutical preparations and urine samples. The % recovery obtained by both (proposed and reference) methods ranged from 99.98 to 100.05 % followed the compliance (100 ± 2 %) with Canadian Health Protection regulatory guidelines. The performance of the proposed method was compared with published papers and found to be acceptable and superior. The proposed method was quite effective as the reference method, and hence can be used as an alternative method for the assay of active caffeine in pharmaceutical preparations and urine samples.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods
Phytochemical quantification and HPLC analysis of Parkia speciosa pod extract

Cheong NDH, Mohamed E, Haron N, Camalxaman SN, Abdullah A, Mohamad Yusof MI, et al.

Med J Malaysia, 2024 Mar;79(Suppl 1):34-39.
PMID: 38555883

INTRODUCTION: Parkia speciosa Hassk., commonly known as bitter bean or twisted cluster bean, is a tropical leguminous plant species native to Southeast Asia. The plant's edible pods have been traditionally used in various cuisines, particularly in Malaysian, Thai, and Indonesian cooking. Apart from being used as a food ingredient, the pods of P. speciosa also have a range of potential applications in other fields, including medicine, agriculture, and industry. The pods are said to have several phytochemicals that hold great therapeutic values such as reducing inflammation, improving digestion, and lowering blood sugar levels. However, there is limited information on the specific phytochemical contents of the pods in the literature. Thus, the aim of this study is to quantify the total phenolic and flavonoid compounds and to determine the concentrations of four selected phytochemical compounds in the P. speciosa pod extract (PSPE).
MATERIALS AND METHODS: Quantification of the total phenolic (TPC) and flavonoid contents (TFC) in PSPE were done via colourimetric methods; and the determination of the concentrations of four specific phytochemicals (gallic acid, caffeic acid, rutin, and quercetin) were done via High- Performance Liquid Chromatography (HPLC).
RESULTS: Colourimetric determination of PSPE showed TPC and TFC values of 84.53±9.40 mg GAE/g and 11.96±4.51 mg QE/g, respectively. Additional analysis of the phytochemicals using HPLC revealed that there were 6.45±3.36 g/kg, 5.91±1.07 g/kg, 0.39±0.84 g/kg, and 0.19±0.47 g/kg of caffeic acid, gallic acid, rutin, and quercetin, respectively.
CONCLUSION: The findings show that PSPE contains substantial amounts of caffeic acid, gallic acid, rutin, and quercetin, which may indicate its potential as antibacterial, anti-inflammatory, anti-lipid, and antiviral medicines.

Matched MeSH terms: Chromatography, High Pressure Liquid/methods

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