Determination of Benzo[a]pyrene in Edible Oil by High Performance Liquid Chromatography-Fluorescence Detector (HPLC-FL)

In this study, an optimized high performance liquid chromatography-fluorescence detector (HPLC-FL) method for the determination of benzo[a]pyrene in edible oil was established. HPLC was performed with Thermo Fisher Scientific C18 column (250 mm×4.6 mm, 5 μm) as the chromatographic column and acetonitrile and water as the mobile phase, and the excitation wavelength and emission wavelength of fluorescence detector were 286 and 430 nm, respectively. The response was high, and the linear range was 0.5-10.0 ng/ml. The lowest limit of detection was 0.11 ng/ml, and the average recovery was 92.5%. This method is suitable for quantitative analysis of benzo[a]pyrene content in edible oil.

Efficient and selective extraction of uranium from seawater based on a novel pulsed liquid chromatography radionuclide separation method

The novel pulsed liquid chromatography radionuclide separation method presented here provides a new and promising strategy for the extraction of uranium from seawater.In this study,a new chromatographic separation method was proposed,and a pulsed nuclide automated separation device was developed,alongside a new chromatographic column.The length of this chromatographic column was 10 m,with an internal warp of 3 mm and a packing size of 1 mm,while the total separation units of the column reached 12,250.The most favorable conditions for the separation of nuclides were then obtained through optimizing the separation conditions of the device:Sample pH in the column=2,sample injection flow rate=5.698 mL/min,chromatographic column heating temperature=60℃.Separation experiments were also carried out for uranium,europium,and sodium ions in mixed solutions;uranium and sodium ions in water samples from the Ganjiang River;and uranium,sodium,and magnesium ions from seawater samples.The separation factors between the different nuclei were then calculated based on the experimental data,and a formula for the separation level was derived.The experimental results showed that the separation factor in the mixed solution of uranium and europium(1:1)was 1.088,while achieving the initial separation of uranium and europium theoretically required a 47-stage separation.Considering the separation factor of 1.50for the uranium and sodium ions in water samples from the Ganjiang River,achieving the initial separation of uranium and sodium ions would have theoretically required at least a 21-stage separation.Furthermore,for the seawater sample separation experiments,the separation factor of uranium and sodium ions was 1.2885;therefore,more than 28 stages of sample separation would be required to achieve uranium extraction from seawater.The novel pulsed liquid chromatography method proposed in this study was innovative in terms of uranium separation and enrichment,while expanding the possibilities of extracting uranium from seawater through chromatography.

Determination of Veterinary Drug Residues in Animal-derived Foods by Liquid Chromatography-Mass Spectrometry

[Objectives]This study was conducted to establish a rapid and effective method for simultaneous extraction of 54 kinds of veterinary drug residues in animal-derived food, including sulfonamides, quinolones, tetracyclines, malachite greens, penicillins, nitroimidazoles, tranquilizers and macrolides, by HPLC-MS. [Methods] The samples were extracted with 80% acetonitrile water(containing 0.1% formic acid), combined with QuEChERS extraction technology and C18 and PSA purification, analyzed by high performance liquid chromatography-mass spectrometry, and quantified by external standard method. The target substances were analyzed on ZORBAX Eclipse C18 chromatographic column using 0.2% formic acid water and 0.2% methanol as mobile phases. The gradient elution mode was used for chromatographic separation and multiple reaction detection. [Results] In the linear range of 0.5-50.0 ng/ml, the linear relationship of the 54 kinds of veterinary drug residues was good, with correlation coefficients(r~2) greater than 0.995, and the detection limits ranged from 0.30 to 1.00 μg/kg. The results showed that the recovery ranged from 75.4% to 118.2% when different matrixes were added for recovery. [Conclusions] This method is simple, efficient, accurate, stable, and highly operable. It is applicable to simultaneous batch screening of veterinary drug residues in animal-derived food, and has high practical application value.

