Angiotensin-I converting enzyme inhibitory (ACEi) peptides were prepared from rohu (Labeo rohita) fish wastes by enzymatic hydrolysis [1.1% alcalase concentration (v/w); 130 min time; 52°C temperature and 0.8:1 solid-liquid ratio]. Ultrasound Assisted Enzymatic Extraction (UAEE) and Microwave Assisted Enzymatic Extraction (MAEE) were performed to enhance degree of hydrolysis (DH), ACEi activity, and peptide yield (PY). The ACEi peptides were examined for stability during thermal processing, varied pH treatments and in vitro gastrointestinal digestion. Peptides were stable at different temperatures (25, 37, 55, 75, and 100°C) except at 121°C which reduced their stability by 7%, 6.9% and 2.2% in UAEE, MAEE and Non-Assisted Enzymatic Extraction (NAEE) peptides, respectively. Similarly, peptides retained ACEi at pH 2 to 8 and subsequent reduction at pH>8. In vitro digestion studies showed stability of all peptides to GI digestion at 0.05% pepsin, 0.05% trypsin, 0.05% chymotrypsin and the combination of 0.025% trypsin and 0.025% chymotrypsin except MAEE peptides showing less stability with pepsin (0.05%). Therefore, it is concluded that these ACEi peptides derived from rohu fish waste, stable to heat, pH and gastrointestinal enzymatic action, can be used as ingredient in functional foods, which undergo thermal processing and digestion.

The presence of a significant amount of toxic metal in seafood has raised human health concerns worldwide. This study investigates the effect of heat processing/cooking methods (boiling, frying, grilling, and microwave cooking) on toxic metals concentration in farmed shrimp and estimates its potential human health risks. Pacific white leg shrimps (Penaeus vannamei) farmed along the Gulf of Mannar region were used to evaluate the toxic metal uptake by consuming raw and cooked shrimps. Toxic heavy metals such as Cd, Hg, and Pb were found below the maximum residual limit (MRL) of 0.5 mg/kg set by the European Union. The concentration of Cu, Zn, and As were below the MRLs of 30 mg/kg, 100 mg/kg, and 76 mg/kg set by the World Health Organization and Food Safety and Standard Authority of India, respectively. The target hazard quotient (THQ) for individual metals and total THQ (TTHQ) for combined metals were 1, indicating no health risk for shrimp consumers. The lifetime cancer risk (LCR) for toxic heavy metals (Cr, Ni, As, Sr, Cd, and Pb) were found between the recommended level of 10-6 to 10-4, indicating no carcinogenic health consequences. With the estimated maximum allowable consumption rates (CRmm), an adult can safely consume 30 raw or cooked shrimp meals/month with no adverse health consequences. Therefore, culinary processes such as boiling and grilling are recommended as suitable methods to reduce toxic metals risk. However, it is recommended to monitor their concentration before and after cooking shrimps collected worldwide and ensure they are within the regulatory limits.

Anti-hypertensive [angiotensin-I converting cnzyme (ACE) inhibitory (ACEi) peptides are enzymatically extracted from biological material. The high cost of enzymes, low peptide yield and long hydrolysis time in enzymatic extraction paved the way for alternative ultrasound assisted enzymatic extraction (UAEE) and microwave-assisted enzymatic extraction (MAEE). In this study, rohu (Labeo rohita) fish wastes were homogenised and treated with ultrasound (40 kHz) and microwave (2450 MHz) before enzymatic hydrolysis using optimised concentration of alcalase® for the extraction of ACEi. The UAEE (sonication temperature: 30-70; optimum: 57.88ºC and time: 10-50; optimum: 63.4 min) and MAEE (microwave power: 180-900, optimum: 335.23 W and time 5-25, optimum: 15 min) were studied for their effect on degree of hydrolysis (DH), ACE inhibition (ACEi) and peptide yield (PY). The efficiency obtained by UAEE (5%) was higher than MAEE (4%) in terms of ACEi. In contrast, MAEE showed higher efficiency with respect to DH and PY (7 and 6.5 %, respectively) than UAEE (2.5 and 4%, respectively). The study indicated that UAEE and MAEE are efficient methods for extraction of ACE-inhibitory peptides from rohu fish waste. However, UAEE peptides shows better ACEi than MAEE. Keywords: ACE-inhibitory peptide, Alcalase, Degree of hydrolysis, Enzymatic extraction, Peptide yield, Rohu processingwaste

