High-performance Liquid Chromatography With Photodiode Array Detection Research Articles

Rapid Determination of Costunolide and Dehydrocostuslactone in Human Plasma Sample and Chinese Patent Medicine Xiang Sha Yang Wei Capsule Using HPLC‐DAD Coupled with Second‐order Calibration

AbstractA novel methodology that combines high performance liquid chromatography with photodiode‐array detector (HPLC‐DAD) coupled with second‐order calibration method based on alternating trilinear decomposition (ATLD) algorithm was used in determination of the effective constituents such as costunolide and dehydrocostuslactone, in plasma sample and Chinese patent medicine Xiang Sha Yang Wei (XSYW) capsule. Complicated systems such as plasma and Chinese patent medicine which have intricate components are tedious to isolate and purify. The problem that chromatographic peaks are heavily overlapped among the analytes and interferents from the background matrices can be resolved, and the satisfactory quantification results have been gained with the help of the ATLD algorithm which utilized "mathematical separation" instead of partial "physical or chemical separation". Meanwhile, HPLC‐MS/MS method was used to validate the accuracy of the proposed determination method.

Determination of Chlorantraniliprole Residues in Grape by High-Performance Liquid Chromatography

An effective analytical method for the residue analysis of a novel insecticide chlorantraniliprole and its dissipation in grape were studied. Chlorantraniliprole residues were extracted from grape samples with ethyl acetate. The extract was cleaned up with QuEChERS (Quick, Easy, Cheap, Effective, Rugged and Safe) method and determined by high-performance liquid chromatography with photodiode array detector (HPLC-DAD). At fortification levels of 0.06, 0.5, and 1.0 mg kg−1 in grape, it was shown that recoveries ranged from 95.11 to 102 % with relative standard deviation (RSD) of 6 to 11 %. The limit of detection (LOD) and limit of quantification (LOQ) were found to be 0.02 and 0.06 mg kg−1, respectively. The dissipation half-life time of chlorantraniliprole residues in grape was 2.70 days. According to maximum residue limit (MRL), the preharvest interval (PHI) of chlorantraniliprole on grape was 4 days after the treatment. Based on the results of this study and the relevant residue regulation, chlorantraniliprole residue levels will be acceptable when applied to grape in Egypt.

Three-phase hollow fiber microextraction based on two immiscible organic solvents for determination of tricyclic antidepressant drugs: Comparison with conventional three-phase hollow fiber microextraction

The aim of this research was to compare the extraction efficiencies of two modes of three-phase hollow fiber microextraction (HF-LLLME) based on aqueous and organic acceptor phases for analysis of tricyclic antidepressant (TCA) drugs. High-performance liquid chromatography with photodiode array detection (HPLC-DAD) was applied for determination of the drugs. In order to examine the ability of the new concept of HF-LLLME based on organic acceptor solvent in comparison with aqueous acceptor phase to extract the analytes, four TCAs were selected. The effect of different extraction conditions (i.e., type of acceptor phase, hollow fiber length, ionic strength, stirring rate, and extraction time) on the extraction efficiency of the TCAs was investigated and optimized using central composite design (CCD) as a powerful tool. Both methods were characterized by good linearity and high repeatability, but HF-LLLME with organic acceptor provided higher extraction efficiency and thus lower limits of detection (LODs). Calibration curves were linear ( r 2 > 0.996) in the range of 0.2–200 μg L −1. LODs for all the TCAs ranged from 0.08 to 0.2 μg L −1 using HPLC-DAD. Also an improvement in sensitivity of several orders of magnitude was achieved using single-ion monitoring GC–MS analyses (0.04 μg L −1) due to compatibility of this technique with GC instrument. The applicability of the proposed HF-LLLME/GC–MS and HPLC-DAD methods was demonstrated by analyzing the drugs in spiked urine and plasma samples. The obtained recoveries of the drugs in the range of 87.9–109.2% indicated the excellent capability of the developed method for extraction of TCAs from complex matrices.

