Enantioseparation and quantitative analysis of three alkaloids in Rhizoma Corydalis by chiral LC-MS/MS.

A chiral liquid chromatography-mass spectrometry (LC-MS/MS) approach was developed and verified for the first time in order to quantify the three alkaloids, namely tetrahydropalmatine (THP), tetrahydroberberine (THB) and tetrahydrocoptisine (THC) in Rhizoma Corydalis. Solid-phase extraction was used to prepare sample solution. Enantiomeric separation was obtained on a Chiralpak IC column using acetonitrile-water-ammonia (90:10: 0.1, v/v/v) as mobile phase. The detection was carried out in multiple reaction monitoring (MRM) mode with positive electrospray ionization source. Transitions of m/z 356.0→192.0, m/z 340.0→176.0 and m/z 324.1→176.0 were monitored for THP, THB and THC respectively. All calibration curves showed good linearity (r2 ≥0.9994) within the test ranges. The lower limit of quantification (LLOQ) was 1.0 ng mL-1 and 1.5 ng mL-1 for (+)- and (-)-THP, 1.0 ng mL-1 and 3.0 ng mL-1 for (+)- and (-)-THB, 1.5 ng mL-1and 1.8 ng mL-1 for (+)- and (-)-THC, respectively. The precision, repeatability, and stability tests all meet the requirements. The average recoveries of the analytes ranged from 98.9 % to 101.3 %. Ultimately, the established method was successfully applied to the quantitative analysis of these alkaloid enantiomers extracted from Rhizoma Corydalis of different habitats.

Association Between LOX-1, LAL, and ACAT1 Gene Single Nucleotide Polymorphisms and Carotid Plaque in a Northern Chinese Population.

Objective: Carotid atherosclerosis is one of the major risk factors for ischemic stroke. The presence of carotid plaque has been widely used to assess the risk of clinical atherosclerotic disease. Lectin-type oxidized LDL (low-density lipoprotein) receptor 1 (LOX-1), lysosomal acid lipase (LAL), and acyl-CoA:cholesterol acyltransferase 1 (ACAT1) are important for lipid accumulation in atherosclerosis. The objective of this study was to investigate the relationship between single nucleotide polymorphisms (SNPs) in the LOX-1, LAL, and ACAT1 genes and the presence of carotid plaque in a Northern Chinese population. Methods: Three polymorphisms in LOX-1 (rs1050286), LAL (rs11203042), and ACAT1 (rs11576517) were identified and genotyped in 215 patients with carotid plaque and 252 controls using the polymerase chain reaction with high-resolution melting analysis. Results: The LOX-1 (rs1050286) AA and LAL (rs11203042) TT genotypes were significantly associated with increased risk of carotid plaque, whereas a ACAT1 (rs11576517) TT genotype was shown to be protective against carotid plaque in a Northern Chinese population (p < 0.05). Even after the Bonferroni correction, the LAL (rs11203042) TT genotype (odds ratio = 3.838, 95% confidence interval = 1.748-8.426, p < 0.001) was still associated with an increased risk for carotid plaque. Conclusions: These results suggest that the LAL (rs11203042) TT genotype is associated with increased risk for carotid plaque in a Northern Chinese population, and that the LOX-1 (rs1050286) AA genotype shows a nonstatistically significant trend towards association. However, no association was found between the ACAT1 (rs11576517) polymorphisms and carotid plaque presence.

The construction of a panel of serum amino acids for the identification of early chronic kidney disease patients.

