High-Performance Liquid Chromatographic Analysis of Sarcosine as a Fluorescent Levofloxacin Derivative.

Sarcosine, N-methyl glycine, could be used as a biomarker for the diagnosis of prostate cancer. It exists in biosamples at low levels; therefore, sensitive methods are necessary for its detection. In this study, we developed a sensitive and selective method for the analysis of sarcosine, based on derivatizing sarcosine with a fluorescent reagent levofloxacin acyl chloride. The resulting derivative is highly responsive to a fluorimetric detector (λex = 290 nm, λem = 460 nm). The sarcosine derivative can be separated from its molecular isomers (α-l-alanine, α-d-alanine and ß-alanine) on a hexyl-phenyl column by gradient elution using sodium acetate buffer (pH 3.8; 50 mM) and tetrahydrofuran as the mobile phase. The method showed a determination range of sarcosine in water over 44.5-1780.0 ng/mL (0.5-20.0 µM) and the limit of detection at 8.9 ng/mL (0.1 µM) (S/N = 3 with 20 µL injected). Intra- and inter-day precision (as % relative standard deviation) and accuracy (as % relative error) were all below 4.8%. Application of the method to the analysis of sarcosine in human urine proved feasible.

Simultaneous determination of regular insulin and insulin aspart by capillary zone electrophoresis and application in drug formulations.

A rapid and simple capillary zone electrophoresis (CZE) with ultraviolet detection has been developed for simultaneous determination of regular insulin and insulin aspart in bulk and commercial injection dosage forms. The simultaneous analysis of the tested drugs was performed in phosphate buffer (80 mM; pH 6.5). The separation of regular insulin and insulin aspart was achieved at 13 kV and detection at 200 nm within 7 min with RSD for the absolute migration time reproducibility of less than 3.8% (n=10). Selectivity, linearity, precision, accuracy, limits of quantification (LOQ) and limits of detection (LOD) were evaluated as the method validation. Calibration plots were linear (r>0.999) over a range of 2.0-60.0 microg/mL for regular insulin and insulin aspart. The LOD were all 1.0 microg/mL (signal-to-noise ratio=3; injection 6.89 kPa, 7s). The small amount of sample required and the expeditiousness of the procedure can be an advantageous alternative to traditional methodology for the quantification of tested drugs in individual pharmaceutical products.

Expression of BUBR1 in human oral potentially malignant disorders and squamous cell carcinoma.

OBJECTIVE: BUBR1 is one of the key components of the spindle assembly checkpoint (SAC) machinery and is activated in response to kinetochore tension. Defects in the SAC contribute to an increased rate of aneuploidization during tumorigenesis. The aim of the present study was to examine the immunohistochemical expression of BUBR1 protein for human oral squamous cell carcinogenesis. STUDY DESIGN: A total of 120 samples of squamous cell carcinoma (SCC, n = 43) and 5 types of potentially malignant disorders (PMDs: oral epithelial dysplasia, n = 11; hyperkeratosis/epithelial hyperplasia, n = 20; lichen planus, n = 16; submucous fibrosis, n = 19; and verrucous hyperplasia, n = 11) of human oral mucosa (1991-2001) from our institution were retrieved and immunohistochemical staining were performed. Normal oral mucosa (n = 9) and fibrous hyperplasia (n = 9) from patients without the aforementioned oral habits were also included in the study. RESULTS: BUBR1 staining was detected at the basal and suprabasal layers in 75 (97.4%) of 77 samples of PMD and 43 (100%) of 43 samples of SCC of oral mucosa but was absent in all samples of normal oral mucosa (n = 9) and fibrous hyperplasia (n = 9). BUBR1 expression of various types of PMD and SCC of oral mucosa was significantly overexpressed as compared respectively with normal mucosa (P < .001) and fibrous hyperplasia (P < .001). Moreover, the expression of oral SCC was significantly higher as compared respectively with the 5 types of oral PMD; on the other hand, BUBR1 expression of verrucous hyperplasia was significantly higher than that of the other 4 types of PMD of oral mucosa (P < .001). CONCLUSION: Our results may interpret that BUBR1 protein is suggested to be one of the contributing factors involved in the pathogenesis of oral SCC. These also hypothesize that BUBR1 protein is a putative biomarker for human oral squamous cell carcinogenesis.

