Author/Authors :
Ghamari, M Department of Food Science and Technology - Ferdowsi University of Mashhad, Mashhad, Iran , Alemzadeh, I Department of Chemical and Petroleum Engineering - Sharif University of Technology, Tehran, Iran , Tabatabaee Yazdi, F Department of Food Science and Technology - Ferdowsi University of Mashhad, Mashhad, Iran , Vossoughi, M Department of Chemical and Petroleum Engineering - Sharif University of Technology, Tehran, Iran , Varidi, M Department of Food Science and Technology - Ferdowsi University of Mashhad, Mashhad, Iran
Abstract :
In this study, Aspergillus niger lipase after extraction of medium culture was precipitated with different percentages of acetone and purified by ion exchange chromatography using SP-sepharose HP and Q-sepharose HP. The process of purification of the anzyme was studied by electrophoresis and the molecular weight was detected and determined by Zymography using overlying containing phenol red and rhodamine B. The results show that the vast majority of lipase from this strain has been precipitated by acetone 70% saturation, and leads to the 1.67 fold the purified enzyme, with special activities 32.8 U/mg and 38.5%efficiency. Using two-phase chromatography, enzyme specific activity reached 246.47 U/mg and 12.59-fold purification were achieved. The results of Zymography and electrophoresis indicate a lipase band with weighing about 30 kDa.
Farsi abstract :
در اين تحقيق ليپاز حاصل از كپك آسپرژيلوس نايجر، پس از استخراج از محيط كشت توسط رسوب دهي با درصد هاي متفاوت استون و كروماتوگرافي تعويض يوني با استفاده از SP-sepharose HP و Q-sepharose HP خالص سازي شد. جهت بررسي روند خالص سازي از الكتروفورز و جهت رديابي آنزيم و تعيين وزن مولكولي آن از زايموگرافي با استفاده از پوشاننده حاوي فنول رد و ردامين B، استفاده شد. نتايج نشان مي دهند بخش اعظم آنزيم ليپاز اين سويه، بوسيله استون 70% اشباع رسوب مي كند كه اين امر باعث تخليص آنزيم به ميزان 1.67برابر، با فعاليت ويژهU/mg 32.8 و بازده 38.5% مي شود. با استفاده از دو مرحله كروماتوگرافي آنزيم با فعاليت ويژهU/mg 246.47 و 12.59 برابر تخليص بدست آمد. نتايج الكتروفورز و زايموگرافي نشان دهنده باند ليپاز با وزن حدود 30 كيلو دالتون مي باشد.
Keywords :
Lipases , Purification , Chromatography , A.niger , Zymography