BACKGROUND: Serum creatinine, urea, and cystatin-c are standardly used for the evaluation of renal function in the clinic. However, some patients have chronic kidney disease but still retain kidney function; a conventional serum index in these patients can be completely normal. Serum amino acid levels can reflect subtle changes in metabolism and are closely related to renal function. Here, we investigated how amino acids change as renal impairment increases. METHODS: Subjects were divided into three groups by renal function glomerular filtration rate: healthy controls, patients with chronic kidney disease with normal kidney function, and patients with chronic kidney disease with decreased kidney function group. We identified 11 amino acids of interest using LC-MS/MS on MRM (+) mode. RESULTS: Statistical analysis indicated that alanine (ALA), valine (VAL), and tyrosine (TYR) decrease with renal function impairment, whereas phenylalanine (PHE) and citrulline (CIT) increase. We tried to construct a diagnostic model utilizing a combination of amino acids capable of identifying early chronic kidney disease patients. The accuracy, specificity, and sensitivity of the combining predictors were 86.9%, 84.6%, and 90.9%, respectively, which is superior to the reported values for serum creatinine, urea, and cystatin-c. CONCLUSION: Our data suggest that serum amino acid levels may supply important information for the early detection of chronic kidney disease. We are the first to establish a diagnostic model utilizing serum levels of multiple amino acids for the diagnosis of patients with early-stage chronic kidney disease.

A sensitive liquid chromatographic-mass spectrometric method for simultaneous quantification of six iridoid glycosides from Zhi-zi-chi Decoction in rat plasma and its application to a pharmacokinetic study.

A sensitive liquid chromatographic-mass spectrometric (LC-MS) method was developed and validated for simultaneous determination of geniposide, geniposidic acid, scandoside methyl ester, gardenoside, deacetyl asperulosidic acid methyl ester and genipin-1-ß-gentiobioside after oral administration of Zhi-zi-chi Decoction in rat plasma. The six iridoid glycosides were extracted from plasma samples by protein precipitation, and then separated on an Apollo C18 column (250 mm × 4.6mm, 5 µm) through the application of a gradient elution. The analytes were monitored in positive electrospray ionization by selected ion monitoring mode (SIM). The lower limits of quantitation (LLOQ) of the six analytes were all lower than 6 ng/mL. The accuracy (relative error, RE%) was between -7.0% and 9.9%, while the intra- and inter-day precisions (relative standard deviation, RSD%) were less than 6.3% and 9.8% for the six analytes, respectively. The developed method was successfully applied to a comparative pharmacokinetic study of the six iridoids in rat plasma after oral administration of Zhi-zi-chi Decoction and Gardenia jasminoides extract.

Determination of phthalate esters in polyvinyl chloride infusion bag by stir bar sorptive extraction combined with GC.

A novel, efficient, and sensitive stir bar sorptive extraction method coupled with GC for the detection of four kinds of phthalate esters in plasticized polyvinyl chloride infusion bag has been developed and validated. Some experimental parameters including stirring speed, stirring time, pH value, salt concentration, desorption mode, desorption solvents, and desorption time were investigated and optimized. Under optimum condition, the validated assay was found to be linear (r > 0.9945) and LODs were between 1.07 and 2.67 ng for the four analytes. The method exhibited excellent precision with RSD varied from 4.5 to 6.1% (n = 5). The recoveries of the four phthalate esters at two different concentrations ranged from 80.5 to 93.4%. The results showed that the validated method could meet the need of determination of targets and was successfully applied to the analysis of phthalate esters in real samples.

Identification of multiple constituents in the traditional Chinese medicine formula Zhi-zi-chi decoction and rat plasma after oral administration by liquid chromatography coupled to quadrupole time-of-flight tandem mass spectrometry.

RATIONALE: Liquid chromatography (LC) coupled to positive electrospray ionization (ESI) tandem mass spectrometry (MS/MS) employing a time-of-flight tandem mass spectrometer was established to identify multi-components of Zhi-zi-chi decoction, a traditional Chinese medicine formula, and the constituents in rat plasma after oral administration of Zhi-zi-chi decoction. METHODS: The LC separation was achieved on a C(18) column. The mobile phase consisted of acetonitrile/0.2% formic acid with gradient program. The quadrupole time-of-flight (Q-TOF) mass spectrometer was operated in the positive ion mode with an electrospray ionization source (ESI+). The capillary voltage of the ion source was set at 4500 V and the capillary exit was 90 V. The nebulizer pressure was maintained at 1.2 bar. Hexapole radio frequencies 1 and 2 were set to 200 Vpp and 250 Vpp, respectively. RESULTS: A total 47 compounds in the Zhi-zi-chi decoction and 24 constituents in rat plasma after oral administration of Zhi-zi-chi decoction were identified. Of the 47 detected compounds in the Zhi-zi-chi decoction, 15 were identified by comparing the retention time and MS data with that of reference compounds and the rest were identified by MS analysis and retrieving the reference literature. Of the identified 24 compounds in rat plasma, 19 were the original form of the compounds absorbed from the 47 detected compounds, and the other five were the metabolites of the compounds existing in the Zhi-zi-chi decoction. CONCLUSIONS: A fast and sensitive LC/Q-TOF MS method has been developed and successfully utilized to screen the active ingredients of a Chinese medical formula, Zhi-zi-chi decoction, for the first time. The results indicated that the 24 compounds identified in rat plasma were the potential active ingredients of Zhi-zi-chi decoction, which provided helpful chemical information for further pharmacology and active mechanism research on Zhi-zi-chi decoction and other traditional Chinese medicines.