Chromatographic Fingerprint and Quantitative Analysis of Seven Bioactive Compounds ofScutellaria barbata

Scutellaria barbata D. Don is widely used as a folk antitumor and anti-inflammatory agent in Asia. However, a simple and global quality control method for S. barbata was lacking. In this study, six phenolic compounds, including P-coumaric acid, scutellarin, apigenin 5-O-β-glucopyranoside, luteolin, apigenin and 4'-hydroxywogonin were obtained from S. barbata by phytochemical investigations. The six compounds plus baicalein show cytotoxcities to the nine human cancer cells, K562, MGC-803, HL60, SH-SY5Y, SW1116, SMMC-7221, SW480, HepG2 and KB. Subsequently, a high-performance liquid chromatography with photodiode array detector (HPLC-DAD) was developed for both fingerprint analysis of S. barbata and quantitative determination of the seven anticancer active compounds in S. barbata. The chromatographic separation was accomplished on an Ultimate™ XB-C18 column (4.6 x 250 mm, 5 µm) in 65 min. For fingerprinting, 26 common peaks were found and selected as characteristic peaks to assess the consistency of S. barbata samples. For quantitative analysis, the seven bioactive compounds showed good regression relationship ( R² > 0.999) within test ranges and the recovery of the method was in the range of 90-105 %. In brief, the present study provides the fingerprint analysis and quantitative methods for global and systematical quality control of S. barbata for its anticancer usage.

Regulation of p21, MMP-1, and MDR-1 Expression in Human Colon Carcinoma HT29 Cells by Tian Xian Liquid, a Chinese Medicinal Formula, In Vitro and In Vivo

Tian-Xian liquid (TXL), a commercially available Chinese medicine decoction, has been used as an anticancer dietary agent for more than 10 years without reported side effects. The safety and quality consistency of TXL and its mechanisms of action on antiproliferation, antimetastasis, and reversion of multidrug resistance (MDR) regimens were explored. In this study, an atomic absorption spectrophotometer and reversed phase high performance liquid chromatography with photodiode array detection (HPLC-DAD) were used to evaluate the main toxic elements and the quality consistency among different batches of TXL extracts, respectively. HT29 human colon cancer cell line and tumor-bearing nude mice were used. TXL was provided by China-Japan Feida Union Company Limited. The effect of TXL on in vitro proliferation of HT29 human colon cancer cell line was examined. The percentages of treated cells distributed in different phases of the cell cycles were analyzed by flow cytometry. Antiproliferative effect after treatment with TXL was assessed by determination of the protein levels of p21, cyclinD1, PCNA, and cdk-2, which are the key regulators for cell cycle progression. Meanwhile, the protein levels of MMP-1 and MDR-1 (multidrug resistance protein-1) were also determined to assess the effect of TXL on antimetastasis and reversion of MDR regimen, respectively. The contents of main toxic elements were lower in TXL extract compared with the standard set by the Department of Health of the Government of Hong Kong Special Administrative Region (SAR). Our HPLC results showed that the relative standard deviations of the amount of the 5 standards were less than 5% in different batches of TXL. Immunoblotting analysis revealed a dramatic induction of cyclin kinase inhibitor p21 as well as an inhibition of cyclinD1, PCNA, and cdk-2 in the TXL-treated in vitro models, thereby, impeding cell progression from G1/S phase. Results obtained from the in vivo study also demonstrated that TXL upregulated the protein level of p21 and downregulated the protein levels of MMP-1 and MDR-1. Results obtained from the present investigation not only demonstrate the safety and quality of TXL extract but also demonstrate that TXL possesses antiproliferative and antimetastatic activities and brings about reversion of MDR on HT29 cell and on xenografted tissue in tumor-implanted nude mice.

Simultaneous determination of psoralen and isopsoralen in plasma and Chinese medicine Xian Ling Gu Bao capsule by using HPLC-DAD coupled with alternating trilinear decomposition algorithm

A method using high performance liquid chromatography with photodiode-array detection (HPLC-DAD) coupled with alternating trilinear decomposition (ATLD) algorithm was proposed for simultaneous determination of psoralen and isopsoralen in plasma and Chinese medicine “Xian Ling Gu Bao” capsule (XLGBC). In this paper, the application of ATLD algorithm into traditional chromatographic method can handle this problem that the chromatographic and spectral peaks are heavily overlapped among the analytes and even between the analytes and interferences from the background matrices. A simple improvement of chromatographic condition like mobile phase is not enough to realize effective separation for the two isomeric compounds, especially in the presence of interferences. However, the ATLD algorithm utilized “mathematical separation” instead of partial “physical or chemical separation” to directly determine the spectral profiles of the analytes of interests in complex system. The satisfactory quantification results have been gained with simple mobile phase. In the analysis of real Chinese medicine samples, the accuracy of the concentrations which were obtained by ATLD was also validated by HPLC–MS method.

