Studies on fish lipases

Lipids constitute a significant portion of the biomass of earth and lipolytic enzymes play a very important role in lipid turn over. Apart from their biological significance, lipolytic enzymes are also very important in the fields of nutrition, food technology, medicine and preparative and analytical lipid biochemistry. Recent developments in the study of proteins and enzymes have largely benefited the study of lipolytic enzymes, that some of these enzymes were isolated in pure form. Even today there is a continuous search for new and potent sources of these lipolytic enzymes. The zest for elucidating the structure and mechanism of action of the enzymes obtained in pure form for biochemist still remains unabated. The literature shows no record of such an effort for the study of lipases from marine sources. The fact that many fishes like oil sardine, mackerel, cat fish, seer etc. contains large amounts of lipid shows the possibility of the existence of lipases in significant amounts necessitating their exhaustive study. Such a study will, not only provide alternate sources for lipase but also will provide methods to curb lipolysis and the resultant rancidity and off flavor development in fish and fishery products.

Partial purification, immobilization and preliminary biochemical characterization of lipases from Rhizomucor pusillus

Lipases have important applications in biotechnological processes, motivating us to produce, purify, immobilize and perform a biochemical characterization of the lipase from Rhizomucor pusillus. The fungus was cultivated by solid state fermentation producing lipolytic activity of about 0.5 U/mL(4U/g). A partial purification by gel filtration chromatography in Se-phacryl S-100 allowed obtaining a yield of about 85% and a purification factor of 5.7. Our results revealed that the purified enzyme is very stable with some significant differences in its properties when compared to crude extract. The crude enzyme extract has an optimum pH and temperature of 7.5 ° C and 40 ° C, respectively. After purification, a shift of the optimum pH from 7 to 8 was observed, as well as a rise in optimumtemperature to 60 ° C and an increase in stability. The enzyme was immobilized on CNBr-Agarose and Octyl-Agarose supports, having the highest immobilization yield of 94% in the second resin. The major advantage of immobilization in hydrophobic media such as Octyl is in its hyper activation, which in this case was over 200%, a very interesting finding. Another advantage of this type of immobilization is the possibility of using the derivatives in biotechnological applications, such as in oil enriched with omega-3 as the results obtained in this study display the hydrolysis of 40% EPA and 7% DHA from sardine oil, promising results compared to the literature.

Modulation of the activity and selectivity of the immobilized lipases by surfactants and solvents

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Value-adding Australian sardines: Factors affecting rates of deterioration in sardine (Sardinops sagax) quality during post-harvest handling

Deficiencies in sardine post-harvest handling methods were seen as major impediments to development of a value-adding sector supplying Australian bait and human consumption markets. Factors affecting sardine deterioration rates in the immediate post-harvest period were investigated and recommendations made for alternative handling procedures to optimise sardine quality. Net to factory sampling showed that post-mortem autolysis was probably caused by digestive enzyme activity contributing to the observed temporal increase in sardine Quality Index. Belly burst was not an issue. Sardine quality could be maintained by reducing tank loading, and rapid temperature reduction using dedicated, on-board value-adding tanks. Fish should be iced between the jetty and the processing factory, and transport bins chilled using an efficient cooling medium such as flow ice.

Triacylglycerol lipases of yeast and plants: more than just hydrolases

In the yeast, mobilization of triacylglycerols (TAG) is facilitated by TGL3, TGL4 and TGL5 gene products. Interestingly, experiments using [32P] orthophosphate as a precursor for complex glycerophospholipids revealed that tgl mutants had a lower steady-state level of these membrane lipids. To understand a possible link between TAG lipolysis and phospholipid metabolism, we performed overexpression studies with Tgl3p and Tgl5p which clearly demonstrated that these two enzymes enhanced the level of phospholipids. Domains and motifs search analyses indicated that yeast TAG hydrolases posses a GXSXG lipase motif but also a HX4D acyltransferase motif. Purified Tgl3p and Tgl5p did not only exhibit TAG lipase activity but also catalyzed acyl-CoA dependent acylation of lyso-phosphatidylethanolamine and lyso-phosphatidic acid (LPA), respectively. Search for lipase/hydrolase homologues in the Arabidopsis thaliana genome led to the identification of At4g24160 which possess three motifs that are conserved across the plant species such as GXSXG motif, a HX4D motif and a probable lipid binding motif V(X)3HGF. Characterization of At4g24160 expressed in bacteria revealed that the presence of an acyl-CoA dependent LPA acyltransferase activity. In addition, the purified recombinant At4g24160 protein hydrolyzed both TAG and phosphatidylcholine. We hypothesize that the plant enzyme may be involved in membrane repair. In summary, our results indicate that these TAG lipases play a dual role and thereby contribute to both anabolic and catabolic processes in yeast and plants.