Background: Seafood analogue is a ready to make value added product prepared out of surimi. Thermal processing of restructured products in retort pouches such as fish ball in curry medium, surimi stew in white tripod, boneless rohu balls in curry. Also no work had done on thermal processing of shrimp analogue products in retort pouches. The main objective of this work was to develop the analogue shrimp product from lizardfish and to compare the heat penetration attributes of analogue shrimp curry and masala using retort pouches and different sterilization methods such as steam / air over pressure retort and water immersion retort.Methods: Analogue shrimp products were prepared and thermally processed in retortable pouches. About 125g of shrimp analogue product and 100g of curry (masala) were filled in retort pouches of size, 150x200mm. Air inside the pouch was exhausted by steam injection followed by heat sealing and processing at 121.1°C in a retort by steam/air over pressure retort and water immersion retort. The difference in the heat penetration characteristics of analogue shrimp products processed in retort by steam/air over pressure retort and water immersion retort were studied. Result: The results showed that minimum heating lag factor and minimum come up time led to faster heating rate which decreased total process time in imitated shrimp curry by steam/ air retort. At the same time the cook value was low in curry medium processed by steam air retort. So finally conclude that imitated shrimp curry processed by steam air retort was good.

Extraction of gelatin from fish wastes and the development of artificial fish bait matrix using the extracted gelatin were attempted in the study. Biochemical composition of fish wastes (heads, skins, fins and scales) and the gelatin derived from the wastes of corresponding body parts and physicochemical properties of fish gelatin were analyzed. Considering high yield (19.8–22.4%), good gel strength (1.99–2.3 N), high melting point (27 °C) and simple extraction process, the scale gelatin was selected for the development of artificial fish bait matrix. To optimize the levels of gelling agent, cross linker and solvent for the preparation of bait matrix, RSM was adopted. Gel strength and insolubility ratio were analyzed to know the suitability of the bait. They were found to have the gel strength ranging from 1.3 N to 23.5 N, insolubility ratio from 55–83%, and melting point from 34.3 °C to 41.1 °C. The optimum levels of ingredients for the preparation of ideal bait matrix were standardized as: gelatin-25.5%, sucrose-22.5% and water-52%. The curing temperature and curing duration were optimized based on the optimum gel strength of 20.84 N and insolubility ratio of 74.5% was 5 °C and 25 h respectively. The study revealed that a developed fish bait matrix suitable for fishing can be made from gelatin derived from fish wastes.

The aim of the study has been to develop a biodegradable film from marine polysaccharides. The optimization of polysaccharides quantity for the composite film was sought by empirical response surface methodology. The Box–Behnken Model Design was applied to optimize the concentration of chitosan (1.0-2.0% (w/v), agar (1.0-2.0% (w/v) and glycerol (0.1-0.5% (w/v) as independent variables to achieve the goal. The overall desirability function fits with the quadratic model (0.862043) at a significant level (p < 0.05) for the optimum concentration of chitosan (1.5% (w/v), agar (2.0% (w/v) and glycerol (0.41% (w/v) to obtain the minimum water vapor permeability (7.25 10-10g m m-2 Pa-1 s-1) and maximum tensile strength (12.21 Ma P), elongation at break (7.32%) and puncture resistance (16.18 N) in the optimized composite film. The absolute residual errors of experimental and predicted responses were between 1.24 and 3.56% acceptable levels. Attenuated total reflection–Fourier transform infrared spectroscopy confirmed the intermolecular non-covalent hydrogen bond between the hydroxyl groups of agar and glycerol with the amino group of chitosan. 3D atomic force microscopy images revealed that the chitosan, agar and glycerol film has layer-by-layer smooth surface properties due to homogenous interaction among the polysaccharides; this provides the film with good mechanical properties and with functional application. Chitosan was found to be responsible for the lower level of water vapor permeability and higher puncture resistance of the film. Tensile strength and elongation at break were influenced by agar and glycerol. The whiteness of the film was negatively affected with the concentration of chitosan.