A non-fluorescent derivative from derivatizing trans, trans-muconic acid with 2-(2-naphthoxy)ethyl-2-(piperidino)ethanesulfonate.

trans, trans-Muconic acid (MA) is a polar metabolite of benzene and used as a biomarker for monitoring human exposure to benzene. Because MA is a trace metabolite, sensitive method is required for its detection. In addition, MA is a highly polar compound with dicarboxyl functions that could incur unfavorable adsorption on silica-based stationary phase usually used for separation. To address these problems, we planned to derivatize MA with a fluorescent reagent 2-(2-naphthoxy)ethyl-2-(piperidino)ethanesulfonate to give a naphthoxy derivative of MA for improving detection sensitivity and chromatographic properties. Surprisingly, the resulting derivative shows no fluorescent activity (lambda(ex): 226 nm; lambda(em): 350 nm). The negative results could be used as an instructive example for discussing on fluorescence quenching.

On-line microdialysis coupled with liquid chromatography for biomedical analysis.

This work presents a review of the microdialysis (MD) sampling technique for on-line coupling with high-performance liquid chromatography (HPLC) for biomedical analysis. MD-HPLC was first used in the field of brain metabolism to study neurotransmission, and this remains its common application in the biomedical field. On-line MD-HPLC hyphenated methods provide advantages over those with off-line MD-based techniques, including simplified sample preparation, automated analyses, avoidance of contamination introduced during the analytical process, and in situ analysis of the extracellular fluid of living organisms. This review outlines the effectiveness of the continuous monitoring of unbound chemicals from tissues, organs, and body fluids by on-line MD-HPLC methods. In addition, a discussion is presented on the application of in vivo on-line MD-HPLC toward obtaining biochemical event information in the extracellular fluid of various tissues and in biological fluids for pharmacokinetic, pharmacodynamic, toxicological, and bioprocess monitoring.

Automated on-line microdialysis sampling coupled with high-performance liquid chromatography for simultaneous determination of malondialdehyde and ofloxacin in whole blood.

We have developed a system that couples an on-line microdialysis (MD) system with flow injection high-performance liquid chromatography (HPLC)-fluorescence detection for simultaneous measurement of the concentrations of malondialdehyde (MDA) and ofloxacin (OFL) in whole blood samples. The sample matrix was first cleaned with an MD system using an MD probe. A continuously flowing dialysate stream was derivatized on-line and auto-injected into a separation column. MDA and OFL were separated through a reverse-phase C18 column (250 mm x 4.6 mm) at a flow rate of 0.8 mL min(-1) and then detected using a fluorescence detector (excitation: 532 nm; emission: 553 nm); the system's components were connected on-line using a valve control. Validation experiments demonstrated good linearity, precision, accuracy, and recovery. The precisions for the determinations of MDA and OFL, measured in terms of relative standard deviations, were 6.5% and 4.6%, respectively, for intra-day assays and 7.5% and 8.7%, respectively, for inter-day assays. The average recoveries of MDA and OFL spiked in plasma were each close to 100%. The use of this on-line MD-HPLC system permitted continuous monitoring of MDA and OFL in OFL-treated whole blood subjected to UV-A irradiation. Based on our results, the UV-A irradiation markedly increased the level of MDA in the OFL-treated whole blood.

A simple micellar electrokinetic capillary chromatographic method for the quantitative analysis of organic expectorants.

A simple and rapid micellar electrokinetic capillary chromatographic method is described for the separation and quantification of five expectorant drugs, including ambroxol (AMB), bromhexine (BRM), carbocysteine, guaiacol and guaifenesin. The drugs were separated in a mixed solution of phosphate buffer (35 mM; pH 3.90) and acetonitrile (75:25, v/v) with sodium dodecylsulfate (120 mM) as the micellar source, and the separated drugs were directly monitored with UV detector (200 nm). Several key parameters affecting the separation and analysis of the drugs were studied and optimized. Based on the corrected peak-area ratios of the drugs to an internal standard (7-hydroxy-4-methyl-coumarin) versus the concentration of the drugs, the established method is applicable to quantify AMB and BRM each over 20-150 microM, carbocysteine over 100-1500 microM, guaiacol and guaifenesin each over 10-150 microM. The detection limits (S/N=3; 0.5 psi, 5 s injection) of the method for the analytes are in the range of 3.0-5.0 microM (except that of carbocysteine at 30 microM). The precision (relative standard deviation) and accuracy (relative error) of the method for the intra-day (n=3) and inter-day (n=5) analyses of the analytes at three levels are all below 4%. The method is speedy with a run time of about 6 min for the analysis of the five analytes. Application of this method to the analysis of AMB and BRM in pharmaceutical preparations or AMB in the urine of a dosed subject proved simple and effective.

