Skip to main content
SpringerLink
Log in

Influence of time, storage temperature and freeze/thaw cycles on the activity of digestive enzymes from gilthead sea bream (Sparus aurata)

  • Published:
Fish Physiology and Biochemistry Aims and scope Submit manuscript

Abstract

In this study, we tested the effects of long-term storage (2 years) at −20 °C and short-term storage (several hours) in ice and freeze/thaw cycles on the activities of pancreatic, gastric and intestinal (brush border and cytosolic) digestive enzymes in a teleost fish species. The results revealed a significant lose in activity of pancreatic (trypsin, chymotrypsin, total alkaline proteases and α-amylase) and intestinal cytosolic (leucine–alanine peptidase) enzymes between 140 and 270 days of storage at −20 °C, whereas in contrast, the activity of all the assayed brush border enzymes remained constant during the first 2 years of storage at −20 °C. During short-term storage conditions, the most stable enzymes assayed were those of the enterocytes of the brush border, which did not show any change in activity after being held for 5 h in ice. Five freezing and thawing cycles did not affect the activity of the intestinal brush border enzymes and the cytosolic ones, whereas the activity of trypsin, α-amylase and bile-salt-activated lipase was significantly affected by the number of freezing and thawing cycles. No changes in pepsin activity were found in samples exposed to 1 and 2 freezing and thawing cycles.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

References

  • Ahsan N, Watabe S (2001) Kinetic and structural properties of two isoforms of trypsin isolated from the viscera of Japanese Anchovy, Engraulis japonicas. J Protein Chem 20(1):49–58

    Article  CAS  PubMed  Google Scholar 

  • Aruee ANA, Simpson BK, Villalonga R (2007) Lipase fraction from the viscera of grey mullet (Mugil cephalus) isolation, partial purification and some biochemical characteristics. Enzyme Microb Technol 40:394–402

    Article  Google Scholar 

  • Asgeirsson B, Bjarnason JB (1991) Structural and kinetic properties of chymotrypsin from Atlantic cod (Gadus morhua). Comparison with bovine chymotrypsin. Comp Biochem Physiol B: Biochem Mol Biol 99(2):327–335

    CAS  Google Scholar 

  • Bessey OA, Lowry OH, Brock MJ (1946) Rapid coloric method for determination of alkaline phosphatase in five cubic millimeters of serum. J Biol Chem 164:321–329

    CAS  PubMed  Google Scholar 

  • Bradford MM (1976) A rapid and sensitive method for the quantification of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  CAS  PubMed  Google Scholar 

  • Brunstein J (2012) Freeze-thaw cycles and nucleic acid stability: what’s safe for your samples? Med Lab Obs 47(9):44–47

    Google Scholar 

  • Chakrabarti I, Gani MA, Chaki KK, Sur R, Misra KK (1995) Digestive enzymes in 11 freshwater teleost fish species in relation to food habit and niche segregation. Comp Biochem Physiol A Physiol 112:167–177

    Article  Google Scholar 

  • Cohen T, Gertler A, Birk Y (1981) Pancreatic proteolytic enzymes from carp (Cyprinus carpio)—I. purification and physical properties of trypsin, chymotrypsin, elastase and carboxypeptidase B. Comp Biochem Physiol B: Biochem Mol Biol 69:639–646

    Google Scholar 

  • Crane RK, Boge G, Rigal A (1979) Isolation of brush border membranes in vesicular form from the intestinal spiral valve of the small dogfish Scyliorhinus canicula. Biochim Biophys Acta 554:264–267

    Article  CAS  PubMed  Google Scholar 

  • Dahkqvist A (1970) Assay of intestinal disaccharidase. Enzym Biol Clin 11:52–66

    Google Scholar 

  • Fernandez I, Moyano FJ, Dıaz M, Martınez T (2001) Characterization of a-amylase activity in five species of Mediterranean sparid fishes. J Exp Mar Biol Ecol 262:1–12

    Article  CAS  Google Scholar 

  • García-Careño FL, Haard NF (1993) Characterization of proteinase classes in langostilla (Pleuroncodes planipes) and crayfish (Pacifastacus astacus) extracts. J Food Biochem 17:97–113

