Antarctic krill, Euphausia superba is one of the promising sources of protein. Proper utilisation of this valuable resource is of extreme importance at a time when most of the conventional resources are reaching or exceeding optimum sustainable limits. One of the main problems in utilising krill is the intense postmortem proteolytic activity. High fluoride content, excessive loss of exudates during storage at non-freezing temperature, blackening and development of unpleasant flavour are the other problems observed during krill processing. Several investigations have been carried out to study the processing aspects and biochemical characteristics of krill (Budzinski et al. 1985). The present work was carried out to study the various aspects of krill processing and product development, besides assessing its biochemical changes, changes in fluoride content and other physical parameters during storage.
Materials and Methods
The krill samples were collected from the Southern Ocean between 57° 53’ -61° 13’S and 31° 40’ -36° 31’ E, on board FORV Sagar Sampada. Only non-feeding and partially fed krill, which were pinkish red in colour, with an average length of 35 mm and above were taken for processing. Feeding krill (green in colour) were removed prior to processing. The duration of trawling was reduced in order to reduce the quantity of catch so as to minimise physical damage. Freshly caught krill was processed immediately to minimise deterioration that could set in after four hours of harvest. The harvested krill was cleaned, weighed and packed in cartons lined with polyethylene sheets, frozen at -35°C and then stored at -30°C for further studies. Apart from this, fresh whole krill was processed to mince, tail meat, coagulate and boiled krill, and then frozen and stored as above. The mince was prepared using Baader 694 meat bone separator. The tail meat was prepared using prototype roller peeling machine .
Krill body proportions comprised of cephalothorax (32-36 per cent) abdomen (26-28 per cent) and carapace (25-27 per cent) of the body weight. The remaining 10-18 per cent was lost on separation. The yield of whole boiled krill, headless krill, tail meat, krill mince and krill coagulate were 80,65,12, 75 and 40 per cent respectively.
The proximate composition of the samples was determined by Association of Official Analytical Chemist (AOAC) (2000) method. Non protein nitrogen (NPN) and fluoride content were determined according to the methods of AOAC (1990). Total volatile base nitrogen (TVB-N) and trimethylamine (TMA) were estimated by the micro diffusion method (Conway, 1950). The α-Amino nitrogen (α -AN) was estimated as per the method of Pope and Stevens (1939). Heavy and trace metals in whole krill and krill head were determined by atomic absorption spectrometry (Radhakrishnan,1993a) and the presence of pesticide residues in the same were analysed by gas chromatography (Radhakrishnan, 1993b).
The proximate composition of krill samples are given in the Table 1. The moisture and fat content of krill was found to be high with low crude protein, as also reported by Grantham, 1977. The fluoride content of whole krill and in the different portions are given in Table 2. An increase of up to 44 per cent of fluoride content in the tail meat was observed during the frozen storage at -30°C for 3 months. Similar observations were made by Christians and Leinemann, 1980 and Christians et al., 1981. The changes in fluoride content of other products from krill on storage were marginal. Bykowski et al., 1981 and Boone and Manthey, 1983 made similar observations. Because of the migration of fluoride from the shell to the meat, it is recommended that the whole catch may be converted to intermediate products like tail meat, mince, coagulate etc., which can be used for preparation of several products for human consumption.
The concentration of various trace metals in whole krill and head are given in Table 3. The concentrations of copper, iron, zinc, chromium were above 10 ppm while those of lead, cadmium, nickel and manganese were less than 1 ppm. Marine crustaceans in Antarctic Ocean have been reported to accumulate as much as 13 mg/kg (dw) of cadmium (Petri and Zauke, 1993). Chinese expedition to Antarctic waters found that E. superba does accumulate a number of trace metals such as Pb, Cu, Cd, Zn, Fe and Mn. The maximum accumulation was in the case of copper and minimum in lead (Qun et al., 1990). Hence the higher concentrations of copper, iron and zinc obtained in the present study are in agreement with previously reported values. The concentrations of pesticides in krill were all below detectable limits and no traces of any organochlorine pesticides were found in any of the samples. The absence of pesticides in Antarctic krill samples analysed in the present study may be probably due to the difference in the areas from which the samples were taken.
