Isolation and characterization of a trypsin fraction from the pyloric ceca of chinook salmon (Oncorhynchus tshawytscha) I. Kurtovic a , S.N. Marshall a, ⁎ , B.K. Simpson b a Seafoods and Marine Extracts Group, Crop and Food Research Limited, P.O. Box 5114, Port Nelson, Nelson, New Zealand b Food Science Department, McGill University (Macdonald Campus) 21,111 Lakeshore Road, Ste. Anne de Bellevue (QC) Canada H9X 3V9 Received 8 August 2005; received in revised form 20 December 2005; accepted 20 December 2005 Available online 2 February 2006 Abstract A trypsin fraction was isolated from the pyloric ceca of New Zealand farmed chinook salmon (Oncorhynchus tshawytscha) by ammonium sulfate fractionation, acetone precipitation and affinity chromatography. The chinook salmon enzyme hydrolyzed the trypsin-specific synthetic substrate benzoyl-DL-arginine-p-nitroanilide (DL-BAPNA), and was inhibited by the general serine protease inhibitor phenyl methyl sulfonyl fluoride (PMSF), and also by the specific trypsin inhibitors — soybean trypsin inhibitor (SBTI) and benzamidine. The enzyme was active over a broad pH range (from 7.5 to at least pH 10.0) at 25 °C and was stable from pH 4.0 to pH 10.0 when incubated at 20 °C, with a maximum at pH 8.0. The optimum temperature for the hydrolysis of DL-BAPNA by the chinook salmon enzyme was 60 °C, however, the enzyme was unstable at temperatures above 40 °C. The molecular mass of the chinook salmon trypsin was estimated as 28 kDa by SDS–PAGE. © 2006 Elsevier Inc. All rights reserved. Keywords: Affinity chromatography; Chinook salmon; Farmed fish; Fish viscera; Oncorhynchus tshawytscha; Proteolytic enzymes; Serine protease; Trypsin 1. Introduction Chinook salmon (Oncorhynchus tshawytscha), also known as Pacific, king or quinnat salmon, is native to the northwest coast of North America and northeast Asia. The species is anadromous, living in saltwater as adults, then swimming up rivers to spawn in fresh water. New Zealand has no native trout or salmon and all of the salmonids in the country have been deliberately imported from elsewhere. First attempts to introduce chinook salmon started in the 1870s but were not really successful until releases occurred from Sacramento River stock between 1901 and 1907 (McDowall, 1995; Grzlewski, 2003). Colonization of the present-day range of the fish occurred within 10 years of these releases (Quinn et al., 2001). Chinook salmon farming is now an important industry in New Zealand. The farmed populations have come from fish originally captured in the rivers of the east coast of the South Island. The main saltwater farms are situated in the Marlbor- ough Sounds, Stewart Island, and Akaroa Harbour, whilst much of the freshwater farming is carried out in the raceways of the South Island hydroelectric schemes. The animals are reared from eggs in freshwater hatcheries, then grown to harvest masses of about 3.0–3.5 kg within 12 to 24 months in saltwater pens. In New Zealand, chinook salmon live in temperate waters ranging from a winter low of about 7 °C to a summer high of around 18 °C, with some variability depending on location (Gillard, 2005). On a world scale the salmon fishery in New Zealand is not large at 7500–8000 tonnes/year (as gilled and gutted fish) (Gillard, 2005). Viscera account for approximately 1000 tonnes/ year and less than 15% of this mass is contributed by pyloric ceca. To a by-products processor, the possible disadvantage of this relatively low volume is compensated for by the generation of very fresh by-products of consistent composition that are produced and processed in a highly regulated and controlled manner. This viscera material and other residuals (e.g. gills, skins, fins, heads and bones) from commercial chinook salmon processing are loaded with useful bioingredients such as enzymes, collagen and omega-3 polyunsaturated fatty acids (n-3 PUFAs). Thus far, the residuals have been used to produce Comparative Biochemistry and Physiology, Part B 143 (2006) 432 – 440 www.elsevier.com/locate/cbpb ⁎ Corresponding author. Tel.: +64 3 5391811; fax: +64 3 5467049. E-mail address: MarshallS@crop.cri.nz (S.N. Marshall). 1096-4959/$ - see front matter © 2006 Elsevier Inc. All rights reserved. doi:10.1016/j.cbpb.2005.12.020