    Article  Google Scholar 

  • Gelman A, Kuz’mina V, Drabkin V, Glatman L (2003) Temperature dependent characteristics of intestinal glycyl-l-leucine dipeptidase in boreal zone fish. Comp Biochem Physiol B: Biochem Mol Biol 136(2):323–329

    Article  CAS  Google Scholar 

  • German DP, Gawlicka AK, Horn MH (2014) Evolution of ontogenetic dietary shifts and associated gut features in prickleback fishes (Teleostei: Stichaeidae). Comp Biochem Physiol B: Biochem Mol Biol 168:12–18

    Article  CAS  Google Scholar 

  • Gisbert E, Giménez G, Fernández I, Kotzamanis Y, Estévez A (2009) Development of digestive enzymes in common dentex Dentex dentex during early ontogeny. Aquaculture 267(3–4):381–387

    Article  Google Scholar 

  • Gisbert E, Morais S, Moyano FJ (2013) Feeding and Digestion. In: Qin JG (ed) Larval fish aquaculture. Nova Publishers, New York, pp 73–124

    Google Scholar 

  • Gjellesvik DR, Lombardo D, Walther BT (1992) Pancreatic bile salt dependent lipase from cod (Gadus morhua): purification and properties. BBA 1124:123–134

    CAS  PubMed  Google Scholar 

  • Görgün S, Akpınar MA (2012) Purification and characterization of lipase from the liver of carp, Cyprinus carpio L. (1758), Living in Lake Tödürge (Sivas, Türkiye). Turk J Fish Aquat Sci 12:207–215

    Google Scholar 

  • Heu MS, Kim HR, Pyeun JH (1995) Comparison of trypsin and chymotrypsin from the viscera of anchovy, Engraulis japonica. Comp Biochem Physiol B Biochem Mol Biol 112(3):557–567

    Article  CAS  PubMed  Google Scholar 

  • Hiranpradit S, Lopez A (1976) Activity and stability of a- and b-amylase at temperatures from 4 °C to −23 °C. J Food Sci 41:138–144

    Article  CAS  Google Scholar 

  • Hjelmeland K, Raa J (1982) Characteristics of two trypsin type isozymes isolated from the arctic fish capelin (Mallotus villosus). Comp Biochem Physiol B: Biochem Mol Biol 71(4):557–562

    Article  CAS  Google Scholar 

  • Holm H, Hanssen LE, Krogdahl A, Florholmen J (1988) High and low inhibitor soybean meals affect human duodenal proteinase activity differently: in vivo comparison with bovine serum albumin. J Nutr 118:515–520

    CAS  PubMed  Google Scholar 

  • Huang K, Takahara S, Kinouchi T, Takeyama M, Ishida T, Ueyama H, Nishi K, Ohkubo I (1997) Alanyl aminopeptidase from human seminal plasma: purification, characterization, and immunohistochemical localization in the male genital tract. J Biochem 122(4):779–787

    Article  CAS  PubMed  Google Scholar 

  • Iijima N, Tanaka S, Ota Y (1998) Purification and characterization of bile-salt activated lipase from the hepatopancreas of red sea bream, Pagrus major. Fish Physiol Biochem 18:59–69

    Article  CAS  Google Scholar 

  • Izvekova GI, Solovyev MM, Kashinskaya EN, Izvekov EI (2013) Variations in the activity of digestive enzymes along the intestine of the burbot Lota lota expressed by different methods. Fish Physiol Biochem 39(5):1181–1193

    Article  CAS  PubMed  Google Scholar 

  • Klomklao S, Benjakul S, Visessanguan W, Kishimura H, Simpson BK, Saeki H (2006) Trypsins from yellowfin tuna (Thunnus albacores) spleen: purification and characterization. Comp Biochem Physiol B: Biochem Mol Biol 144:47–56

    Article  Google Scholar 

  • Krogdahl Å, Gajardo K, Kortner TM, Penn M, Gu M, Berge GM, Bakke AM (2015) Soya saponins induce enteritis in Atlantic salmon (Salmo salar L.). J Agric Food Chem 63(15):3887–3902

    Article  CAS  PubMed  Google Scholar 

  • Kumar A, Venkatesu P (2012) Overview of the stability of α-chymotrypsin in different solvent media. Chem Rev 112(7):4283–4307

