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1399860 
Journal Article 
Three-phase partitioning of trypsin inhibitor from legume seeds 
Wati, RK; Theppakorn, T; Benjakul, S; Rawdkuen, S 
2009 
1 
Process Biochemistry
ISSN: 1359-5113
EISSN: 0032-9592 
Elsevier Science, The Boulevard Kidlington Oxford OX5 1GB UK 
44 
12 
1307-1314 
English 
WOS:000272071900001 
Three-phase partitioning (TPP) was used to isolate trypsin
inhibitors from navy bean (NB), red kidney bean (RK) and adzuki bean (AZ) from the Royal Project
Foundation in Thailand. The method was to mix the crude extract with solid ammonium sulfate (30%
saturation, w/v) and tert-butanol (t-butanol) in order to obtain the three phases. The trypsin
inhibitor was purified to 5-, 14- and 7-fold with 315%, 441% and 228% recovery for NB, RK and AZ,
respectively. The SDS-PAGE showed the major inhibitor band with the molecular weights (MWs) of
132,118 and 13 kDa for NB, RK and AZ, respectively. The fractions from NB and AZ showed higher pH
stability compared to that of the RK, and they had the optimum pH ranges of 7-9. The highest
relative inhibitory activity of the fractions of NB and RK were found at 50 degrees C, and all
fractions were quite stable at 90 degrees C for 60 min of incubation. Increasing the
concentration of salt (up to 3%, w/v) did not significantly decrease the inhibitory activity of
all fractions (p > 0.05). The trypsin inhibitors from the three legumes were unable to inhibit
the autolysis of Pacific whiting and arrowtooth flounder muscles. (C) 2009 Elsevier Ltd. All
rights reserved. 
Adzuki bean; Autolysis; Navy bean; Red kidney bean; Three-phase partitioning; Trypsin inhibitor 
IRIS
• tert-Butanol
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