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

Yıl 2009, Sayı: 40, 65 - 75, 27.04.2011

Öz

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.

Kaynakça

  • 1. Esen, A., (1993). β-Glucosidases: Overview. In: Esen, A. (Ed.), β-Glucosidases-Biochemistry and Molecular Biology, ACS Symposium Series 533, American Chemical Society, Washington, DC, pp.1-14.
  • 2. Poulton, J.E. (1990). Cyanogenesis in plants, Plant Physiol., 94: 401-405.
  • 3. Kleczkowski, K., Schell, J. (1995). Phytohormone conjugates: nature and function, Crit. Rev. Plant Sci., 14: 283-298.
  • 4. Akiyama, T., Kaku, H., Shibuya, N. (1998). A cell wall-bound β-glucosidase from germinated rice: Purification and properties, Phytochemistry, 48: 49-54.
  • 5. Sue, M., Ishihara, A., Iwamura, H. (2000). Purification and characterization of a β-glucosidase from rye (Secale cereale L.) seedlings, Plant Sci., 155: 67-74.
  • 6. Sue, M., Ishihara, A., Iwamura, H. (2000). Purification and characterization of a hydroxamic acid glucoside β-glucosidase from wheat (Triticum aestivum L.) seedlings, Planta, 210: 432-438.
  • 7. Cameron, R.G., Manthey, J.A., Baker, R.A., Grohmann, K. (2001). Purification and characterization of a β-glucosidase from Citrus sinensis var. Valencia fruit tissue, J. Agric. Food Chem., 49: 4457-4462.
  • 8. Lecas, M., Gunata, Z.Y., Sapis, J-C., Bayonove, C.L. (1991). Purification and partial characterization of β-glucosidase from grape, Phytochemistry, 30: 451- 454.
  • 9. Gerardi, C., Blando, F., Santino, A., Zacheo, G. (2001). Purification and characterization of β-glucosidase abundantly expressed in ripe sweet cherry (Prunus avium L.) fruit, Plant Sci., 160: 795-805.
  • 10. Rashid, M.H., Siddiqui, K.S. (1997). Purification and characterization of a β-glucosidase from Aspergillus niger, Folia Microbiol., 42: 544-550.
  • 11. Venturi, L.L., Polizeli, M.L., Terenzi, H.F., Furriel, R.P.M., Jorge, J.A. (2002). Extracellular β-D-glucosidase from Chaetomium thermophilum var. coprophilum: production, purification and some biochemical properties, J. Basic Microbiol., 42: 55-66.
  • 12. Capasso, F., Borrelli, F., Capasso, R., Di Carlo, G., Izzo, A.A., Pinto, L., Mascolo, N., Castaldo, S., Longo, R. (1998). Aloe and its therapeutic use, Phytother. Res., 12: S124-S127.
  • 13. Heggers, J.P., Kucukcelebi, A., Stabenau, C.J., Ko, F., Broemeling, L.D., Robson, M.C., Winters, W.D. (1995). Wound healing effects of Aloe gel and other topical antibacterial agents on rat skin, Phytother. Res., 9: 455-457.
  • 14. Koo, M.W.L. (1994). Aloe Vera: Antiulcer and antidiabetic effects, Phytother. Res., 8: 461-464.
  • 15. Saito, H. (1993). Purification of active substances of Aloe arborescens Miller and their biological and pharmacological activity, Phytother. Res., 7: S14-S19.
  • 16. Akev, N., Turkay, G., Can, A., Gurel, A., Yildiz, F., Yardibi, H., Ekiz, E.E., Uzun, H. (2007). Effect of Aloe vera leaf pulp extract on Ehrlich ascites tumours in mice, Eur. J. Cancer Prev., 16: 151-157.
  • 17. Beppu, H., Nagamuro, Y., Fujita, K. (1993). Hypoglycaemic and antidiabetic effects in mice of Aloe arborescens Miller var. natalensis Berger, Phytother. Res., 7: S37-S42.
  • 18. Okyar, A., Can, A., Akev, N., Baktir, G., Sütlüpinar, N. (2001). Effect of Aloe vera leaves on blood glucose level in type I and type II diabetic rat models, Phytother. Res., 15: 157-161.
  • 19. Yeoh, H.H., Tan, T.K., Koh, S.K. (1986). Kinetic properties of β-glucosidase from Aspergillus ornatus, Appl. Microbiol. Biotechnol., 25: 25-28.
  • 20. Odoux, E., Chauwin, A., Brillouet, J-M. (2003). Purification and characterization of vanilla bean (Vanilla planifolia Andrews) β-D-glucosidase, J. Agric. Food Chem., 51: 3168-3173.
  • 21. Hartmann-Schreier, J., Schreier P., (1986). Purification and partial characterization of β-glucosidase from papaya fruit, Phytochemistry, 25: 2271- 2274. 75
  • 22. Hsieh, M-C., Graham, T.L. (2001). Partial purification and characterization of a soybean β-glucosidase with high specific activity towards isoflavone conjugates, Phytochemistry, 58: 995-1005.
  • 23. Leah, R., Kigel, J., Svendsen, I., Mundy, J. (1995). Biochemical and molecular characterization of a barley seed β-glucosidase, J. Biol. Chem., 270: 15789- 15797.
  • 24. Sivakumar, G., Bati, C.B., Ucella, N. (2007). Demethyloleuropein and -glucosidase activity in olive fruits, Biotechnol. J., 2: 381-385.
  • 25. Tiselius, A., Hjertén, S., Levin, Ö. (1956). Protein chromatography on calcium phosphate columns, Arch. Biochem. Biophys., 65: 132-135.
  • 26. Lowry, O.H., Rosebrough, N.J., Farr, A.L., Randall, R.J. (1951). Protein measurement with the Folin phenol regent, J. Biol. Chem., 193: 265-275.
  • 27. Warburg, O., Christian, W. (1941). Isolierung und Kristallisation des Gärungsferments Enolase, Biochem. Z., 310: 384-421.
  • 28. Ornstein, L. (1964). Disc electrophoresis-I. Background and theory, Ann. N.Y. Acad. Sci., 121: 321-349.
  • 29. Davis, B.J. (1964). Disc electrophoresis-II. Method and application to human serum proteins, Ann. N.Y. Acad. Sci., 121: 404-427.
  • 30. Coughlan, M.P. (1988). Staining techniques for the detection of the individual components of cellulolytic enzyme systems, Methods Enzymol., 160: 135-144.