HPLC-DAD characterisation of phenolic compounds from Andean oca ( Oxalis tuberosa Mol.) tubers and their contribution to the antioxidant capacity

A qualitative and quantitative characterisation of the main non-anthocyanin phenolic compounds from two different colored oca ( Oxalis tuberosa Mol.) genotypes with potential antioxidant capacity was carried out by high performance liquid chromatography with photodiode array detection (HPLC-DAD). Phenolic compounds were fractionated in two main fractions: an aqueous (Faq) and a ethyl acetate fraction (Fea). In addition, the contribution of these phenolic fractions to the antioxidant capacity was evaluated. The Faq revealed the presence of caffeic, vanillic and cinnamic acid derivatives, flavan-3-ols and flavones derivatives, as the main non-anthocyanin phenolic compounds for both genotypes. Anthocyanins for the purple genotype were significantly present in this fraction. Acid hydrolysis revealed the presence of vanillic, caffeic and cinnamic acids and malvidin in Faq. The Fea was composed mainly of caffeic and cinnamic acid derivatives as well as flavan-3-ols, flavones and flavanone derivatives. Based on their UV–Vis spectral data the flavan-3-ols, flavones and flavanones detected in both fractions seem to correspond to bound forms of catechin, luteolin and apigenin and naringenin, respectively. The Faq fractions were the major contributors to the ABTS antioxidant capacity (77–82%). The results obtained in the present study suggest that oca tubers could potentially be considered beneficial for human health and for potential industrial applications.

Beer constituents inhibit prostate cancer cells proliferation

A qualitative and quantitative characterisation of the main non-anthocyanin phenolic compounds from two different colored oca (Oxalis tuberosa Mol.) genotypes with potential antioxidant capacity was carried out by high performance liquid chromatography with photodiode array detection (HPLC-DAD). Phenolic compounds were fractionated in two main fractions: an aqueous (Faq) and a ethyl acetate fraction (Fea). In addition, the contribution of these phenolic fractions to the antioxidant capacity was evaluated. The Faq revealed the presence of caffeic, vanillic and cinnamic acid derivatives, flavan-3-ols and flavones derivatives, as the main non-anthocyanin phenolic compounds for both genotypes. Anthocyanins for the purple genotype were significantly present in this fraction. Acid hydrolysis revealed the presence of vanillic, caffeic and cinnamic acids and malvidin in Faq. The Fea was composed mainly of caffeic and cinnamic acid derivatives as well as flavan-3-ols, flavones and flavanone derivatives. Based on their UV–Vis spectral data the flavan-3-ols, flavones and flavanones detected in both fractions seem to correspond to bound forms of catechin, luteolin and apigenin and naringenin, respectively. The Faq fractions were the major contributors to the ABTS antioxidant capacity (77–82%). The results obtained in the present study suggest that oca tubers could potentially be considered beneficial for human health and for potential industrial applications.

Application of liquid–liquid–liquid microextraction and ion-pair liquid chromatography coupled with photodiode array detection for the determination of chlorophenols in water

A method termed as liquid–liquid–liquid microextraction was utilized to extract chlorophenols from water. The extracted chlorophenols, present in anionic form, were then separated, identified, and quantitated by ion-pair high-performance liquid chromatography with photodiode array detection (HPLC/DAD). For trace chlorophenol determination using HPLC/DAD, the chlorophenolate anion provides a better ultraviolet spectrum for quantitative and qualitative analyses than does uncharged chlorophenol. This is due to the auxochromic effect of the phenolate anion. In the study, experimental conditions such as organic phase identity, acceptor phase volume, sample agitation, extraction time, acceptor phase NaOH concentration, donor phase HCl concentration, salt addition, and UV absorption wavelength were optimized. Relative standard deviations (RSD, 2.3–5.4%), coefficients of determination ( r 2 0.9994–0.9999), and detection limits (0.049–0.081 ng mL −1) of the proposed method were investigated under the selected conditions. The method was successfully applied to analyses of reservoir and tap water samples, and the relative recoveries of chlorophenols from the spiked reservoir and tap water samples were 94.1–100.4% and 87.8–101.2%, respectively. The proposed method is capable of identifying and quantitating each analyte to 0.5 ng mL −1, confirming the HPLC/DAD technique to be quite robust for monitoring trace levels of chlorophenols in water samples.

