Effect of trypsin inhibitor from Crotalaria pallida seeds on Callosobruchus maculatus (cowpea weevil) and Ceratitis capitata (fruit fly).
A proteinaceous trypsin inhibitor was purified from Crotalaria pallida seeds by ammonium sulfate precipitation, affinity chromatography on immobilized trypsin-Sepharose and TCA precipitation. The trypsin inhibitor, named CpaTI, had M(r) of 32.5 kDa as determined by SDS-PAGE and was composed of two subunits with 27.7 and 5.6 kDa linked by disulfide bridges. CpaTI was stable at 50 degrees C and lost 40% of activity at 100 degrees C. CpaTI was also stable from pH 2 to 12 at 37 degrees C. CpaTI weakly inhibited chymotrypsin and elastase and its inhibition of papain, a cysteine proteinase, were indicative of its bi-functionality. CpaTI inhibited, in different degrees, digestive enzymes from Spodoptera frugiperda, Alabama argillacea, Plodiainterpunctella, Anthonomus grandis and Zabrotes subfasciatus guts. In vitro and in vivo susceptibility of Callosobruchus maculatus and Ceratitis capitata to CpaTI was evaluated. C. maculatus and C. capitata enzymes were strongly susceptible, 74.4+/-15.8% and 100.0+/-7.3%, respectively, to CpaTI. When CpaTI was added to artificial diets and offered to both insect larvae, the results showed that C. maculatus was more susceptible to CpaTI with an LD(50) of 3.0 and ED(50) of 2.17%. C. capitata larvae were more resistant to CpaTI, in disagreement with the in vitro effects. The larvae were more affected at lower concentrations, causing 27% mortality and 44.4% mass decrease. The action was constant at 2-4% (w/w) with 15% mortality and 38% mass decrease.
Gomes
Carlos E M
CE
Laboratório de Química e Função de Proteínas Bioativas, Departamento de Bioquímica, Centro de Biociências, Universidade Federal do Rio Grande do Norte, Natal, RN, Brazil.
Barbosa
Aulus E A D
AE
Macedo
Leonardo L P
LL
Pitanga
Joelma C M
JC
Moura
Fabiano T
FT
Oliveira
Adeliana S
AS
Moura
Raniere M
RM
Queiroz
Alexandre F S
AF
Macedo
Francisco P
FP
Andrade
Lúcia B S
LB
Vidal
Márcia S
MS
Sales
Mauricio P
MP
D000818:Animals; D033622:Ceratitis capitata / Q000201:enzymology* / Q000254:growth & development; D002918:Chymotrypsin / Q000037:antagonists & inhibitors; D029864:Crotalaria / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D007306:Insecticides / Q000302:isolation & purification; D007814:Larva / Q000201:enzymology; D007928:Lethal Dose 50; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors; D012639:Seeds / Q000378:metabolism; D013696:Temperature; D014361:Trypsin Inhibitors / Q000302:isolation & purification; D048889:Weevils / Q000201:enzymology* / Q000254:growth & development
D007306:Insecticides; D014361:Trypsin Inhibitors; D002918:Chymotrypsin; D010196:Pancreatic Elastase