On-line microdialysis-nano-Au/TiO2-high-performance liquid chromatography system for the simultaneous determination of cobalt and nickel in water.

An on-line configuration of microdialysis (MD), Au/TiO2 nanoparticle preconcentration, and high-performance liquid chromatography-ultraviolet (HPLC-UV) detection method was developed for the simultaneous measurement of cobalt (Co) and nickel (Ni) concentrations in water. The sample matrix was first cleaned with an MD system using a MD probe. A continuously flowing dialysate stream was introduced into tubing coated with Au/TiO2 nanoparticles to adsorb metals, followed by elution by an acidic eluent. The enriched samples were derivatized on-line using 8-hydroxyquinoline. The separation of Co and Ni were achieved by using a LC-C18 column. The three aforementioned system components were connected on-line using a valve control. The UV detection was performed at 319nm. Validation experiments demonstrate good linearity, precision, accuracy, and recovery. The proposed method offers a simple and reliable procedure to determine the levels of Co and Ni in environmental water samples. Moreover, the methodology described in this study adheres to the concept of green chemistry, including the absence of organic solvents in the MD sampling and extraction processes. To the best of our knowledge, the proposed method is the first reported on-line connection of MD, Au/TiO2 nanoparticle tubing, and HPLC devices for the measurement of Co and Ni concentrations in water.

Simultaneous determination of malondialdehyde and ofloxacin in plasma using an isocratic high-performance liquid chromatography/fluorescence detection system.

This study establishes the applicability of using high-performance liquid chromatography (HPLC) with fluorescence detection for the simultaneous determination of malondialdehyde (MDA) and ofloxacin (OFL). The MDA and OFL were separated through a reverse-phase C18 column (250 mm x 4.6 mm) at a flow rate of 1.0 mL min(-1) and then detected using a fluorescence detector (excitation: 532 nm; emission: 553 nm). The separation conditions were optimized by varying the concentration and pH of the phosphate buffer and the percentage of organic solvent; the optimal mobile phase was a mixture of 50 mM phosphate buffer (adjusted to pH 5.8 with potassium hydroxide) and methanol (45:55, v/v). The retention times of MDA and OFL were 3.6 and 5.9 min, respectively, with detection limits (at a signal-to-noise ratio of 3) of 0.015 and 4.0 microM, respectively. This method afforded linear responses between the MDA and OFL concentrations and the HPLC peak areas within the ranges 0.15-2.43 microM and 0.06-1.0 mM, respectively. The precisions of the determinations of MDA and OFL, measured in terms of relative standard deviations, were 1.6-5.0% and 1.9-3.6%, respectively, for intra-day assays and 1.0-4.3% and 0.3-1.8%, respectively, for inter-day assays. The average recoveries of MDA and OFL spiked in plasma were 100.4% and 98.8%, respectively. To the best of our knowledge, this paper describes the first practical analytical approach toward simultaneously monitoring the levels of MDA and OFL in plasma. The OFL-induced oxidative stress measured using this method indicated that OFL treatment did not markedly increase the level of MDA.

A simple and sensitive liquid chromatographic method for the analysis of free docosanoic, tetracosanoic and hexacosanoic acids in human plasma as fluorescent derivatives.

Docosanoic (C22), tetracosanoic (C24) and hexacosanoic (C26) acids are saturated very-long-chain fatty acids (VLCFA) present at trace levels in biosamples. VLCFA can be used as potential biomarkers for the diagnosis of hereditary diseases such as X-linked adrenoleukodystrophy. Because the analytes to be detected are at trace levels, a sensitive fluorimetric liquid chromatographic method was developed to analyze VLCFA in plasma. The method is simple based on extracting VLCFA from plasma with toluene, and the obtained toluene extract was subject to the derivatization of VLCFA with a fluorescent reagent 2-(2-naphthoxy)ethyl-2-(piperidino)ethanesulfonate (NOEPES) without solvent evaporation/replacement. The resulting fluorescent derivatives were monitored by fluorimetric detection (excitation at 225nm and emission at 360nm), giving a high sensitivity with the limit of detection about 5.0nM (S/N=3, 10microL injected) of the analytes. Application of the method to the analysis of VLCFA in the plasma of patients with adrenoleukodystrophy proved practical and effective.