Determination of Voriconazole in Human Plasma by Liquid Chromatography Tandem Mass Spectrometry: Application in Therapeutic Drug Monitoring

A sensitive, accurate and robust Liquid Chromatography Tandem Mass Spectrometry method has been developed and validated to measure voriconazole trough levels in human plasma. The plasma samples were mixed with fluconazole as an Internal Standard and directed to protein precipitation and drug extraction. An aliquot of 1 μl was injected into the chromatographic system and separated by the Acquity BEH C18 column at a flow rate of 0.30 ml/min in a gradient mobile phase consisting of acetonitrile, Ultrapure water (UPW), methanol and formic acid. Voriconazole was detected by a Triple Quadrupole Detector (TQD) operating on Multiple Reaction Monitoring (MRM) and a positive ion mode Electrospray ionization (ESI) Q1 mass: 350.1 m/z, Q3 mass: 281.1 m/z. Method linearity of the calibration curve (0.10 - 8.00 μg/ml) indicated a correlation coefficient r ≥ 0.99. The intra and inter-assay accuracy was within 85% - 115% and the intra and inter-assay precision was ≤5.76%. Voriconazole recovery percentage was between 97.69 - 119.62%. The method was successively applied in routine voriconazole TDM.

A High-Performance Liquid Chromatography Method for the Simultaneous Determination of Five Index Components in Danhong Injection

The purpose of this study was to establish a high-performance liquid chromatography (HPLC) method for the simultaneous determination of sodium danshensu, protocatechuic aldehyde, rosmarinic acid, salvianolic acid B, and 4-coumaric acid in Danhong injection. The chromatographic method employed was as follows: the column was a Welch Ultimate XB-C18 column (250 mm × 4.6 mm, 10 μm), the mobile phase was a gradient elution of 0.4% formic acid aqueous solution (A) and acetonitrile (B), the detection wavelengths were 280 nm for sodium danshensu, protocatechuic aldehyde, and salvianolic acid B and 326 nm for 4-coumaric acid and rosmarinic acid, the sample volume was 10 μL, the flow rate was 1.0 mL/min, and the column temperature was 35°C. This method can realize the separation and determination of sodium danshensu, protocatechuic aldehyde, rosmarinic acid, salvianolic acid B, and 4-coumaric acid within 50 minutes. The linear relationships of the five peak areas and their concentrations are good (R2> 0.9997). The precision RSD values are all less than 1.0%. The reproducibility RSD values are all less than 1.3%. The stability RSD values are all less than 2.2%. The recovery values ranged from 92.4% to 99.4%. This method is simple, accurate, and reproducible. It can be used for the determination of sodium danshensu, protocatechuic aldehyde, rosmarinic acid, salvianolic acid B, and 4-coumaric acid in Danhong injection.

Determination of egg and milk allergen in food products by liquid chromatography-tandem mass spectrometry based on signature peptides and isotope-labeled internal standard

The aim of this work was to develop a liquid chromatography-tandem mass spectrometry method for the determination of milk allergen and egg allergen in food products.Signature peptides GGLEPINFQTAADQAR,VGINYWLAHK,VLVLDTDYK,FFVAPFPEVFGK,and NAVPITPTLNR were confirmed and synthesized as the quantitative peptide of ovalbumin,α-lactalbumin,β-lactoglobulin,α_(S1)-casein andα_(S2)-casein,the relative isotope-labeled internal standards were used in the quantitative analysis.Linear range was in the range of0.5-5000.0 nmol/L for egg and milk allergen in bread,cake,cookie,rice crust and wheat flour samples with free from egg and milk,the limits of detection of milk allergens and egg allergen were in the range between0.94 mg/100 g and 56.71 mg/100 g,limits of quantification of milk allergens and egg allergen were in the range between 2.36 mg/100 g and 141.78 mg/100 g.The recoveries ranged from 76.7%to 122.8%,the relative standard deviations were in the range of 1.60%-15.60%.The developed method has been successfully used for the detection of egg and milk allergen in various food samples.

