Liquid Chromatography-electrospray Ionization Tandem Mass Spectrometry for Simultaneous Determination of Metformin and Glimepiride in Beagle Dog Plasma and Bioequivalence Study

A sensitive and selective liquid chromatography-electrospray ionization tandem mass spectrometry（LC- ES1-MS/MS） was used for the simultaneous determination of metformin and glimepiride in beagle dog plasma with glipizide as internal standard（IS）. After simplified protein precipitation with methanol, both the analytes and IS were chromatographed on a Zorbax CN column via gradient elution with methanol（containing 5 mmol/L ammonium ace- tate） and 5 mmol/L aqueous ammonium acetate as the mobile phase. Detection was performed by multiple reaction monitoring（MRM） scanning via ESI source operated in positive ionization mode. Specificity, linearity, accuracy, pre- cision, recovery, matrix effect and stability were validated for metformin and glimepiride in beagle dog plasma. The calibration curves were linear in a concentration range of 10-10000 ng/mL for metformin and 4-4000 ng/mL for glimepiride with both correlation coefficients higher than 0.99. The recoveries obtained for the analytes and IS were all between 82.7% and 101.2%. The method exhibited excellent performance in terms of selectivity, robustness, short analytical time and simplicity of sample preparation. Finally, the proposed method was applied to a bioequivalence study of self-made bilayer tablet and commercial formulation containing 500 mg of metformin and 1 mg of glimepi- ride in beagle dogs.

Simultaneous determination of seven flavonoids in Epimedium by liquid chromatography-tandem mass spectrometry method

In this paper, a sensitive and specific liquid chromatography-electrospray ionisation-mass spectrometry （LC-ESI-MS） method has been developed and validated for the identification and determination of seven flavonoids, namely epimedin A, epimedin B, epimedin C, icariin, sagittatoside B, 2＂-O-rhamnosyl icariside II, and baohuoside I in Epimedium from different sources.

LC-ESI-MS/MS,a modified method for simultaneous quantification of isoflavonoids,flavonoids,alkaloids and saponins in a Chinese herbal preparation Gegen-Qinlian decoction

A sensitive and reliable liquid chromatography-electrospray ionisation-tandem mass spectrometry(LC-ESI-MS/MS)method was established to simultaneously quantitate four categories of compounds(isoflavonoids,flavonoids,alkaloids and saponins)in Gegen-Qinlian decoction(GQD).These compounds were separated by a Shiseido CAPCELL PAK C18 column with a linear gradient consisting of 0.1%(v/v)formic acid in water(A)and 0.1%(v/v)formic acid in acetonitrile(B),and delivered at a flow rate of 0.3 mL/min.All the analytes were determined by electrospray positive ionization tandem mass spectrometry in a multiple reaction monitoring(MRM)mode.Linearity,accuracy,precision,recovery and stability of the method were evaluated with the validation over the range of 4.0-538 5 ng/mL.The proposed method was applied to the analysis of a Chinese herbal preparation GQD successfully.

Investigation of Breaking Efficiency of N-Glycosidic Bond Between Sulfonamide and Honey

Sulfonamide residue in honey existed in a form bonded to sugar via the N-glycosidic bond.It would result in the possible underestimation of concentration of sulfonamide if it is not decomposed by chemical methods.However,in China's Mainland and Taiwan（P.R.China）,the regulation for sulfonamide residue analysis does not include hydrolysis and has been applied to a very broad range of samples,for example,egg,milk,meat,seafood as well as honey.This paper demonstrates the necessity of hydrolysis of it prior to extracting honey.The breaking efficiencies of N-glycosidic bond were investigated by 2 mol/L hydrochloric acid,pure methanol and 0.5 mol/L hydrochloric acid in methanol,respectively.It was found that acid plays the key role in breaking the N-glycosidic bond,and it was also noticed that the dissolution of liberated sulfonamide in methanol could carry the reaction forward in favor of breaking the N-glycosidic bond.

Chemical profiling of principle active and toxic constituents in herbs containing aristolochic acids

Objective:To clear the amounts of the principal active/toxic components in herbs containing aristolochic acids(HCAAs),which are still used as medicine and/or seasoning in many ethnic minority areas of China.Methods:In this study,six major active and toxic components in HCAAs were extracted with ultrasonic extraction.With 6-O-methyl guanosine as internal standard,the target compounds were analyzed qualitatively and quantitatively by using ultrahigh performance liquid chromatography-electrospray ionization-tandem mass spectrometry(UPLC-ESI-MS/MS)with multiple reaction monitoringinformation dependent acquisition-enhanced production ion scanning mode(MRM-IDA-EPI)combined with dynamic background subtraction(DBS)function.Results:The method showed good linearity in the linear range of the six analytes.The limit range of detection was from 0.01 ng/mL to 0.27 ng/mL.All of the detection repeatability,extraction repeatability and accuracy of the method were good.After extraction,the samples remained stable at 15℃ within 24 h.Six analytes were all found in samples except aristolactam(AL)in sample 2,and the contents varied greatly.The contents of these compounds decreased in fruits,leaves and stems of Aristolochia delavayi successively.Conclusion:This method has the advantages of less sample dosage,simple operation,short analysis cycle,high sensitivity,specificity and accuracy.It laid a good foundation for guiding the safety of HCAAs,the indepth study of pharmacological and toxicological effects and the scientific and standardized processing and compatibility of HCAAs.