The present study intended to develop improved biopolymer film from seaweed polysaccharides. The quantity optimization of polysaccharides for the composite film was sought by empirical response surface methodology. To achieve the goal, Box–Behnken model was applied to the concentration of three independent variables, viz. agar (1.0–2.0% w/v), alginate (1.0–2.0% w/v) and carrageenan (1.0–2.0% (w/v). The glycerol was used as a plasticizer and kept constant (25% w/w) for total solid mass. The overall desirability function fits with the quadratic model at 99.78% level of significance for the optimization of agar (1.99% w/v), alginate (1.45 w/v) and carrageenan (2.0% w/v) to reach minimum water vapor permeability and maximum tensile strength, elongation at break and puncture resistance. The absolute residual error (1.04–3.37%) of experimental and predicted response was also validated. Attenuated total reflection-Fourier transform infrared spectroscopy confirmed the interactions such as stretching at 2900 cm−1 region corresponded to C–H stretching vibration and an intensity peak observed at 1200 cm−1 of AAC film corresponded to sulfate ester groups. The shift in crystalline nature of composite film was confirmed by XRD. The 3D image of atomic force microscopy showed layer-by-layer assembly of intermolecules at 310-nm resolution, and the characterized smooth surface has more functional application. The carrageenan and agar are found to be more responsible for the film properties such as moisture content, thickness, whiteness index, transparency, swelling and erosion than alginate.

A PCR-RFLP was developed by targeting the mitochondrial 12S rRNA region to authenticate the six species viz., Lutjanus fulvus, L. rivulatus, L. quinquelineatus, L. fulviflanuna, L. madras and L. decussatus available along the Indian coast. A newly designed primer set, 12SU-F/12SU-R, was used for PCR amplification to obtain a product size of 550 bp, which was then digested using three endonucleases viz., BseDI, MnlI and MspAll. All the six species were unambiguously differentiated by BseDI enzyme. MnlI differentiated only four species viz., L fulvus, L. quinquelineatus, L. fulvifiamma and L. madras; while MspAI differentiated four species viz., L. rivulatus, L. quinquelineatus, L. fulviflamma and L. madras from the rest. The developed PCR-RFLP protocol using BseDI enzyme can therefore be easily adopted by regulatory authorities to the prevention of possible adulteration of species for red snappers.

An attempt was made to explore the possibilities of better utilization of Lizard fish (Saurida tumbil) for the development of valuable imitation shrimp products. Minced meat, surimi and imitation shrimp products were prepared from Lizard fish. The aluminum and polylactic acid shrimp moulds were developed and to produce the imitation shrimp products. In the present study, the imitated shrimp products was observed for the wholesomeness.

PCR-SSCP and PCR-FINS methods were developed to authenticate nine species of snapper. A highly variable mitochondrial D-loop region was chosen as a target region and three new primers viz., one forward (Fish-DL-F) and two reverse primers (Fish-DL-R and DLFR) were designed. PCR-SSCP was first targeted to amplify 515 bp D-loop fragment using Fish-DL-F/Fish-DL-R primers, but it could not differentiate all the species. Another reverse primer (DLFR) could amplify a 360 bp region, which had differentiated all the species. A FINS method was also developed for snapper authentication using the same D-loop region of 515 bp. The developed FINS was validated with 20 market snapper samples that showed 100% specificity. This study helped to generate nucleotide sequences of D-loop region for 6 species of snapper for the first time for deposition in GenBank. Hence, to authenticate the snapper species, the developed PCR-SSCP and FINS were more specific and reliable.

A PCR-RFLP protocol was developed for authenticating nine different snapper species by targeting the 515 bp fragment in highly polymorphic mitochondrial D-loop region with a newly designed primer Fish-DL-F/Fish-DL-R, followed by restriction digestion using a single enzyme, Tsp5091. Individual species could be differentiated by 3-5 major bands. Very closely related species like L. fulvus and L. fulviflamma gave similar pattern due to high (94%) identity, while the other seven species were clearly differentiated. The protocol was also found successful in distinguishing the species in frozen, cooked and fried snappers. This protocol was validated with snapper samples procured from the fish market of Thoothukudi, India. Hence, the method can be efficiently used to authenticate snapper species within 8 h by regulatory authorities to avoid seafood species substitution. (C) 2018 Elsevier Ltd. All rights reserved.