Simultaneous Determination of 14 β-Receptor Agonists Residues in Mutton by High Performance Liquid Chromatography-Tandem Mass Spectrometry (HPLC-MS/MS)

[Objectives]A high performance liquid chromatography-tandem mass spectrometry(HPLC-MS/MS)method was established for the determination of 14β-receptor agonist residues in mutton.[Methods]Samples were hydrolyzed byβ-glucuronidase and extracted with 5%acetic acid-acetonitrile(1:99,V/V)solution.An Eclipse plus C 18 column was used for separation,and the MRM mode was used for qualitative analysis,and the external standard method was used for quantitative analysis of matrix standard solutions.[Results]Under the optimal conditions,the retention time of the 14 kinds ofβ-receptor agonists ranged from 1.0 to 9.5 min.When the mass concentration was in the range of 0.05-0.50μg/ml,the linear relationship ofβ-receptor agonists was good,with correlation coefficients(r)≥0.9992.The detection limits of the method were in the range of 0.04-0.87μg/kg,and the quantitative limits were in the range of 0.35-1.86μg/kg.The average recovery values were in the range of 82.8%-108.9%,with RSDs(n=6)in the range of 1.9%-6.7%.[Conclusions]The method is simple,sensitive,reproducible,accurate,and can be used for simultaneous determination of the 14 kinds ofβ-receptor agonist residues in mutton.

Determination of Oleuropein in Cosmetics by Ultra Performance Liquid Chromatography-Triple Quadrupole Tandem Mass Spectrometry

An ultrahigh performance liquid chromatography-triple quadrupole tandem mass spectrometry(UPLC-MS/MS)was established to quickly and accurately determine the content of oleuropein in cosmetics.The samples were extracted with methanol-aqueous solution,and the mobile phase with methanol-formic acid solution(0.1 mol/L)=40∶60 was separated by Agilent ZORBAX Eclipse Plus C18(2.1 mm×50 mm×1.8μm-Micron)column temperature 30℃,flow rate 0.3 mL/min.The MS end was detected by electrospray negative mode ionization(ESI-)and multiple reaction monitoring(MRM)mode.The results show a good linear relationship in the range of 0.002~5 mg/L,with a correlation coefficient R2 of 0.999,5.Method recovery range from 84.2%~107.6%and the relative standard deviation RSD is 5.8%.The detection time is 5 min,the detection limit is 0.000,6 mg/L,and the limit of quantification is 0.002 mg/L.This method has the advantages of convenient operation,low quantification limit,high precision and good repeatability,and is suitable for measuring the content of oleuropein in many kinds of cosmetics.

Analysis of ganciclovir and its related substances using high performance liquid chromatography and liquid chromatography-mass spectrometry methods

Objective High performance liquid chromatography(HPLC)and liquid chromatography-mass spectrometry(LC/MS)methods were developed for the determination of ganciclovir and its related substances.Methods A Hypersil ODS2 column(4.6 mm×250 mm,5 μm)was used with a mobile phase of 0.02 M potassium dihydrogen phosphate buffer(pH 6.0)-methanol(92∶8)at a flow rate of 1.0 mL/min,and UV detector set at 254 nm was used for monitoring the eluents.Results The method was simple,rapid,selective and capable of separating all related substances at trace level with a detection limit of 0.04 μg/mL.It has been validated with respect to accuracy,precision,linearity,and limits of detection and quantification.The linearity range was 10.2-153.0 μg/mL with r=0.9998.The percentage recoveries ranged from 96.7% to 101.6%,and RSD was 1.24%-1.96%(n=5).Conclusion The method was found to be suitable not only for monitoring the reactions during the process development but also for quality control of ganciclovir.For identification of related substances,LC/MS was used.The mainly related substances of ganciclovir active pharmaceutical ingredients(API)were determined as guanine,(1,3-dioxolan-4-yl)methyl acetate,and diacetyl guanine.