A sensitive liquid chromatography-mass spectrometry method for simultaneous determination of two active chromones from Saposhnikovia root in rat plasma and urine.

A sensitive and efficient liquid chromatography-mass spectrometry method was developed and validated for the simultaneous determination of two active chromones (prim-O-glucosylcimifugin and 4'-O-D-glucosyl-5-O-methylvisamminol) from Saposhnikovia root in rat plasma and urine. The plasma or urine samples were prepared by protein precipitation. Chromatographic separation of the two active chromones from matrix interferences was achieved on an Angilent TC-C(18) column with a mobile phase consisted of methanol, water and 0.1% formic acid. Puerarin was added as the internal standard. The method was validated with the concentration range 1.0-100 ng/mL in rat plasma and 10-1000 ng/mL in urine for prim-O-glucosylcimifugin, 1.5-150 ng/mL in plasma and 15-1500 ng/mL in urine for 4'-O-D-glucosyl-5-O-methylvisamminol. The lower limit of quantitation (LLOQ) of prim-O-glucosylcimifugin and 4'-O-D-glucosyl-5-O-methylvisamminol was 1.0 and 1.5 ng/mL in plasma, 10 and 15 ng/mL in urine, respectively. The intra- and inter-day precision across three validation days over the entire concentration range was lower than 9.0% as terms of relative standard deviation (R.S.D.). Accuracy determined at three quality control concentrations (2.0, 25 and 75 ng/mL for prim-O-glucosylcimifugin; 3.0, 37.5 and 112.5 ng/mL for 4'-O-D-glucosyl-5-O-methylvisamminol) ranged from -1.9 to 3.9% as terms of relative error (R.E.). The LC-ESI-MS method was further applied to assess pharmacokinetics and urine excretion of the two chromones after oral administration of Fangfeng extract to rats. Practical utility of this new LC-MS method was confirmed in pilot pharmacokinetic studies in rats following oral administration.

Determination of nifedipine in human plasma and its use in bioequivalence study.

A sensitive atmospheric pressure chemical ionization liquid chromatographic-mass spectrometric (APCI-LC-MS) assay with positive ion mode has been developed for the determination of nifedipine in human plasma. In this method, nifedipine was extracted from human plasma using diethyl ether with dimethoxanate as the internal standard. Analysis was achieved on a BDS C(18) column with methanol-H(2)O (66:34, v/v) as the mobile phase. Sustained-release nifedipine tablets from DiSha (test, Weihai, China) and from GuoFeng (reference, Qingdao, China) were evaluated following a single 20mg oral dose to 20 healthy volunteers. Bioequivalence between the products was determined by calculating 90% confidence intervals (90% CI) for the ratio of C(max), AUC(0-t) and AUC(0-infinity) values for the test and reference products, using logarithmic transformed data. The 90% confidence intervals for the ratio of C(max) (86.6-105.2%), AUC(0-t) (97.8-110.9%) and AUC(0-infinity) (96.5-110.4%) values for the test and reference products are within the interval (80.0-125.0% for AUC, and 70-143% for C(max)), proposed by State of Food and Drug Administration [SFDA, 2005. Guidance for Bioavailability and Bioequivalence Studies for Chemical Drug Products in Human Being. China, p. 19]. It was concluded that the two sustained-release nifedipine tablets are bioequivalent in their rate and extent of absorption and, thus, may be used interchangeably.