Fatty acid analysis
Krill lipids were extracted from samples with chloroform-methanol mixture as per Folch et al., 1957. Cholesterol in the unsaponifiable fraction was estimated by the ferric chloride method (Rudel and Morris, 1973). Methyl esters of fatty acids in the saponifiable fraction were prepared (Metcalfe et al., 1966) and analysed on a Chrompack CP 9001 gas chromatograph equipped with an Alltech AT225 capillary column (0.53 mm id and 30 m length) and flame ionisation detector.
Eighty per cent of krill lipids are reported to be phospholipids (Kolakowska, 1988). The proportion of unsaponifiable fraction of lipids was 7.26 per cent, which was comparable with earlier findings (Watanabe et. al.,1976). The cholesterol concentration was 101.7 mg per 100 g of whole krill and 33.4 mg/100 g of tail meat. The concentration of various fatty acids in the saponifiable fraction of krill lipids and cholesterol are given in Table 4. Thirty five per cent of the fatty acids in krill tail meat were saturated, 19 per cent monounsaturated and 36 per cent polyunsaturated (Table 5). Among the saturated fatty acids, palmitic acid (C16:0) was the dominant fatty acid amounting to 28 per cent. Oleic acid (C18:1 n-9), eicosapentaenoic acid (C20:5 n-3) and docosahexaenoic acid (C22:6 n-3) were the major unsaturated fatty acids contributing 12, 25 and 10 per cent, respectively, of the total fatty acids. Palmitic acid, eicosapentaenoic acid and docosahexaenoic acid together accounted for about 60 per cent of the total fatty acids. Stearic acid (C18:0) was not present in detectable levels. Krill lipids on the whole have fatty acid compositions similar to fish lipids, except for the near absence of C18:0.
Amino acid analysis
Amino acid concentration was determined by Ishida and others (1981) method. Amino acid profile of proteins was analysed after hydrolysing the samples in 6M HCl for 24 h at 110°C in evacuated sealed tubes. Samples were injected in to high-performance liquid chromatography (Shimadzu Amino Acid Analyser LCIOAS Model) after filtering through 0.45 µm syringe filter. Separation and quantification of amino acids was carried out with a cation exchange column. Samples were filtered and injected appropriate quantities in to the HPLC system as per the specification of the injector. The eluted amino acids were derivatised post column with o-phthalaldehyde (for fluorescence detector) and hypochlorite for imino acids. Tryptophan was determined after alkaline hydrolysis with 5 per cent (w/v) NaOH (Sastry and Tammuru, 1985). Nutritional studies of the krill meat were carried out as per the method described by Raghunath et al., 1995.
The amino acid profile of whole krill, krill tail meat and krill mince are given in Table 6. Protein of the krill tail meat appeared to be quite balanced in its amino acid composition. All the essential amino acids were present in adequate amounts. Whole krill had a slightly lower concentration of sulphur containing amino acids like cysteine and methionine. The nutritional parameters of krill tail meat are given in Table 7. Krill meat was evidently a good dietary source of protein. Experimental feeding trials in albino rats indicated that there were no ill effects associated with dietary intake of krill in relation to hyperlipidemia.
Various products were developed from krill to find out suitable methods to utilise the krill meat. The whole krill was dried to find out the acceptability of dried krill after partial thawing by boiling in 0,1,2 and 5 per cent brine (1:1 ratio of krill and brine) for 5 minutes. The thawed krill was drained and dried under sun. After drying, the head, tail and legs were removed and packed in polyethylene bags. The samples were then stored for a period of 3 months at ambient temperature (±28°C) for shelf life studies. The dried samples were analysed for total nitrogen (TN), non protein nitrogen (NPN), moisture, fat, ash, salt (AOAC, 2000) and total volatile basic nitrogen (TVN) (Conway, 1947).
