Dertermination Of Reduced Glutathione (GSH) And Thiols In Proteins(P-SH)in Fruit And Vegetable Seeds

Organisms contain a variety of antioxidants,which thiol compounds of containing cysteine residues played an important role in maintaining oxidation reduction atmosphere in organism.According to reports in the literature,the level of thiol compounds in organism is closely linked with many diseases,its content has become a sign of many pathological judgment.Due to the accelerated the pace of life,most people are often in a state of high pressure,it also stimulate the increase of reactive oxide species in the human body,which accelerate the aging of the body,at the same time,the excessive reactive oxide species are also easy to induce genetic mutation and cause malignant diseases such as cancer.According to study,the human body through intaking exterior antioxidant,such as reduced glutathione(GSH),N-acetylcysteine,can obviously improve the level of GSH content in human blood,which plays an important role to improve the human medical treatment.Therefore,exploiting an accurate,sensitive,economic analysis method to detect the content of thiol compounds of containing cysteine residues rapidly,and to screen biological samples in rich of thiol compounds as to exterior antioxidant supplements which has practical significance for human medical treatment,disease prevention.This research using 5,5'-dithio-bis-2-nitrobenzoic acid(DTNB)as derivatizing agent that selective and quantitative generate derivatization product.The contents of reduced glutathione(GSH)and protein thiols in seeds of vegetable and fruit were detected by pre-column derivatization HPLC and spectrometry method with concomitant cell rupture sample pretreatments and screening four seeds in rich of GSH and protein thiols.HPLC determination results showed that detected seeds in grapefruit,pumpkin,cucumber,wax gourd contained relatively abundant GSH and thiols in P-SH.The amounts detected are:GSH=1.7304?mol/g,P-SH=1.2028 ?mol/g for grapefruit seed,GSH=1.0445 ?mol/g,P-SH=1.8878 ?mol/g for pumpkin seed respectively.However,GSH was not detected in Cucumber seed and in wax gourd's seeds which contain relatively abundant protein thiols,The amounts detected are 4.0325 ?mol/g and 4.1623 ?mol/g respectively.Samples are detected by Fluorescence and UV spectroscopy reported in literatures,founded that determination results deviate slightly chromatographic results,show the sample may have certain spectral interferences.In this study,the recoveries of the HPLC method are 99.92%-101.32%,and detection limits are 0.1386 ?mol/g for GSH and 0.2184 ?mol/g for protein thiols,basic Conform to the accurate measurement requirements.Due to the complexity of biological sample composition,these fruit and vegetable seeds contain plenty of oil flavonoids,vitamin and minerals,etc.In order to explore other chemicals in the fruit and vegetable seed influence the determination results of the experiment,grapefruit seed and pumpkin seeds as samples in this study,the interference of flavonoids and Fatty acid detected using HPLC and spectrometry respectively.The results show that;it is bigger to detected the content of thiol compounds by the interference of flavonoids using spectrometry,but its content is detected by HPLC was no significant influence;At the same time,Fatty acid in the sample without interference or interference is relatively small.From what has been discussed above,this research provides important analytical data for further screening biological samples in rich of thiol compounds from plants using accurate analysis method as well as the content of thiol compounds in fruit and vegetables.