Determination of Ethylene Thiourea in Urine by HPLC-DAD

Ethylene thiourea (ETU) is a metabolite of ethylenebisdithiocarbamates (EBDCs); it is the best indicator of exposure to these fungicides. Therefore, high-performance liquid chromatography with photodiode-array detection (HPLC-DAD) was optimized and validated for the determination of ETU in human urines. Urine samples were extracted by solid-phase extraction using Extrelut and analyzed using HPLC-DAD set at 231 nm. The analyses were carried out using a mobile phase of 0.01 M phosphate buffer (pH 4.5) on a C18 Uptisphere NEC-5-20, 250- x 4.6-mm x 5-microm column. The internal standard used was 4-pyridinecarboxylic acid hydrazide. The method was successfully validated in compliance with requirements set by the International Committee on Harmonization 1996. The lower limit of quantitation was at 1 microg/L, and the linearity was studied from 1 to 100 microg/L. There were 272 urine samples collected from farmers exposed to EBDCs in different regions in France analyzed in this study.

Squalene in hair—a natural reference substance for the improved interpretation of fatty acid ethyl ester concentrations with respect to alcohol misuse

Fatty acid ethyl esters (FAEE) are incorporated into hair mainly from sebum. For this reason, the use of their concentration C FAEE as marker of excessive alcohol consumption is complicated by interindividual differences of the activity of the sebum glands and of elimination by hair care and hair cosmetics. Furthermore, an influence of the investigated hair length due to increasing accumulation from proximal to distal was found. Therefore, it was examined whether these sources of error can be avoided if in addition to C FAEE the relative FAEE concentrationa C FAEE/ C SQ related to squalene SQ as a natural reference compound were used for interpretation. Sebum containes about 10–20% SQ. A sensitive and reliable method for the determination of SQ in addition to FAEE from the same hair extracts by high performance liquid chromatography with photo diode array detector (HPLC-DAD) was developed. The concentrations of ethyl myristate, ethyl palmitate, ethyl oleate, ethyl stearate and squalene were determined and C FAEE/ C SQ was calculated for 13 teetotallers, 16 social drinkers, 12 fatalities with excessive alcohol abuse at life time and 9 cases with unclear alcohol anamnesis. C SQ ranged from 0.02 to 1.97 μg/mg (mean 0.67 μg/mg). From the results follows that squalene enables a control of the lipid content of hair and a correction of C FAEE in cases with deviations from the usual lipid content in a similar way as creatinine in urine. Preliminary values of C FAEE/ C SQ were suggested for the upper limit for teetotallers (< 0.6 ng/μg) and the lower limit for excessive alcohol abuse (> 1.5 ng/μg). However, the relative concentration C FAEE/ C SQ cannot completely replace the absolute concentration C FAEE, and both should regularly be used for an improved interpretation with respect to alcohol abuse.

Determination of Folpet, Procymidone, and Triazophos in Groundwater by HPLC Using Partial Least Squares and Principal Component Regression

Three multivariate calibration methods, partial least squares (PLS-1 and PLS-2) and principal component regression (PCR), full spectrum calibration methods, were applied to the simultaneous determination of the three pesticides folpet, procymidone and triazophos, in mixtures, by high performance liquid chromatography with photodiode array detection (HPLC-DAD). The effects of several preprocessing techniques are discussed in order to optimize the calibration matrices by the PLS and PCR methods. The use of mean-centering and smoothing function chromatograms allows better prediction of the samples. The average recoveries from the different mixtures assayed ranged between 79.6 % and 114.0 %. No advantages were found for the prior differentiation step.

HPLC and TLC profiles of phenylalkylamines of khat (Catha edulis Forsk.) confiscated in Switzerland

Khat, the leaves of Catha edulis Forsk., is widely used as a “natural amphetamine” in East Africa and the Arab Peninsula. In recent years, air-transported khat appeared on local khat markets in Europe (e.g. France, Italy, Great Britain). Due to increasing seizures of khat imported by ethnic communities and emigrants the psychoactive drug has been banned in 1992 by the Swiss Health Authorities. The demand for monitoring the psychotropic potential of the illicit drug made it necessary to establish specific, sensitive and rapid methods for the determination of the alkaloids, mainly the psychoactive principle S-(−)-cathinone. The S-(−)-cathinone concentration measured by high-performance liquid chromatography with photodiode array detection (HPLC-DAD) and alternatively by instrumental thin-layer chromatography (TLC) varied between 2182 and 4506 μg/g dried material. It is typical for khat originating from Kenya and demonstrates its high psychoactivity. The S,S-(+)-norpseudoephedrine and R,S-(−)-norephedrine content ranged from 2114 to 4094 and 192 to 1345 μg/g, respectively.