Comprehensive analysis of insulin products complex disulfide bonds structure by high resolution mass spectrum / 药学学报

The correct pairing of disulfide bonds maintains the correct folding mode and high-level structure formation of peptides and protein drugs, which is crucial for the quality control of products. In order to ensure that the disulfide bonds are correctly paired, disulfide bond analysis is an essential part of peptides and protein drug characterization. Mass spectrometry can be used to analyze disulfide bonds. However, insulin and its analogues have two pairs of disulfide bonds without restriction enzyme cutting site. Conventional collision-induced dissociation (CID) and high-energy induced cleavage (HCD) cannot accurately locate the complex disulfide bond. In our study, three methods were used to localize the complex disulfide, including enzyme digestion combined with key peptide fragment in source decay (ISD) fragmentation method, enzyme digestion combined with partial reduction alkylation method, intact protein source ISD and electron transfer dissociation (ETD) cleavage method, The applicability of insulin aspart, insulin lispro and insulin glargine were also investigated. This study provides a new way for the quality control of disulfide bonding mode of insulin and its analogues, and also provides a reference for the disulfide bond localization of peptides or proteins containing this complex disulfide bond.

Effect and mechanism of leonurine on pressure overload-induced cardiac hypertrophy in rats / 中国中药杂志

To investigate the effects of leonurine(Leo) on abdominal aortic constriction(AAC)-induced cardiac hypertrophy in rats and its mechanism. A rat model of pressure overload-induced cardiac hypertrophy was established by AAC method. After 27-d intervention with high-dose(30 mg·kg~(-1)) and low-dose(15 mg·kg~(-1)) Leo or positive control drug losartan(5 mg·kg~(-1)), the cardiac function was evaluated by hemodynamic method, followed by the recording of left ventricular systolic pressure(LVSP), left ventricular end-diastolic pressure(LVESP), as well as the maximum rate of increase and decrease in left ventricular pressure(±dp/dt_(max)). The degree of left ventricular hypertrophy was assessed based on heart weight index(HWI) and left ventricular mass index(LVWI). Myocardial tissue changes and the myocardial cell diameter(MD) were measured after hematoxylin-eosin(HE) staining. The contents of angiotensin Ⅱ(AngⅡ) and angiotensin Ⅱ type 1 receptor(AT1 R) in myocardial tissue were detected by ELISA. The level of Ca~(2+) in myocardial tissue was determined by colorimetry. The protein expression levels of phospholipase C(PLC), inositol triphosphate(IP3), AngⅡ, and AT1 R were assayed by Western blot. Real-time quantitative PCR(qRT-PCR) was employed to determine the mRNA expression levels of β-myosin heavy chain(β-MHC), atrial natriuretic factor(ANF), AngⅡ, and AT1 R. Compared with the model group, Leo decreased the LVSP, LVEDP, HWI, LVWI and MD values, but increased ±dp/dt_(max) of the left ventricle. Meanwhile, it improved the pathological morphology of myocardial tissue, reduced cardiac hypertrophy, edema, and inflammatory cell infiltration, decreased the protein expression levels of PLC, IP3, AngⅡ, AT1 R, as well as the mRNA expression levels of β-MHC, ANF, AngⅡ, AT1 R, c-fos, and c-Myc in myocardial tissue. Leo inhibited AAC-induced cardiac hypertrophy possibly by influencing the RAS system.

Correlation analysis between foliar endophytic fungi of Salvia miltiorrhiza and effective components / 中国中药杂志

The aim of this study was to comprehensively investigate the correlations between foliar fungal endophyte communities and effective components accumulations in Salvia miltiorrhiza. Foliar samples of S. miltiorrhiza were collected in 5 different areas. Their fungal endophyte communities and effective component contents were determined by denaturing gradient gel electrophoresis (DGGE) and high performance liquid chromatography (HPLC), respectively. The results showed that, for characteristics of foliar fungal endophyte communities and effective component contents, there were both similarities and differences among the five samples. Correlation analysis of DGGEs' band and 24 effective components revealed a significant correlations (P < 0.01). For examples, 4 bands (15, 18, 23 and 26) were all significantly correlated with the accumulations of caffeic acid, salvianolic acid B, salvianolic acid C and dihydrotanshinone I.