    Article  CAS  PubMed  Google Scholar 

  • Kurtovic I, Marshall SN, Simpson BK (2006) Isolation and characterization of a trypsin fraction from the pyloric ceca of chinook salmon (Oncorhynchus tshawytscha). Comp Biochem Physiol B: Biochem Mol Biol 143:432–440

    Article  CAS  Google Scholar 

  • Kurtovic I, Marshall SN, Zhao X, Simpson BK (2009) Lipases from mammals and fishes. Rev Fish Sci 17(1):18–40

    Article  CAS  Google Scholar 

  • Kurtovic I, Marshall SN, Zhao X, Simpson BK (2010) Purification and properties of digestive lipases from Chinook salmon (Oncorhynchus tshawytscha) and New Zealand hoki (Macruronus novaezelandiae). Fish Physiol Biochem 36(4):1041–1060

    Article  CAS  PubMed  Google Scholar 

  • Kuz’mina VV (1996) Influence of age digestive enzyme activity in some freshwater teleosts. Aquaculture 148:25–37

    Article  Google Scholar 

  • Lazo JP, Darias MJ, Gisbert E (2011) Ontogeny of the digestive tract. In: Holt GJ (ed) Larval fish nutrition. Wiley-Blackwell, West Sussex, p 3–46

    Chapter  Google Scholar 

  • Lee DH, Kim JW, Jeon SY, Park BK, Han BG (2010) Proteomic analysis of the effect of storage temperature on human serum. Ann Clin Lab Sci 40(1):61–70

    CAS  PubMed  Google Scholar 

  • Maroux S, Louvard D, Baratti J (1973) The aminopeptidase from hog-intestinal brush border. Biochim Biophys Acta 321:282–295

    Article  CAS  PubMed  Google Scholar 

  • Métais P, Bieth J (1968) Détermination de l'α-amylase. Ann Biol Clin 26:133–142

    Google Scholar 

  • Muller OH, Yamanouchi I (1958) Stable solutions of trypsin. Arch Biochem Biophys 76(2):328–341

    Article  CAS  PubMed  Google Scholar 

  • Nicholson JA, Kim YS (1975) A one-step l-amino acid oxidase assay for intestinal peptide hydrolase activity. Anal Biochem 63:110–117

    Article  CAS  Google Scholar 

  • Nolasco N, Moyano-Lopez F, Vega-Villasante F (2011) Partial characterization of pyloric-duodenal lipase of gilthead seabream (Sparus aurata). Fish Physiol Biochem 37(1):43–52

    Article  CAS  PubMed  Google Scholar 

  • Perera E, Moyano FJ, Díaz M, Perdomo-Morales R, Montero-Alejo V, Rodriguez-Viera L, Alonso E, Carrillo O, Galich GS (2008) Changes in digestive enzymes through developmental and molt stages in the spiny lobster, Panulirus argus. Comp Biochem Physiol B: Biochem Mol Biol 151(3):250–260

    Article  Google Scholar 

  • Raae AJ, Walter BT (1989) Purification and characterization of chymotrypsin, trypsin and elastase like proteinases from cod (Gadus morhua L.). Comp Biochem Physiol B: Biochem Mol Biol 93(2):317–324

    CAS  Google Scholar 

  • Rajagopalan TG, Moore S, Stein WH (1966) Pepsin from pepsinogen. Preparation and properties. J Biol Chem 241:4940–4950

    CAS  PubMed  Google Scholar 

  • Reshkin SJ, Cassano G, Womersley C, Ahearn GA (1988) Preservation of glucose transport and enzyme activity in fish intestinal brush-border and basolateral membrane vesicles. J Exp Biol 140:123–135

    CAS  Google Scholar 

  • Rothman S, Liebow C, Isenman L (2002) Conservation of digestive enzymes. Physiol Rev 82(1):1–18

    Article  CAS  PubMed  Google Scholar 

  • Rotllant G, Moyano FJ, Andrés M, Díaz M, Estévez A, Gisbert E (2008) Evaluation of fluorogenic substrates in the assessment of digestive enzymes in a decapod crustacean Maja brachydactyla larvae. Aquaculture 282:90–96