The present study is the first concerning gillnet selectivity in Thoothukudi waters, Tamil Nadu, India. Selectivity estimates were made for Sphyraena forsteri. Sampling took places at three landing centers in Thoothukudi Coast from the period between April 2012 to March 2013. Samples were collected with gillnets of mesh sizes of 28 mm, 50 mm and 52 mm. Selection factors for S. forsteri is 7.04 whereas the commercial significant length group is 40.42 cm. Optimum size for this species is 5.82 cm.

Length frequency of barracudas, Sphyraena jello (Sphyraenidae) caught in gillnet with mesh size of 28, 50 and 52 mm operated in the Thoothukudi coast, Tamilnadu was recorded using this the selectivity parameters of the nets of these mesh sizes were assured. The size range of S. jello caught was 13-82 cm. The estimated selection factor ranged as 8.27 and the selection length ranged from 48 to 50 cm. Gillnets with mesh size less than 28 mm did not facilitate the escapement of S. jello which had not attained the stage of first maturity.

Listeria monocytogenes was screened from different seafood contact surfaces in five sampling sites of fishing harbour, fish landing centers, seafood processing plants, fish market, and fish curing yards of Tuticorin Coast of India. 115 swab samples were collected and tested for the occurrence of L. monocytogenes by conventional and molecular methods. Overall, 5.22% of swab samples collected were positive for L. monocytogenes. The fishing harbour had high incidence (10.3%) of L. monocytogenes followed by fish landing centers (5.9%), and seafood processing plants (4.1%). Boat deck, fish transport tricycle were the two seafood contact surfaces in fishing harbour, which had the occurrence of L. monocytogenes. The swab samples from fish market and fish curing yards were negative for L. monocytogenes. All the isolated colonies of L. monocytogenes were confirmed by PCR assay targeting virulent hlyA gene. The DNA of all the isolates yielded a product of 174 bp on PCR amplification in comparison with L. monocytogenes Type culture (MTCC 1143). The results clearly indicated the occurrence of L. monocytogenes in seafood contact surfaces along the Tuticorin Coast of India.

Seafood serves as a valuable protein source for human population. Among the seafood, tuna is considered as one of the commercially important species worldwide. Five tuna species namely Euthynnus affinis, Auxis thazard, Katsuwonus pelamis, Thunnus albacares and Thunnus obesus are commonly landed in the Tuticorin coast of Tamil Nadu. Among the five species, the exportable quality and grade of the meat is as follows: T. albacares> T. obesus> K. pelamis> E. affinis> A. thazard. In recent years, mislabeling is done by replacing high value tuna meat with low value tuna meat or with other low value fish meat to earn illegally. Usually, the species identification is done based on the morphological features, but this cannot be applied for the processed fish. PCR based identification methods have gained importance in the identification of fish species. PCR-AFLP is one of the molecular based methods, which can differentiate even closely related species. In this study, SE-AFLP method was employed to differentiate the above five tuna species viz., E. affinis, A. thazard, K. pelamis, T. albacares and T. obesus. Species specific AFLP marker was obtained in the primer combination of EcoR1 for fresh tuna. Band Sharing Index (BSI) analysis was also performed to find the similarities and variation among the five tuna species. AFLP profile of unknown tuna products was compared with the standard AFLP profile and the tuna species authentication was done by analysing BSI score.

Proximate profiles such as protein, carbohydrate, lipid and moisture content is often necessary to ensure that they meet the nutritional requirements essential for the growth and health of a living being. Hence, in the present study, the proximate compositions of three cuttlefish species from Thoothukudi coast were evaluated. Sepia prabahari had highest moisture level with a value 85.08±0.32 and S. ramani showed the lowest value (81.41±0.21). Protein content was highest in S. ramani with a value 16.474±0.46 and S. prabahari showed the lowest value (12.59±0.49). Ash was found to be higher in S. ramani (1.20±0.21) than the other two species. Carbohydrates were high in S. pharaonis (1.35±0.42) and low in S. ramani (0.71±0.10). Fat content ranged from 0.58±0.05 to 0.751±0.04 in the cuttlefish species. Thus, the present study showed that among the three cuttlefish species, Sepia ramani with highest protein content is nutritionally preferred.

