To explore if trypsin could catalyze the degradation of non-protein molecule deltamethrin, we compared in vitro hydrolytic reactions of deltamethrin in the presence and absence of trypsin with ultraviolet-visible (UV/Vis) spectrophotometry and gas chromatography-mass spectrometry (GC/MS). In addition, acute oral toxicity of the degradation products was determined in Wistar rats. The results show that the absorption peak of deltamethrin is around 264 nm, while the absorption peaks of deltamethrin degradation products are around 250 nm and 296 nm. In our GC setting, the retention time of undegraded deltamethrin was 37.968 min, while those of deltamethrin degradation products were 15.289 min and 18.730 min. The LD50 of deltamethrin in Wistar rats is 55 mg/kg, while that of deltamethrin degradation products is 3358 mg/kg in female rats and 1045 mg/kg in male rates (61-fold and 19-fold reductions in toxicity), suggesting that trypsin could directly degrade deltamethrin, which significantly reduces the toxicity of deltamethrin. These results expand people's understanding of the functions of proteases and point to potential applications of trypsin as an attractive agent to control residual pesticides in the environment and on agricultural products.
References
[1]
Krem MM, Di Cera E (2001) Molecular markers of serine protease evolution. EMBO J 20: 3036–3045. doi: 10.1093/emboj/20.12.3036
[2]
Rawlings ND (1998b) Introduction: clan SC containing peptidases with the α/β hydrolase fold. In Barrett,A.J., Rawlings,N.D. and Woessner,J.F. (eds), Handbook of Proteolytic Enzymes. Academic Press, San Diego, CA, pp. 369–372.
[3]
Freeman HJ, Kim YS (1978) Digestion and absorption of protein. Annu Rev Med 29: 99–116. doi: 10.1146/annurev.me.29.020178.000531
[4]
Erickson RH, Kim YS (1990) Digestion and absorption of dietary protein. Annu Rev Med 41: 133–139. doi: 10.1146/annurev.me.41.020190.001025
[5]
Muhlia-Almazan A, Sanchez-Paz A, Garcia-Carreno FL (2008) Invertebrate trypsins: a review. J Comp Physiol B 178: 655–672. doi: 10.1007/s00360-008-0263-y
[6]
Wu HW, Tian HS, Wu GL, Langdon G, Kurtis J, et al. (2004) Culex pipiens pallens:identification of genes differentially expressed in deltamethrin-resistant and -susceptible strains. Pestic Biochem Phys 79(3): 75–83. doi: 10.1016/j.pestbp.2004.04.004
[7]
Gong MQ, Shen B, Gu Y, Tian HS, Ma L, et al. (2005) Serine proteinase over-expression in relation to deltamethrin resistance in Culex pipiens pallens. Arch Biochem Biophys 438(1): 53–62. doi: 10.1016/j.abb.2005.03.021
[8]
Leng G, Gries W (2005) Simultaneous determination of pyrethroid and pyrethrin metabolites in human urine by gas chromatography-high resolution mass spectrometry. J Chromatogr B 814: 285–94. doi: 10.1016/j.jchromb.2004.10.044
[9]
McLaren AD, Estermann EF (1957) Influence of pH on the activity of chymotrypsin at a solid-liquid interface. Arch Biochem Biophys 68(1): 157–60. doi: 10.1016/0003-9861(57)90336-3
[10]
Fredholt K, Savolainen J, Friis GJ (1999) alpha-Chymotrypsin-catalyzed degradation of desmopressin (dDAVP):influence of pH,concentration and various cyclodextrins. Int J Pharm 178(2): 223–9. doi: 10.1016/s0378-5173(98)00377-9
[11]
Ramesh A, Ravi PE (2004) Electron ionization gas chromatography-mass spectrometric determination of residues of thirteen pyrethroid pesticides in whole blood. J Chromatogr B 802(2): 371–6. doi: 10.1016/j.jchromb.2003.12.016
[12]
Modi SJ, LaCourse WR (2006) Monitoring carbohydrate enzymatic reactions by quantitative in vitro microdialysis. J chromatogr A 1118(1): 125–33. doi: 10.1016/j.chroma.2006.02.045
[13]
Leng G, Gries W (2005) Simultaneous determination of pyrethroid and pyrethrin metabolites in human urine by gas chromatography-high resolution mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci 814: 285–294. doi: 10.1016/j.jchromb.2004.10.044
[14]
Ross MK, Edwards CC, Potter PM (2006) Hydrolytic metabolism of pyrethroids by human and other mammalian carboxylesterases. Biochem pharmacol 71(5): 657–69. doi: 10.1016/j.bcp.2005.11.020
[15]
Kundu N, Roy S (1970) Esterase activity of chymotrypsin on O-DNP-L-tyrosine ethyl ester substrates. Nat 226: 1171. doi: 10.1038/2261171a0
[16]
Kushida K, Kato T, Hojo H (2001) High-performance liquid chromatographic- fluorimetric assay of chymotrypsin-like esterase activity. J Chromatogr B 762(2): 137–45. doi: 10.1016/s0378-4347(01)00353-x
[17]
WHO (1992) Vector resistance to pesticides.Fifteenth Report of the WHO Expert Committee on Vector Biology and Control. World Health Organ Tech Rep ser 818: 1–62. doi: 10.1017/s0266467400008531
[18]
Yang QG, Sun L, Zhang DH, Qian J, Sun Y, et al. (2008) Partial characterization of deltamethrin metabolism catalyzed by chymotrypsin. Toxicol In Vitro 22(6): 1528–33. doi: 10.1016/j.tiv.2008.05.007
[19]
Chen S, Lai K, Li Y, Hu M, Zhang Y, et al. (2011) Biodegradation of deltamethrin and its hydrolysis product 3-phenoxybenzaldehyde by a newly isolated Streptomyces aureus strain HP-S-01. Appl Microbiol Biotechnol 90(4): 1471–83. doi: 10.1007/s00253-011-3136-3
[20]
Zheng YZ, Lan WS, Qiao CL, Mulchandani A, Chen W (2007) Decontamination of vegetables sprayed with organophosphate pesticides by organophosphorus hydrolase and carboxylesterase (B1). Appl Biochem Biotechnol 136: 233–241. doi: 10.1007/s12010-007-9022-x
