PARTIAL PURIFICATION AND CHARACTERIZATION OF -GLUCOSIDASE FROM ALOE VERA L. LEAVES

Abstract

The partial purification of β-glucosidase from the pulp of Aloe vera L. Burm. fil. (sarisabir) leaves and some of its kinetic properties is presented. The fresh leaves of A. vera were used; the gel portion was separated and the remaining leaf pulps were cut into small pieces. The crude extract was prepared by homogenization of the leaf pulps in phosphate buffered saline (PBS), pH 7.4 and subsequent centrifugation. β-Glucosidase active fraction was precipitated by 30%-65% ammonium sulphate from the crude extract. Hydroxylapatite column chromatography resulted in a single peak showing β-glucosidase activity eluted with 200 mM phosphate buffer. The partially purified enzyme showed two protein and a single activity band in polyacrylamide gel electrophoresis. It was found that the enzyme exhibited maximum activity at 50oC and at pH 4.4. Km and Vmax values for 4-nitrophenyl-β-D-glucopyranoside were 6.8x10-4 M and 4,58x10-3 U, respectively. When β-glucosidase activity was investigated throughout the year, it was found that the activity increased in winter and decreased in summer.

Keywords

• glycosidases
• -glucosidase
• Aloe vera
• purification
• seasonal variation

PARTIAL PURIFICATION AND CHARACTERIZATION OF -GLUCOSIDASE FROM ALOE VERA L. LEAVES

Abstract

Aloe vera L. Burm. fil. (sarısabır) yapraklarının pulpa kısmından β-glukozidaz kısmen saflaştırıldı ve bazı kinetik özellikleri incelendi. A.vera‟nın taze yaprakları kullanıldı; jel kısmı ayrıldı ve kalan yaprak pulpaları küçük parçalara kesildi. Yaprak pulpalarının fosfatla tamponlanmış %0.9 NaCl (PBS), pH 7.4 ile homojenizasyonunun ardından santrifüje edilmesiyle ham ekstre hazırlandı. β-Glukozidaz aktivitesi gösteren fraksiyon ham ekstreden %30-%65 amonyum sülfat ile çöktürüldü. Hidroksilapatit kolon kromatografisi sonucunda 200 mM fosfat tamponu ile β-glukozidaz aktivitesi gösteren tek bir pik elde edildi. Kısmen saflaştırılan enzim poliakrilamid jel elektroforezinde iki protein ve bir aktivite bandı gösterdi. Enzimin en yüksek aktiviteyi 50C‟de ve pH 4.4‟te gösterdiği ve 4-nitrofenil-β-D-glukopiranozide karşı Km değerinin 6.8x10-4 M, Vmax değerinin ise 4.58x10-3 U olduğu saptandı. β-Glukozidaz aktivitesi yıl boyunca incelendiğinde aktivitenin kış aylarında arttığı, yaz aylarında ise azaldığı bulundu.

Keywords

• glycosidases
• -glucosidase
• Aloe vera
• purification
• seasonal variation

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