PTP1B And ALR2Targeted In Screening Drugs

Diabetes Mellitus and Diabetic chronic complications had been the fifth fataldisease. Recently, the number of Diabetes Mellitus patients is increasing year byyear, and the age of the patients is continually decreasing. Diabetes Mellitus hasbeen a serious disease which has an impact on the quality of daily life. However, theexisting drug against the Diabetes Mellitus has obvious defects, such as dose-dependent, serious side effects and so on. Therefore finding the suitable treatment ofDiabetes Mellitus has become a top priority in the area of drug development. Wehave studied the Diabetes Mellitus and Diabetic chronic complications targets:Protein Tyrosine Phosphatase-1B and Aldose reductase.We constructed humanized ALR2plasmid with the PET-30a vector and expressedin E. coli BL21(DE3). We got the purified cloning ALR2with ion exchanged anddynamic tracking, and the purified ALR2could be used for inhibitor screening. Thestudy of the ferment nature indicated that when substrate was DL-glyceraldehyde, themost appropriate pH value of ALR2was6.2, the most optimal reaction temperaturewas37oC, and the most optimal ion intensity was0.3mol/L, Km=0.265mmol/L,vMAX=0.064μmol·L-1·min-1.We screened the inhibitors against the target PTP1B and ALR2in vitro fromdiverse traditional Chinese medicine. We found that Illicium difengpi extract wasan efficient competitive inhibitor of PTP1B and ALR2with the IC50of59.32μg/mLand60.25μg/mL.We analyzed the active ingredients of Illicium difengpi with the ultrafiltrationmass spectrometry. HPLC–UV analysis was conducted with a C18column (150mm×4.6mm，5μm). The flow rate was0.5mL/min. The volume of injection for LC–MS analysis was10μL and the wavelength for detection was254nm. Themobile phase consisted of acetonitrile (A) and deionised water with0.5%(v/v)acetic acid(B). The gradient elution was set as follows:0-20min20%A-20%A；20-35min20%A-38%A；35-50min38%A-45%A;50-55min45%A-60%A；55-65min60%A-75%A；65-70min75%A-85%A；70-80min85%A-100%A；80-90min100%A-100%A.We have found7compounds binding with PTP1B and ALR2. Five of the sevencompounds have the same fragments carbonyl. One of the other two compounds hasthe carboxyl and another one cannot be analyzed only by MSn. We have concludedthat the compounds with carbonyl group on the hexatomic ring maybe have thepotential of the efficient inhibitor of PTP1B and ALR2. And furfuran besides maybe an advantage factor.