Screening and Identifying of Nephrotoxic Compounds in Lithospermum erythrorhizon Using Live-cell Fluorescence Imaging and Liquid Chromatography Coupled with Tandem Mass Spectrometry

In order to identify the potential nephrotoxic compounds in traditional Chinese medicine Lithospermum erythrorhizon,it was separated into serial fractions according to their polarities.An in vitro method was utilized to determine the nephrotoxicity of these fractions with the help of fluorescence image analysis.As a result,the primary fraction A05 and its secondary fractions C06 ＂C09 and C12 ＂C14 were found to have significant toxicity to LLC-PK1 cell line,as determined by the survive rate less than 20% after they were treated with these fractions.These potential nephrotoxic fractions were further analyzed by multistage and high resolution mass spectrometry.The main compounds in these fractions were tentatively identified to be acetylshikonin,isobutyrylshikonin,β,β＇-dimethyla-cryloylshikonin,and isovalerylshikonin,which may bring nephrotoxicity.

Simultaneous Determination of Tetramethylpyrazine and Aspirin in a New Compound Formulation by Liquid Chromatography

Aim To establish a reversed-phase liquid chromatographic (LC) method forsimultaneous determination of tetramethylpyrazine (TMP) and aspirin in a new compound formulation.Methods Chromatographic separation of the two drugs was achieved on a Diamonsil C_(18) column, usinga binary mixture of methanol-1.5% acetic acid (35:65, V/V, pH = 3.1) as mobile phase at a flow rateof 1.0 mL·min^(-1). Results Separation was completed in less than 12 min. Benzoic acid was used asthe internal standard. Recoveries at levels corresponding to 80 % to 120 % of the label claim ofthe formulation ranged from 99.6 to 100.3 % for aspirin and from 99.9 to 101.3% for TMP. The linearrange was 12.6 - 150.9 μg·mL^(-1)(r= 0.9997, n = 5) for aspirin and 25.0- 300.0 μg·mL^(-1) (r =0.9999, n = 5) for TMP. Conclusion The method developed can be used for the simultaneousdetermination of TMP and aspirin in pharmaceutical preparations.

High Performance Liquid Chromatography-Electrospray Ionization-Mass Spectrometric Analysis of Bilobalide and Ginkgolides in Ginkgo biloba L. Leaves

The ginkgo terpenoids including bilobalide and ginkgolides are the main pharmaceutical components in the leaves or extracts of Ginkgo biloba L. In this paper, the analysis of bilobalide and ginkgolides in leaves of Ginkgo biloba L. by high performance liquid chromatography (HPLC)-electrospray ionization (ESI)-mass spectrometry (MS) was carried out. The separation was performed on Inertsil ODS3 column with methanol-water (36:64) as mobile phase, with 1 mL·min -1 of flow rate at 35℃. Then the mass spectrum analysis was conducted by ZMD micromass electrospray ionization (ESI)-mass spectrometer (MS). The HPLC total ion chromatogram and selected ion chromatogram (with 325, 407, 423, 439 of m/z) of the sample and ESI-/MS mass spectra of the peaks in the chromatograms were obtained. So bilobalide, ginkgolide A, B, C and J in Ginkgo biloba L. leaves were identified. The method is easy and rapid, with a good accuracy.

Novel method for the determination of five carbamate pesticides in water samples by dispersive liquid-liquid microextraction combined with high performance liquid chromatography

A novel method for the determination of five carbamate pesticides （metolcarb, carbofuran, carbaryl, isoprocard and diethofencard） in water samples was developed by dispersive liquid-liquid microextraction （DLLME） coupled with high performance liquid chromatography-diode array detector （HPLC-DAD）. Some experimental parameters that influence the extraction efficiency were studied and optimized to obtain the best extraction results. Under the optimum conditions for the method, the calibration curve was linear in the concentration range from 5 to 1000 ng mL^-1 for all the five carbamate pesticides, with the correlation coefficients （r^2） varying from 0.9984 to 0.9994. Good enrichment factors were achieved ranging from 80 to 177- fold, depending on the compound. The limits of detection （LODs） （S/N = 3） were ranged from 0.1 to 0.5 ng mL^-1. The method has been successfully applied to the analysis of the pesticide residues in environmental water samples.