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

Yıl 2009, Sayı: 40, 65 - 75, 27.04.2011

Öz

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.

Kaynakça

  • 1. Esen, A., (1993). β-Glucosidases: Overview. In: Esen, A. (Ed.), β-Glucosidases-Biochemistry and Molecular Biology, ACS Symposium Series 533, American Chemical Society, Washington, DC, pp.1-14.
  • 2. Poulton, J.E. (1990). Cyanogenesis in plants, Plant Physiol., 94: 401-405.
  • 3. Kleczkowski, K., Schell, J. (1995). Phytohormone conjugates: nature and function, Crit. Rev. Plant Sci., 14: 283-298.
  • 4. Akiyama, T., Kaku, H., Shibuya, N. (1998). A cell wall-bound β-glucosidase from germinated rice: Purification and properties, Phytochemistry, 48: 49-54.
  • 5. Sue, M., Ishihara, A., Iwamura, H. (2000). Purification and characterization of a β-glucosidase from rye (Secale cereale L.) seedlings, Plant Sci., 155: 67-74.
  • 6. Sue, M., Ishihara, A., Iwamura, H. (2000). Purification and characterization of a hydroxamic acid glucoside β-glucosidase from wheat (Triticum aestivum L.) seedlings, Planta, 210: 432-438.
  • 7. Cameron, R.G., Manthey, J.A., Baker, R.A., Grohmann, K. (2001). Purification and characterization of a β-glucosidase from Citrus sinensis var. Valencia fruit tissue, J. Agric. Food Chem., 49: 4457-4462.
  • 8. Lecas, M., Gunata, Z.Y., Sapis, J-C., Bayonove, C.L. (1991). Purification and partial characterization of β-glucosidase from grape, Phytochemistry, 30: 451- 454.
  • 9. Gerardi, C., Blando, F., Santino, A., Zacheo, G. (2001). Purification and characterization of β-glucosidase abundantly expressed in ripe sweet cherry (Prunus avium L.) fruit, Plant Sci., 160: 795-805.
  • 10. Rashid, M.H., Siddiqui, K.S. (1997). Purification and characterization of a β-glucosidase from Aspergillus niger, Folia Microbiol., 42: 544-550.
  • 11. Venturi, L.L., Polizeli, M.L., Terenzi, H.F., Furriel, R.P.M., Jorge, J.A. (2002). Extracellular β-D-glucosidase from Chaetomium thermophilum var. coprophilum: production, purification and some biochemical properties, J. Basic Microbiol., 42: 55-66.
  • 12. Capasso, F., Borrelli, F., Capasso, R., Di Carlo, G., Izzo, A.A., Pinto, L., Mascolo, N., Castaldo, S., Longo, R. (1998). Aloe and its therapeutic use, Phytother. Res., 12: S124-S127.
  • 13. Heggers, J.P., Kucukcelebi, A., Stabenau, C.J., Ko, F., Broemeling, L.D., Robson, M.C., Winters, W.D. (1995). Wound healing effects of Aloe gel and other topical antibacterial agents on rat skin, Phytother. Res., 9: 455-457.
  • 14. Koo, M.W.L. (1994). Aloe Vera: Antiulcer and antidiabetic effects, Phytother. Res., 8: 461-464.