Fluorescent high-performance liquid chromatographic analysis of vigabatrin enantiomers after derivatizing with naproxen acyl chloride.

Vigabatrin is widely used as an anticonvulsant in the treatment of seizures. Vigabatrin is usually supplied as racemate in formulation, but only the (S)-(+)-enantiomer of vigabatrin is pharmacologically active. A simple and sensitive liquid chromatographic method is described for the separation and quantification of vigabatrin enantiomers. The method is based on derivatizing racemic vigabatrin with a fluorescent chiral reagent (naproxen acyl chloride). The resulting diastereomeric derivatives are highly responsive to a fluorimetric detector (lambda(ex)=230 nm, lambda(em)=350 nm). The lower quantitation limit of the method is attainable at 25 nM for (S)-(+)-vigabatrin or (R)-(-)-vigabatrin with a detection limit of about 2.5 nM (S/N=3 with 10 microl injected). Application of the method to the analysis of vigabatrin in serum of dosed patients proved feasible.

Trends in the use of lipid-lowering drugs by outpatients with diabetes in Taiwan, 1997-2003.

PURPOSE: To analyze recent trends in LLD use by outpatients with diabetes in Taiwan over a 7-year period and to see whether the trends are consistent with clinical trial outcomes and published guidelines. METHODS: A cross-sectional survey was implemented using the National Health Insurance Research Database between January 1997 and December 2003. Adult outpatients who had diagnoses of diabetes and dyslipidemia and who had concurrent antidiabetic drug and LLD claim were identified. The prescribing trends were described in terms of the prescribing rates and patterns of LLDs in each study year. RESULTS: Between 1997 and 2000, fibrates were the most widely prescribed major class of LLDs, while the rates showed a significant decline over time. The proportions of statin use increased 3.3-fold from 20.8% of all LLDs in 1997 to 69.0% in 2003. With the continuous increase in the use of statins, by 2001, they replaced fibrates as the most heavily used LLDs. Monotherapy was the most prominent prescribing pattern comprising 98.1% and 98.3% of all LLD prescriptions in 1997 and 2003, respectively. Among patients with diabetic dyslipidemia plus CHD, the use of statins increased almost threefold instead of a large reduction in the percentage with non-statin class. CONCLUSIONS: The prescribing rates of LLDs shifted from the fibrates to statins, especially in newer statin. The majority of patients were maintained on monotherapy. These findings may imply that management of hyperlipidemia in patients with diabetes had a positive trend towards recent clinical trial outcomes and guideline's recommendation.

Effects of zinc compound on body weight and recovery of bone marrow in mice treated with total body irradiation.

This study aimed to investigate if zinc compound would have effects on body weight loss and bone marrow suppression induced by total body irradiation (TBI). ICR mice were divided randomly into two groups and treated with test or control compounds. The test compound contained zinc (amino acid chelated with bovine prostate extract), and the control was reverse osmosis pure water (RO water). One week after receiving the treatment, mice were unirradiated, or irradiated with 6 or 3 Gy by 6 MV photon beams to the total body. Body weight changes were examined at regular intervals. Three and 5 weeks after the radiation, animals were sacrificed to examine the histologic changes in the bone marrow. Lower body weight in the period of 1-5 weeks after radiation and poor survival rate were found after the 6 Gy TBI, as compared with the 3 Gy groups. The median survival time after 6 Gy and 3 Gy TBI for mice given the test compound were 26 and 76 days, respectively, and the corresponding figures were 14 and 70 days, respectively, for mice given the control compound (p < 0.00001). With zinc supplement, the mean body weight in mice which received the same dose of radiation was 7-8 g heavier than in the water-supplement groups during the second and third weeks (p < 0.05). Hence, there was no statistically significant difference in survival rate between zinc and water supplement in mice given the same dose of irradiation. Histopathologically, there was less recovery of bone marrow cells in the 6 Gy groups compared with the 3 Gy groups. In the 3 Gy water-supplement group, the nucleated cells and megakaryocytes were recovered in the fifth week when recovery was still not seen in the 6 Gy group. With zinc supplement, these cells were recovered in the third week. In this study, we found that zinc is beneficial to body weight in mice treated with TBI. Histologic examination of bone marrow showed better recovery of bone marrow cells in groups of mice fed with zinc. This study suggests that zinc can be used as supplements in cancer patients receiving radiotherapy to reduce radiation-induced complications.