Dispersive Liquid-liquid Microextraction Combined with High-performance Liquid Chromatography for the Determination of Clozapine and Chlorpromazine in Urine

A simple method has been proposed for the determination of clozapine （CLZ） and chlorpromazine （CPZ） in human urine by dispersive liquid-liquid microextraction （DLLME） in combination with high-performance liquid chromatography-ultraviolet detector （HPLC-UV）. All important variables influencing the extraction efficiency, such as pH, types of the extraction solvent and the disperser solvent and their volume, ionic strength and centrifugation time were investigated and optimized. Under the optimal conditions, the limit of detection （LODs） and quantification （LOQs） of the method were 13 and 39 ng/mL for CLZ, and 2 and 6 ng/mL for CPZ, respectively. The relative standard deviations （RSDs） of the targets were less than 5.1% （C=0.100 μg/mL, n=9）. Good linear behaviors over the tested concentration ranges were obtained with the values of R20.999 for the targets. The absolute extraction efficiencies of CLZ and CPZ from the spiked blank urine samples were 98.3% and 97.8%, respectively. The applicability of the technique was validated by analyzing urine samples and the mean recoveries for spiked urine samples ranged from 93.3% to 105.0%. The method was successfully applied for the determination of CLZ and CPZ in real human urine.

Studies on Chromatography Fingerprint of Hongqi by High-performance Liquid Chromatography

Chromatography fingerprint (CFP) of 10 samples of hongqi were studied. 23 common peaks were analyzed, their average similarity was 97.29%. CFP were positioned with main index composition such as formononetin, calycosin and then the contents of index composition were determined. The character and exclusive of CFP of 10 samples of hongqi were clear. CFP and content determination of index composition of hongqi could be used to evaluate the quality of hongqi comprehensively.

Bio-analytical method development and validation of Rasagiline by high performance liquid chromatography tandem mass spectrometry detection and its application to pharmacokinetic study

The most suitable bio-analytical method based on liquid liquid extraction has been developed and validated for quantification of Rasagiline in human plasma. Rasagiline-13C3 mesylate was used as an internal standard for Rasagiline. Zorbax Eclipse Plus C18 （2.1 mm × 50 mm, 3.5 um） column provided chromatographic separation of analyte followed by detection with mass spectrometry. The method involved simple isocratic chromatographic condition and mass spectrometric detection in the positive ionization mode using an API-4000 system. The lotal run time was 3.0 min. The proposed method has been validated with the linear range of 5 12000 pg/mL for Rasagiline. The intra-run and inter-run precision values were within 1.3% 2.9% and 1.6% 2.2% respectively for Rasagiline. The overall recovery for Rasagiline and Rasagiline-13C3 mesylate analog was 96.9% and 96.7% respectively. This validated method was successfully applied to the bioequivalence and pharmacokinetic study of human volunteers under fasting condition.

Determination of trans-resveratrol in mouse liver by high performance liquid chromatography

To develop a sensitive high performance liquid chromatography （HPLC） assay for the determination of trans-resveratrol in mouse liver. The whole liver of a mouse was removed from the body, homogenated, and extracted by ethyl acetate. The organic layer was isolated and evaporated to dryness, the residue was reconstituted in 0.2 mL mobile phase for centrifugation, and 50 uL of the supernatant was injected into the/-IPLC instrument. The sample was separated on a Shimadzu ODS column （150 mm × 4.6 mm, 5 um） at 35 ℃ and detected by ultraviolet （UV） detector at the wavelength of 305 nm. The mobile phase consisted of methanol and 0.1 mol/L acetic acid （4：6, v/v） with the flow-rote at 1 mL/min. The limit of detection was 3.0 ng/g in liver homogenate with a signal/noise ratio of 3：1. The linear range of the calibration curve was 5.0-120.0 ng/g. The mean recoveries at the concentrations of 6, 10 and 80 ng/g were 102%, 96.0% and 91.5%, respectively. The RSDs for inter- and intra-day assays were less than 5%. Compared with other reported methods, this method was faster and more sensitive. It was also proved to be of good linearity, selectivity, accuracy and precision, and can be efficiently applied to the pharmacoldnetic study of trans-resveratrol in mouse liver.