  • 15. Saito, H. (1993). Purification of active substances of Aloe arborescens Miller and their biological and pharmacological activity, Phytother. Res., 7: S14-S19.
  • 16. Akev, N., Turkay, G., Can, A., Gurel, A., Yildiz, F., Yardibi, H., Ekiz, E.E., Uzun, H. (2007). Effect of Aloe vera leaf pulp extract on Ehrlich ascites tumours in mice, Eur. J. Cancer Prev., 16: 151-157.
  • 17. Beppu, H., Nagamuro, Y., Fujita, K. (1993). Hypoglycaemic and antidiabetic effects in mice of Aloe arborescens Miller var. natalensis Berger, Phytother. Res., 7: S37-S42.
  • 18. Okyar, A., Can, A., Akev, N., Baktir, G., Sütlüpinar, N. (2001). Effect of Aloe vera leaves on blood glucose level in type I and type II diabetic rat models, Phytother. Res., 15: 157-161.
  • 19. Yeoh, H.H., Tan, T.K., Koh, S.K. (1986). Kinetic properties of β-glucosidase from Aspergillus ornatus, Appl. Microbiol. Biotechnol., 25: 25-28.
  • 20. Odoux, E., Chauwin, A., Brillouet, J-M. (2003). Purification and characterization of vanilla bean (Vanilla planifolia Andrews) β-D-glucosidase, J. Agric. Food Chem., 51: 3168-3173.
  • 21. Hartmann-Schreier, J., Schreier P., (1986). Purification and partial characterization of β-glucosidase from papaya fruit, Phytochemistry, 25: 2271- 2274. 75
  • 22. Hsieh, M-C., Graham, T.L. (2001). Partial purification and characterization of a soybean β-glucosidase with high specific activity towards isoflavone conjugates, Phytochemistry, 58: 995-1005.
  • 23. Leah, R., Kigel, J., Svendsen, I., Mundy, J. (1995). Biochemical and molecular characterization of a barley seed β-glucosidase, J. Biol. Chem., 270: 15789- 15797.
  • 24. Sivakumar, G., Bati, C.B., Ucella, N. (2007). Demethyloleuropein and -glucosidase activity in olive fruits, Biotechnol. J., 2: 381-385.
  • 25. Tiselius, A., Hjertén, S., Levin, Ö. (1956). Protein chromatography on calcium phosphate columns, Arch. Biochem. Biophys., 65: 132-135.
  • 26. Lowry, O.H., Rosebrough, N.J., Farr, A.L., Randall, R.J. (1951). Protein measurement with the Folin phenol regent, J. Biol. Chem., 193: 265-275.
  • 27. Warburg, O., Christian, W. (1941). Isolierung und Kristallisation des Gärungsferments Enolase, Biochem. Z., 310: 384-421.
  • 28. Ornstein, L. (1964). Disc electrophoresis-I. Background and theory, Ann. N.Y. Acad. Sci., 121: 321-349.
  • 29. Davis, B.J. (1964). Disc electrophoresis-II. Method and application to human serum proteins, Ann. N.Y. Acad. Sci., 121: 404-427.
  • 30. Coughlan, M.P. (1988). Staining techniques for the detection of the individual components of cellulolytic enzyme systems, Methods Enzymol., 160: 135-144.
Toplam 30 adet kaynakça vardır.