A sensitive liquid chromatographic method for the analysis of isovaleric and valeric acids in urine as fluorescent derivatives.

A simple and sensitive isocratic liquid chromatographic method was developed for the analysis of isovaleric and valeric acids in human urine as biomarkers in metabolic acidosis. The method is based on the derivatization of isovaleric and valeric acids with a fluorescent reagent 2-(2-naphthoxy)ethyl-2-(piperidino)ethanesulfonate for labeling the analytes with the naphthoxy fluorophore. The resulting fluorescent derivatives of isovaleric and valeric acids were separated on a phenyl-hexyl column, using a mixed solvent of methanol-water-tetrahydrofuran (55:31:14, v/v) as the mobile phase. The separated derivatives were monitored with a fluorimetric detector (excitation at 225 nm and emission at 360 nm). The linear range of the method for the determination of isovaleric acid or valeric acid derivative was over 0.2 approximately 8.0 microM. The detection limit (signal to noise ratio=3 with 10 microl injected) of isovaleric acid or valeric acid was about 0.04 microM. Application of the method to the analysis of isovaleric acid in the urine of a patient with isovaleric acidemia proved feasible.

Combining high-performance liquid chromatography with on-line microdialysis sampling for the simultaneous determination of ascorbyl glucoside, kojic acid, and niacinamide in bleaching cosmetics.

We have used on-line microdialysis sampling coupled with high-performance liquid chromatography and UV-vis detection to simultaneously determine the contents of ascorbyl glucoside (AA-2G), kojic acid (KA), and niacinamide (VitB(3)) in commercial bleaching cosmetics. Our results indicate that AA-2G, KA, and VitB(3) separated well within 4.5 min on a reverse-phase Hypersil Fluophase PFP column when eluting with 0.020 M phosphate buffer solution in 40% (v/v) methanol at pH 5.5. The calibration curves were linear over the ranges 0.068-304, 0.071-284, and 0.024-488 microg mL(-1) for AA-2G, KA, and VitB(3), respectively, with correlation coefficients for the linear regression analyses falling within the range 0.9982-0.9999. The detection limits for AA-2G, KA, and VitB(3) were 0.01, 0.01, and 0.007 microg mL(-1), respectively. The detection wavelength was robust when the levels of the analytes in the samples were high (0.1-2%). The analytes were all detected using ultraviolet light (254 nm). The compounds diffuse through the membrane more readily when KA and VitB(3) are in their molecular forms and AA-2G is ionized. The recoveries were in the range 92-106% with good reproducibility (R.S.D.=3.9-8.7%). We used this procedure to assay six commercially available bleaching cosmetics; our results confirmed not only the precision of the method but also the claims made on the labels of the cosmetics. This approach provides a very simple means to determine the contents of AA-2G, KA, and VitB(3) in various dosages in bleaching cosmetics.

Highly sensitive analysis of cholesterol and sitosterol in foods and human biosamples by liquid chromatography with fluorescence detection.

A simple and sensitive method is described for the quantitative analysis of important animal and plant sterols (cholesterol and sitosterol) by liquid chromatography with fluorimetric detection. The method is based on the derivatization of cholesterol and sitosterol with a fluorescent reagent (naproxen acyl chloride) in toluene. The resulting derivatives were isocratically separated on a C(8) column with a mixed solvent of methanol-isopropanol-water (90:5:5, v/v) as a mobile phase and monitored with a fluorimetric detector (excitation 231 nm and emission 352 nm). The linear range for the quantitation of cholesterol or sitosterol was 0.1-2.0 microM with a detection limit (S/N=3 with 10 microl injected) of about 25 nM. Recoveries of cholesterol spiked in milk (n=5) ranged over 99-104% with relative standard deviations (RSD) less than 6.0%. Application of the method to the analysis of cholesterol or sitosterol in milk, saliva and urine proved simple and feasible.

Trends in the use of antihypertensive drugs by outpatients with diabetes in Taiwan, 1997-2003.