HPLC-ESI-MS Analysis of Flavonoids Obtained from Tissue Culture of Dracaena cambodiana

Dragon＇s blood is a traditional medicine and used in many countries with different cultures because of its various therapeutic properties. The main bioactive constituents of dragon＇s blood are flavonoids, which exhibit various pharmaceutical activities, such as haemostatic, analgesic, anticoagulant, and antimicrobial activities and so on, and have attracted the attention of researchers on the development of new drugs. However, the formation process of dragon＇s blood in nature is very slow, which cannot meet the demand of pharmaceutical uses. Dracaena cambodiana （D. cambodiana） is one of the resource plants of dragon＇s blood na, 6-benzylaminopurine was added in the medium, then red During the course of tissue culture of D. cambodiasecretion was discovered in the cultured medium. Twelve compounds（1--12）, including 11 flavonoids, were determined via the analysis of constituents of the cultured medium by high performance liquid chromatography-electrospray ionization-mass spectrometry（HPLC-ESI-MS） and the results were compared with that of the standards isolated from dragon＇s blood. Among compounds 1-12, 4,4＇- dihydroxy-2-methoxydihydrochalcone（5）, 4,4＇-dihydroxy-2,6-dimethoxydihydrochalcone（6） and （2S）-7,3＇-dihydroxy- 4＇-methoxyflavane（9） were major compounds of the red secretion, with contents of 15.70, 7.10 and 57.23 mg/L, respectively. Therefore, it is promising that flavonoids from dragon＇s blood can be obtained from the tissue culture of its resource plants for the purpose of drug development.

New insights on the utilization of ultrasonicated mustard seed cake:chemical composition and antagonistic potential for root-knot nematode,Meloidogyne javanica

This study focused,for the first time,on the effect of ultrasonic features on the extraction efficiency of secondary metabolites in mustard seed cake(MSC).The nematostatic potential of sonicated seed cake was examined against the second-stage juveniles(J2s)of root-knot nematode,Meloidogyne javanica.The results show that a 35 ppm(parts per million)concentration of a sonicated extract(SE)sample of MSC caused 65%J2s mortality at 18 h exposure period in vitro.It also significantly suppressed the root-knot index(RKI=0.94)in tomato roots.The lethal concentration values for SE were 51.76,29.79,and 13.34 ppm,respectively,at 6,12,and 18 h of the exposure period,and the lethal concentration values for the non-sonicated extract(NSE)sample were 116.95,76.38,and 55.59 ppm,respectively,at similar exposure time.Sinapine and gluconapin were identified as the major compounds in ultrasonic-assisted MSC.Because of the high extraction efficiency of metabolites in the SE,all treat‐ments of SE were shown to be antagonistic to J2s.Thus,this study of ultrasonication activity-based profiling of MSC may help generate target-based compounds at a scale relevant to the control of disease caused by nematodes in economic crops.

Studies on Intestinal Transport of Ginsenoside Compatibility with Veratrum nigrum via Caco-2 Cell Monolayer Model Coupled with UPLC-ESI-MS Method

The Caco-2 cells have been recognized as effective tools to be applied to imitating the drug absorption in human intestine for the transport of drug. Herein, Caco-2 cell monolayer model was used to study the transport of the ginsenoside compatibility with Veratrum nigrum in different proportions. A specific high performance liquid chro- matography-electrospray ionization-mass spectrometry（HPLC-ESI-MS） method was developed for the semiquantita- tive determination of ginsenoside in intestinal transport with Dioscin as an internal standard. For the Caco-2 model constructed, two influencing factors were investigated, including time and concentration. The results suggest that the absorption of ginsenoside Re, Rgl, Rbl, Rc, Rb2 and Rd are time- and concentration-dependent and the excretions of Rbl, Rc, Rb2 and Rd have a relatronship with some transport proteins. The bioavailability of the ginsenosides has reduced compared to the single Panax ginseng extract when compatibility with a certain amount of Veratrum nigrum.

Pheromone 3kPZS evokes context-dependent serotonin sexual dimorphism in the brain of the sea lamprey (Petromyzon marinus)

Mature male sea lampreys(Petromyzon marinus)release a sex pheromone,3-keto-petromyzonol sulfate(3kPZS),that induces sexually dimorphic behavioral responses in conspecifics.However,the neural mechanism of such responses is mostly unknown.We examined the neurotransmitter serotonin(5-hydroxytryptamine,5-HT)and the expression of 5-HT1A receptors in the forebrain and brainstem of sea lamprey exposed to the vehicle(0.91 ppm methanol)or 10−10 M 3kPZS for 2 h using high performance liquid chromatography-electrospray ionization tandem mass spectrometry,immunohistochemistry and real-time quantitative polymerase chain reaction.Exposure to 3kPZS for 2 h increased 5-HT concentration in the forebrain of adult females,whereas 5-HT was not detected in the forebrain of adult males.On the contrary,3kPZS exposure decreased 5-HT concentration in the brainstem of adult females and had no effect in adult males.Pheromone exposure evoked context-dependent sexual dimorphism in brain 5-HT1A receptor immunoreactivity,but had no effect on 5-HT1A mRNA concentrations in the brain with 2 h exposure time.It appears that in sea lamprey pheromone 3kPZS affects the 5-HT system in the brain in a context-dependent,sexually dimorphic manner.