Simultaneous determination of 15 pesticide residues in Chinese cabbage and cucumber by liquid chromatography-tandem mass spectrometry utilizing online turbulent flow chromatography

In this experiment,a liquid chromatography tandem mass spectrometry method was built to determine 15 pesticide residues in Chinese cabbage and cucumber samples based on online turbulent flow chromatography purification.After modified quick,easy,cheap,effective,rugged,and safe(QuEChERS)extraction,extracts were directly injected to the TLX(TurboFlow Liquid Xcalibur)system and brought to TurboFlow™columns for on-line purification and then transferred to analytical column for further separation and analysis.TurboFlow™columns types,transfer flow rate,and transfer time were optimized.Limits of detection and limits of quantification of the method obtained for 15 pesticide residues were ranged between 0.2–1.0μg/kg and 0.5–2.0μg/kg in Chinese cabbage and cucumber samples.Recoveries of pesticide residues were in range of 75.3%–103.7%.Matrix effects for 15 pesticides were in range of 5.6%–106.6%.The developed method has been successfully used for the determination of 15 pesticide residues in real samples.

Determination of Steroid Hormone Residues in Fish Tissues Using Gel Permeation Chromatography and Ultra Performance Liquid Chromatography-Tandem Mass Spectrometry

A highly sensitive method was developed for the simultaneous determination of 8 steroid hormones in high-fat fish tissues using ultra high performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS).The 8 steroid hormones were extracted from the tissues with diethyl ether.Differing from other common purification methods,the extract solutions were cleaned by gel permeation chromatography(GPC) using ethyl acetate-cyclohexane solution(1:1,v/v) as the mobile phase.The separation of target compounds was carried out by a BEH C18 column and a gradient elution consisting of acetonitrile and 0.2% aqueous formic acid(v/v).The compounds were detected under the multiple reaction monitoring(MRM) mode and quantified with external standard method.This method was validated with respect to linearity,specificity,accuracy and precision.A linearity with correlation coefficient larger than 0.995 was achieved in the range of 0.5 to 50 ng m L^(-1).The average recoveries at the spiked levels of 1.0,5.0,and 10.0 μg kg^(–1) varied between 81.7% and 90.8%,with the relative standard deviations(n=5) ranged from 3.50% to 10.0%.The limit of quantification(LOQ) for 8 steroid hormones ranged from 0.2 to 1.5 μg kg^(-1).It was concluded that this method can be successfully applied for the determination of 8 steroid hormones in complicated matrices including high-fat fish tissues.

Comparison of liquid-liquid extraction-thin layer chromatography with solid-phase extraction-high-performance thin layer chromatography in detection of urinary morphine

Liquid-liquid extraction-thin layer chromatography （LLE-TLC） has been a common and routine combined method for detection of drugs in biological materials. Solid-phase extraction （SPE） is gradually replacing the tra- ditional LLE method. High performance thin layer chromatography （HPTLC） has several advantages over TLC. The present work studied the higher efficiency of a new SPE-HPTLC method over that of a routine LLE-TLC method, in extraction and detection of urinary morphine. Fifty-eight urine samples, primarily identified as mor- phine-positive samples by a strip test, ＇were re-screened by LLE-TLC and SPE-HPTLC. The results of LLE-TLC and SPE-HPTLC were then compared with each other. The results showed that the SPE-HPTLC detected 74% of total samples as morphine-positive samples whereas the LLE-TLC detected 48% of the same samples. We further discussed the effect of codeine abuse on TLC analysis of urinary morphine. Regarding the importance of morphine detection in urine, the present combined SPE-HPTLC method is suggested as a replacement method for detection of urinary morphine by many reference laboratories.