    Article  CAS  Google Scholar 

  • Ryan CA, Clary JJ, Tomimatsu Y (1965) Chicken chymotrypsin and turkey trypsin part II: physical and enzymic properties. Arch Biochem Biophys 110:175–183

    Article  CAS  PubMed  Google Scholar 

  • Solovyev MM, Kashinskaya EN, Izvekova GI, Gisbert E, Glupov VV (2014) Feeding habits and ontogenic changes in digestive enzyme patterns in five freshwater teleosts. J Fish Biol 85(5):1395–1412

    Article  CAS  PubMed  Google Scholar 

  • Sorensen SH, Noren O, Sjöström H, Danielsen M (1982) Amphiphilic pig intestinal microvillus maltase/glucoamylase. Structure and specificity. Eur J Biochem 126:559–568

    Article  CAS  PubMed  Google Scholar 

  • Stefansson B, Helgadóttir L, Olafsdottir S, Gudmundsdottir A, Bjarnason JB (2010) Characterization of cold-adapted Atlantic cod (Gadus morhua) trypsin I—kinetic parameters, autolysis and thermal stability. Comp Biochem Physiol B: Biochem Mol Biol 155:186–194

    Article  Google Scholar 

  • Stiefel JS, Keller PJ (1973) Preparation and some properties of human pancreatic amylase including a comparison with human parotid amylase. BBA 302:345–361

    CAS  PubMed  Google Scholar 

  • Tamiya T, Okahashi N, Sakuma R, Aoyama T, Akahane T, Juichiro JM (1985) Freeze denaturation of enzymes and its prevention with additives. Physiol Rev 22:446–456

    CAS  Google Scholar 

  • Ugolev AM, Egorova VV, Kuz’mina VV, Grusdkov AA (1983) Comparative-molecular characterization of membrane digestion in fish and mammals. Comp Biochem Physiol B: Biochem Mol Biol 76(3):627–635

    CAS  Google Scholar 

  • Worthington Biochemical Corporation (1972) Worthington enzyme manual: enzymes, enzyme reagents, related biochemicals. Worthington Biochemical Corp., Freehold

    Google Scholar 

  • Wu G-P, Cao M-J, Chen Y, Liu B-X, Su W-J (2008) Leucine aminopeptidase from Red sea bream (Pagrus major) skeletal muscle: purification, characterization, cellular location, and tissue distribution. J Agric Food Chem 56:9653–9660

    Article  CAS  PubMed  Google Scholar 

  • Xiong YL (1997) Protein denaturation and functionality losses. In: Erickson MC, Hung Y-C (eds) Quality in frozen food. Chapman & Hall, New York, pp 111–141

    Chapter  Google Scholar 

Download references

Author information

Author notes

    Authors and Affiliations

    1. Institute of Systematics and Ecology of Animals Siberian Branch of Russian Academy of Sciences, 11 Frunze St., Novosibirsk, 630091, Russia

      Mikhail Solovyev

    2. Centre de Sant Carles de la Ràpita, Institut de Recerca i Tecnologia Agroalimentaries (IRTA), Crta. Poble Nou del Delta km 5.5, 43540, Sant Carles de la Ràpita, Spain

      Enric Gisbert

    Authors
    1. Mikhail Solovyev
      View author publications

      You can also search for this author in PubMed Google Scholar

    2. Enric Gisbert
      View author publications

      You can also search for this author in PubMed Google Scholar

    Corresponding author

    Correspondence to Mikhail Solovyev.

    Additional information

    Mikhail Solovyev and Enric Gisbert have contributed equally to the study.

    Rights and permissions

    Reprints and permissions

    About this article

    Check for updates. Verify currency and authenticity via CrossMark

    Cite this article

    Solovyev, M., Gisbert, E. Influence of time, storage temperature and freeze/thaw cycles on the activity of digestive enzymes from gilthead sea bream (Sparus aurata). Fish Physiol Biochem 42, 1383–1394 (2016). https://doi.org/10.1007/s10695-016-0226-2

    Download citation

    • Received:

    • Accepted:

    • Published:

    • Issue Date:

    • DOI: https://doi.org/10.1007/s10695-016-0226-2

    Keywords

    Search

    Navigation