Inhibitors of human and bovine trypsin and chymotrypsin in fenugreek (Trigonella foenum-graecum L.) seeds

Demonstration and purification
  • Jürgen K. P. Weder
  • Kristine Haußner
Original Paper

Summary

Fenugreek contained proteinase inhibitors inhibiting 5–9 mg human trypsin, 5–7 mg bovine trypsin, 2–6 mg human chymotrypsin, and 1–3 mg bovine chymotrypsin per g seed material. About 30 inhibitors were electrophoretically detected, and 23 of them, inhibiting all the four enzymes, were characterized by means of their isoelectric points: a group of acid inhibitors (TFI-A1 to A10, pI 4.48–5.12), a group of neutral inhibitors (TFI-N1 to-N6, pI 5.91–6.71), and a group of basic inhibitors (TFI-B1 to-B7, pI 7.76–9.77). To eliminate the galactomannans which complicate further purification, coarsely ground seeds were separated by density into two fractions, seed coats+endosperm and cotyledons+embryos (C+E). Isolation of the fenugreek inhibitors by extraction of fraction C+E, followed by ammonium sulfate fractionation and affinity chromatography on anhydrotrypsin-Sepharose, resulted in an about 700-fold enrichment.

Inhibitoren für Human- and Rinder-Trypsin und -Chymotrypsin in Bockshornkleesamen (Trigonella foenum-graecum L.) Nachweis und Reinigung

Zusammenfassung

Bockshornkleesamen enthalten Proteinaseinhibitoren, die pro g Samenmaterial 5–9 mg Humantrypsin, 5–7 mg Rindertrypsin, 2–6 mg Humanchymotrypsin und 1–3 mg Rinderchymotrypsin hemmen. Von den ca. 30 elektrophoretisch nachweisbaren Inhibitoren wurden 23 durch ihre isoelektrischen Punkte charakterisiert, die alle vier genannten Enzyme hemmen. Sie bilden eine Gruppe saurer Inhibitoren (TFI-A1 bis-A10, pI 4,48–5,12), eine Gruppe neutraler Inhibitoren (TFI-N1 bis-N6, pI 5,91–6,71) und eine Gruppe basischer Inhibitoren (TFI-B1 bis-B7, pl 7,76–9,77). Zur Ausschaltung der die weitere Aufarbeitung stark störenden Galaktomannane wurden die grob gemahlenen Samen durch Dichteklassierung in die zwei Fraktionen Samenschale+ Endosperm und Keimling+Embryo (C+E) getrennt und die Inhibitoren aus der Fraktion C+E extrahiert. Durch Ammoniumsulfatfraktionierung und anschließende Affinitätschromatographie an Anhydrotrypsin-Se-pharose zur Abtrennung der Begleitproteine wurde insgesamt eine ca. 700fache Anreicherung erreicht.

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Copyright information

© Springer-Verlag 1991

Authors and Affiliations

  • Jürgen K. P. Weder
    • 1
  • Kristine Haußner
    • 1
  1. 1.Institut für LebensmittelchemieTechnische Universität MünchenGarchingGermany

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