Ayrıntılar

Birincil Dil İngilizce
Konular Eczacılık ve İlaç Bilimleri
Bölüm Makaleler
Yazarlar

Tuğba Yılmaz Bu kişi benim

Ayşe Can Bu kişi benim

Yayımlanma Tarihi 27 Nisan 2011
Yayımlandığı Sayı Yıl 2009 Sayı: 40

Kaynak Göster

APA Yılmaz, T., & Can, A. (2011). PARTIAL PURIFICATION AND CHARACTERIZATION OF -GLUCOSIDASE FROM ALOE VERA L. LEAVES. Journal of Faculty of Pharmacy of Istanbul University(40), 65-75.
AMA Yılmaz T, Can A. PARTIAL PURIFICATION AND CHARACTERIZATION OF -GLUCOSIDASE FROM ALOE VERA L. LEAVES. Journal of Faculty of Pharmacy of Istanbul University. Mayıs 2011;(40):65-75.
Chicago Yılmaz, Tuğba, ve Ayşe Can. “PARTIAL PURIFICATION AND CHARACTERIZATION OF -GLUCOSIDASE FROM ALOE VERA L. LEAVES”. Journal of Faculty of Pharmacy of Istanbul University, sy. 40 (Mayıs 2011): 65-75.
EndNote Yılmaz T, Can A (01 Mayıs 2011) PARTIAL PURIFICATION AND CHARACTERIZATION OF -GLUCOSIDASE FROM ALOE VERA L. LEAVES. Journal of Faculty of Pharmacy of Istanbul University 40 65–75.
IEEE T. Yılmaz ve A. Can, “PARTIAL PURIFICATION AND CHARACTERIZATION OF -GLUCOSIDASE FROM ALOE VERA L. LEAVES”, Journal of Faculty of Pharmacy of Istanbul University, sy. 40, ss. 65–75, Mayıs 2011.
ISNAD Yılmaz, Tuğba - Can, Ayşe. “PARTIAL PURIFICATION AND CHARACTERIZATION OF -GLUCOSIDASE FROM ALOE VERA L. LEAVES”. Journal of Faculty of Pharmacy of Istanbul University 40 (Mayıs 2011), 65-75.
JAMA Yılmaz T, Can A. PARTIAL PURIFICATION AND CHARACTERIZATION OF -GLUCOSIDASE FROM ALOE VERA L. LEAVES. Journal of Faculty of Pharmacy of Istanbul University. 2011;:65–75.
MLA Yılmaz, Tuğba ve Ayşe Can. “PARTIAL PURIFICATION AND CHARACTERIZATION OF -GLUCOSIDASE FROM ALOE VERA L. LEAVES”. Journal of Faculty of Pharmacy of Istanbul University, sy. 40, 2011, ss. 65-75.
Vancouver Yılmaz T, Can A. PARTIAL PURIFICATION AND CHARACTERIZATION OF -GLUCOSIDASE FROM ALOE VERA L. LEAVES. Journal of Faculty of Pharmacy of Istanbul University. 2011(40):65-7.