Cruise Report 51-S-1 of N.B. Scofield: Number S-21 of the Cooperative Sardine Research Program

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Economic Effects of Global Warming under Stock Growth Uncertainty: The European Sardine Fishery

Global warming of the oceans is expected to alter the environmental conditions that determine the growth of a fishery resource. Most climate change studies are based on models and scenarios that focus on economic growth, or they concentrate on simulating the potential losses or cost to fisheries due to climate change. However, analysis that addresses model optimization problems to better understand of the complex dynamics of climate change and marine ecosystems is still lacking. In this paper a simple algorithm to compute transitional dynamics in order to quantify the effect of climate change on the European sardine fishery is presented. The model results indicate that global warming will not necessarily lead to a monotonic decrease in the expected biomass levels. Our results show that if the resource is exploited optimally then in the short run, increases in the surface temperature of the fishery ground are compatible with higher expected biomass and economic profit.

Síntese de sebacato de dioctila empregando lipases

A crescente demanda por lubrificantes obtidos a partir de fontes renováveis vem incentivando a pesquisa por alternativas sustentáveis. O objetivo principal deste trabalho foi investigar a síntese de sebacato de dioctila a partir da reação de esterificação entre o ácido sebácico e o 1-octanol empregando biocatalisadores e catalisador químico convencional (ácido sulfúrico). Alguns parâmetros reacionais foram estudados: tipo de lipase comercial imobilizada (Novozym 435, Lipozyme RM IM e Lipozyme TL IM), temperatura, razão molar ácido/álcool, concentração de lipase, métodos de remoção da água do meio reacional. A reutilização da lipase Novozym 435 também foi avaliada. A conversão da reação foi determinada por cromatografia em fase gasosa. A lipase Novozym 435 apresentou os melhores resultados: 100% de conversão de ácido sebácico quando foi empregada razão molar ácido:álcool de 1:5 e 5% m/m de lipase, após 150 minutos de reação a 100C. O emprego de peneira molecular e vácuo não aumentou a conversão do ácido sebácico. O produto final foi caracterizado com relação à viscosidade, ao índice de viscosidade, ao ponto de fulgor, ao ponto de fluidez e ao índice de neutralização, e apresentou comportamento semelhante a um óleo naftênico

Prediction and confirmation of seasonal migration of Pacific sardine (Sardinops sagax) in the California Current Ecosystem

During the last century, the population of Pacific sardine (Sardinops sagax) in the California Current Ecosystem has exhibited large fluctuations in abundance and migration behavior. From approximately 1900 to 1940, the abundance of sardine reached 3.6 million metric tons and the “northern stock” migrated from offshore of California in the spring to the coastal areas near Oregon, Washington, and Vancouver Island in the summer. In the 1940s, the sardine stock collapsed and the few remaining sardine schools concentrated in the coastal region off southern California, year-round, for the next 50 years. The stock gradually recovered in the late 1980s and resumed its seasonal migration between regions off southern California and Canada. Recently, a model was developed which predicts the potential habitat for the northern stock of Pacific sardine and its seasonal dynamics. The habitat predictions were successfully validated using data from sardine surveys using the daily egg production method; scientific trawl surveys off the Columbia River mouth; and commercial sardine landings off Oregon, Washington, and Vancouver Island. Here, the predictions of the potential habitat and seasonal migration of the northern stock of sardine are validated using data from “acoustic–trawl” surveys of the entire west coast of the United States during the spring and summer of 2008. The estimates of sardine biomass and lengths from the two surveys are not significantly different between spring and summer, indicating that they are representative of the entire stock. The results also confirm that the model of potential sardine habitat can be used to optimally apply survey effort and thus minimize random and systematic sampling error in the biomass estimates. Furthermore, the acoustic–trawl survey data are useful to estimate concurrently the distributions and abundances of other pelagic fishes.

Distributions and abundances of Pacific sardine (Sardinops sagax) and other pelagic fishes in the California Current Ecosystem during spring 2006, 2008, and 2010, estimated from acoustic–trawl surveys