Collagen, collagen-chitosan (C1), collagen-calcium acetate (C2) and collagen-chitosan-calcium acetate (C3) films were prepared from shark catfish (Pangasius pangasius) skin, and were investigated for their mechanical and functional properties. Collagen was characterized as type I and contained three identical chains. C3 film had higher tensile strength (10.4MPa) and Young's modulus (570MPa). Composite films had low swelling rates and solubility and lasted for 6 days before complete biodegradation. Fourier transform infrared spectra suggested that the interactions of collagen with chitosan and calcium salts affected the secondary structure and molecular order of collagen, particularly the triple helical structure. Scanning electron microscopy micrograph of collagen films depicted the formation of spindle-like structures with chitosan, granules with calcium acetate salts and porous structures in C3 films. Therefore, it was concluded that the collagen films had better mechanical, functional and in-vitro biodegradation properties by the addition of calcium and chitosan. These results demonstrate the potential application of composite collagen films from shark catfish skin in biomedical and pharmaceutical industries.Practical ApplicationsTo reuse fish-processing waste for biomedical materials, multicomposite collagen films were prepared from shark catfish skin and examined for their physicofunctional and mechanical properties. The purified collagen was combined with calcium and chitosan to improve the film properties. These results suggest that collagen films had better mechanical, in-vitro biodegradation and functional properties by the addition of calcium and chitosan. Accordingly, the prepared multicomposite fish collagen films could be suitable materials in biomedical and pharmaceutical industries as alternatives to mammalian collagen films.

Squid protein hydrolysates (SPH) were prepared from the Indian squid Loligo duvauceli using papain. Response surface methodology (RSM) was used for optimization of hydrolysis conditions, including temperature, time, and the enzyme-substrate ratio using DPPH radical scavenging activity as a response. The amino acid composition of SPH was compared with raw squid muscle. In vitro antioxidant activities were evaluated based on reducing power, metal chelation, ABTS, hydroxyl radical, and superoxide anion radical scavenging assays. SPH exhibited good ABTS radical scavenging activities of 96.50±0.90%, superoxide anion radical scavenging activities of 96.4±0.89%, reducing powers of 0.71±0.02, moderate hydroxyl radical scavenging activities of 64.03±2.11%, and metal chelating activities of 52.04±1.02%. In vivo antioxidant activities determined using a sardine minced model system showed 42% reduction in formation of secondary oxidative products as thiobarbituric acid reactive substances (TBARS), almost equivalent to reduction by ascorbic acid of 41.42% at 400 ppm.

Squids perish rapidly like other fishes and require some treatment to maintain the quality for export. The microbial quality of frozen squid (Sepioteuthis lessoniana, Lesson 1830) treated with the commercial food grade chemicals imported from Spain was therefore studied. Dressed squids were treated with the chemicals Hidratech_4A (0.4%) and Whitech_3 (0.25%) dissolved in chilled freshwater (STCF) and chilled saltwater (STCS). Chilled squids not treated served as control (SNTC). Squids were quick frozen at -40°C in contact plate freezer and stored at-20±1°C. Samples were tested raw, after pre-processing, treatments and freezing and during storage at monthly intervals for 7 months. The microbial quality evaluation included Total Plate Count (TPC), Escherichia coli, Staphylococcus aureus, Vibrio cholerae and Salmonella. Study revealed a better quality of treated samples than control. Microbiological quality of STCS was better than STCF and SNTC. E. coli counts decreased after treatment. Salmonella and V. cholerae were absent.

A PCR-RFLP method was used to identify five species of raw and processed groupers belonging to the genus, Epinephelus viz. E. areolatus, E. bleekeri, E. faveatus, E. longispinis and E. undulosus. DNA extracted from all the species using phenol-chloroform method amplified the mt 16S rRNA gene, unique for the genus, Epinephelus, using GenF and EpiR primers at 300bp. This fragment did not amplify the DNA of even closely related grouper genus, Cephalopholis, making the method more specific the grouper genus, Epinephelus. The specificity was further ascertained by performing the internal control that targeted 18S rRNA, which was eukaryotic specific. Digestion of PCR products with three selected restriction enzymes viz. SduI, BciVI, and Sau3AI followed by PAGE yielded species specific patterns that enabled identification of three grouper species viz. E. bleekeri, E. faveatus and E. longispinis, while the other two species viz. E. areolatus and E. undulosus gave similar pattern in the gel, although minor variations in the molecular sizes of digested products were noticed. Chilled, frozen and cooked groupers also gave similar PCR-RFLP pattern to that of raw groupers making this method suitable even for processed products.