Another attempt was made to prepare surimi from krill by mixing it with Catla catla mince. Fresh C. catla were obtained from a local fish farm at Cochin and brought to the laboratory in iced condition without delay. Upon arrival at the laboratory the samples were de-iced and washed with chilled (4oC) potable water. The samples were then filleted and minced in a meat bone separator. The minced meat was washed in chilled water and surimi was prepared as described by Muraleedharan, et al., 1997. The mince from krill was mixed with washed C. catla mince at 10, 20, 30 and 40 per cent (w /w) proportions in a Hobart mixer for 25 min at 15°C and the chemical, physical and organoleptic properties of the mixtures were studied. Moisture, total nitrogen, fat and ash contents of the prepared surimi were determined by the AOAC (2000) method. Salt soluble nitrogen (SSN) was analysed by the method of Dyer et al., 1950.
Fish paste was prepared by incorporating 10, 20 and 30 per cent (w/w) krill mince with C. catla and made into a paste using a silent cutter. The sensory quality aspects and spreadability were studied as described by Muraleedharan, et al., 1996. Value added fish products such as fish burger, fish cutlets (Joseph et al.,1984) and fish wafers (Shenoy et. al., 1983) were also prepared by incorporating cooked krill mince at 10, 20, 30 and 40 per cent with cooked Nemipterus japonicus meat. Sensory characteristics of the products were evaluated using a trained taste panel consisting of ten members.
The yield of dried krill meat varied from 6 to 10 per cent of the initial wet krill samples. The krill cooked in 5 per cent brine showed the highest with samples cooked in water showed the least. The dried product prepared by cooking in water became powdery after two months of storage while the product cooked in salt solution did not show any change in their shape and physical characteristics. The dried products prepared after cooking in 1 and 2 per cent salt solution had a dark pinkish appearance while the samples cooked in 5 per cent brine showed a light pinkish appearance.
The chemical composition of dried krill products are given in the Table 8. The product cooked in water and dried had high total nitrogen and 45 per cent of the TN was NPN. The TVN value was also very high. The TVN contents of the other three samples were below 100mg/100g. The salt content of the dried product showed a proportional increase depending on the salt content of the brine used for cooking.
Table 9 shows the change in the chemical, physical and sensory quality of C. catla mince incorporated krill meat. Colour of the mince increased from off-white to pink and red upon increasing the proportion of krill meat. Odour also changed to unpleasant upon increasing the concentration of krill meat. The soft texture of C. catla surimi became fibrous coarse by incorporation of krill mince at a rate of 40 per cent and above. The deformation of the heat set gel on application of external force decreased with corresponding drop in the rupture force, indicating appreciable decrease of gel strength. Heat set salt stabilised krill mince was found to be a very weak. Hence, krill mince alone could not be subjected to gel strength measurement. This clearly shows that the myofibrillar proteins in the krill meat have undergone significant hydrolysis during three months of storage, making it unsuitable for surimi preparations. The studies also indicated that the incorporation of krill meat in C. catla mince resulted in significant reduction of quality of the mince. The sensory characteristics of different products prepared by incorporating krill mince to fish meat are given in Table 10. The acceptability of the products reduced with an increase in krill meat concentration in the product.
Based on the results it is suggested that the whole catch may be immediately converted into intermediate products like head less, tail meat, mince, coagulate etc., to reduce the migration of fluoride from the shell to the meat. The biochemical composition of the samples showed that krill meat is rich in C18:1, C20:5, and C22:6 fatty acids. Protein of krill meat is balanced in its amino acid composition and all the essential amino acids are in adequate amounts. Various value added products can be prepared by incorporating krill mince to the fish mince in lower quantities. The acceptability of products reduced as a result of increasing the krill concentrations in the fish mince.
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