The abundances and distributions of coastal pelagic fish species in the California Current Ecosystem from San Diego to southern Vancouver Island, were estimated from combined acoustic and trawl surveys conducted in the spring of 2006, 2008, and 2010. Pacific sardine (Sardinops sagax), jack mackerel (Trachurus symmetricus), and Pacific mackerel (Scomber japonicus) were the dominant coastal pelagic fish species, in that order. Northern anchovy (Engraulis mordax) and Pacific herring (Clupea pallasii) were sampled only sporadically and therefore estimates for these species were unreliable. The estimates of sardine biomass compared well with those of the annual assessments and confirmed a declining trajectory of the “northern stock” since 2006. During the sampling period, the biomass of jack mackerel was stable or increasing, and that of Pacific mackerel was low and variable. The uncertainties in these estimates are mostly the result of spatial patchiness which increased from sardine to mackerels to anchovy and herring. Future surveys of coastal pelagic fish species in the California Current Ecosystem should benefit from adaptive sampling based on modeled habitat; increased echosounder and trawl sampling, particularly for the most patchy and nearshore species; and directed-trawl sampling for improved species identification and estimations of their acoustic target stren

Otolith morphometrics and population structure of Pacific sardine (Sardinops sagax) along the west coast of North America

The broad distribution of Pacific sardine (Sardinops sagax) along the Pacif ic coast of North America makes it difficult for fisheries managers to identify regional stocks of this dominant small pelagic species. An investigation of morphometric characteristics of otoliths of Pacific sardine across most of their range revealed regional differences in populations. In a survey of over 2000 otoliths, all ages (with an emphasis on age-1 recruits) were compared. Principal components analysis, multivariate analysis of variance, and a novel method derived from regression and residuals calculations, termed perimeter-weight profiles (PWPs), revealed otolith similarities and differences. The results of the different approaches to statistical comparisons did not always agree. Sardine otoliths from Mexican waters were generally lighter and more lobate than those from U.S. and Canadian populations. Age-1 otoliths from northern California in 2006–07 tended to be heavier and smoother than those from other areas, including year-class cohorts from southern California. Comparisons of age-groups and year-classes of northern California otoliths with the use of the PWP models indicated signif icant trends in year-to-year patterns. In conjunction with other established indices of population structure, otolith PWPs are a useful tool for identifying local and regional stocks of Pacific sardine and may help distinguish populations of other fish species as well.

Biomass and reproduction of Pacific sardine (Sardinops sagax) off the Pacific northwestern United States, 2003–2005

The Pacific sardine (Sardinops sagax) is distributed along the west coast of North America from Baja California to British Columbia. This article presents estimates of biomass, spawning biomass, and related biological parameters based on four trawl-ichthyoplankton surveys conducted during July 2003 –March 2005 off Oregon and Washington. The trawl-based biomass estimates, serving as relative abundance, were 198,600 t (coefficient of variation [CV] = 0.51) in July 2003, 20,100 t (0.8) in March 2004, 77,900 t (0.34) in July 2004, and 30,100 t (0.72) in March 2005 over an area close to 200,000 km2. The biomass estimates, high in July and low in March, are a strong indication of migration in and out of this area. Sardine spawn in July off the Pacific Northwest (PNW) coast and none of the sampled fish had spawned in March. The estimated spawning biomass for July 2003 and July 2004 was 39,184 t (0.57) and 84,120 t (0.93), respectively. The average active female sardine in the PNW spawned every 20–40 days compared to every 6–8 days off California. The spawning habitat was located in the southeastern area off the PNW coast, a shift from the northwest area off the PNW coast in the 1990s. Egg production in off the PNW for 2003–04 was lower than that off California and that in the 1990s. Because the biomass of Pacific sardine off the PNW appears to be supported heavily by migratory fish from California, the sustainability of the local PNW population relies on the stability of the population off California, and on local oceanographic conditions for local residence.

Degeneration of postovulatory follicles in the Iberian sardine Sardina pilchardus: structural changes and factors affecting resorption

Inaccuracy in the aging of postovulatory follicles (POFs) and in estimating the effect of temperature on the resorption rate of POFs may introduce bias in the determination of the daily spawning age classes with the daily egg production method (DEPM). To explore the above two bias problems with f ield-collected European pilchard (Sardina pilchardus, known regionally as the Iberian sardine), a method was developed in which the time elapsed from spawning (POF age) was estimated from the size of POFs (i.e., from the cross-sectional area in histological sections). The potential effect of the preservative type and embedding material on POF size and the effect of ambient water temperature on POF resorption rate are taken into account with this method. A highly significant loglinear relationship was found between POF area and age; POF area shrank by approximately 50% per day. POFs were also shown to shrink faster at higher temperatures (approximately 3% per degree), but this temperature effect is unlikely to be an important source of bias in the assignment of females to daily spawning classes. The embedding material was also shown to influence the size of POFs, the latter being significantly larger in resin than in paraffin sections. In conclusion, the size of POFs provides an indirect, reliable estimation of the time elapsed from spawning and may thus be used to test both the validity of POF staging criteria for identifying daily classes of spawners and the effect of other factors (such as temperature and laboratory processing) in applications of the DEPM to S. pilchardus and other fish species.