PURPOSE: To analyze trends in AHD-use by diabetic outpatients in Taiwan over a 7-year period (1997-2003) and to see whether the trends are consistent with clinical trial outcomes and published guidelines. METHODS: A cross-sectional survey was implemented using National Health Insurance Research Database between January 1997 and December 2003. Adult outpatients who had diagnoses of diabetes and hypertension and who had concurrent antidiabetic and antihypertensive drug claim were identified. The prescribing trends were described in terms of the prescribing rates and patterns of AHDs in each study year. RESULTS: Of the AHDs, CCBs were the most widely prescribed class throughout the study period but the prescribing rates declined considerably over the study period. A significant downward trend was also observed for beta-blockers and other classes. Drugs acting on the RAS were the only one class showing a significant increase in prescribing rates with time. The prescribing patterns for monotherapy regimen decreased over time while those for two-, three-, and four or more drug regimens increased over time. Monotherapies maintained with CCBs, beta-blockers, diuretics, and other classes steadily declined but those maintained with drugs acting on the RAS markedly increased. CONCLUSIONS: The use of drugs acting on the RAS showed a marked increasing trend over the course of the study. Physicians' prescribing patterns for AHD are increasingly involving multi-drug regimens. These findings may imply that management of hypertension in patients with diabetes had a positive trend toward to new clinical trial outcomes and guideline's recommendation.

Real-time dynamic monitoring of nonprotein-bound copper in the blood.

To obtain real-time dynamic changes of non-protein-bound copper in the blood, we have developed an online microdialysis sampling system coupled with a flow-injection graphite furnace-atomic absorption spectrometer (FI-GFAAS). The analytical performances of the online system such as linearity, limit of detection, precision, and spiked recoveries were validated. Before the in vivo experiments, the in vivo recovery was conducted. The levels of non-protein-bound Cu in the blood of living rabbits were evaluated before and after administering them with 5 mg/kg body weight of CuSO4 by the online microdialysis-FI-GFAAS system. The results showed that the average basal concentration of non-protein-bound Cu in the blood of living rabbits was 16.2 microg/L (n = 3). Furthermore, the levels of non-protein-bound Cu in the blood of living rabbits were observed after a long delay following intravenous injection of CuSO4. The non-protein-bound Cu reached the maximum value at 125 min after injection. Our present study might provide the in vivo, direct observation that different metals have their own binding characteristics with proteins when transported into the blood of living organisms.

Micellar electrokinetic capillary chromatographic method for the separation and quantitation of multiple amino acids as naphthoxy derivatives in pharmaceutical formulations.

The MEKC method is described for the quantitative analysis of 17 amino acids (AA) in pharmaceutical products. The method is based on simply derivatizing the AA with (2-naphthoxy)acetyl chloride under mild conditions. The resulting derivatives were separated by MEKC with borate buffer (35 mM; pH 9.50) including 150 mM SDS at the applied voltage of 25 kV in an uncoated capillary (effective length, 40 cm) and monitored by UV at 230 nm. The detection limits of the amino acid derivatives were in the range of 3.0-8.0 microM (S/N = 3, injection 5.0 s, 6 895 Pa). The precision (RSD) and accuracy (relative error) of the method for intra- and interday analyses of the analytes are all below 5.2%. The amino acid derivatives are stable at room temperature for 33 h studied and the molar absorptivity of the alanine derivative (used as a model) is stable over a wide pH range of 3.00-12.00. This is favorable for monitoring the derivatives in various pH by CE or LC. Application of the method to the analysis of multiple AA in a liquid injection formulation proved satisfactory.

Micellar electrokinetic capillary chromatographic method for the quantitative analysis of uricosuric and antigout drugs in pharmaceutical preparations.

A simple MEKC method is described for the separation and quantification of seven widely used uricosuric and antigout drugs, including allopurinol (AP), benzbromarone (BZB), colchicine (COL), orotic acid (OA), oxypurinol (OP), probenecid (PB), and sulfinpyrazone (SPZ). The drugs were separated in a BGE of borate buffer (45 mM; pH 9.00) with SDS (20 mM) as the micellar source and the separated drugs were directly monitored with a UV detector (214 nm). Several parameters affecting the separation and analysis of the drugs were studied. Based on the normalized peak-area ratios of the drugs to an internal standard versus the concentration of the drugs, the method is applicable to quantify BZB, COL, and SPZ (each 5-200 microM), AP, OA, OP, and PB (each 10-200 microM) with detection limits (S/N = 3, 0.5 psi, 5 s injection) in the range of 0.6-4.0 microM. The precision (RSD; n = 5) and accuracy (relative error; n = 5) of the method for intraday and interday analyses of the analytes at three levels (30, 120, and 180 microM) are below 4% (n = 3). The method was demonstrated to be suitable for the analysis of AP and COL in commercial tablets with speed and simplicity.