GREEN EXTRACTION AND CHARACTERIZATION OF GELATIN COMING FROM UNSORTED DEHYDRATED CANNED TUNA SIDE STREAMS

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Introduction This study, funded by the EcoeFISHent project [1], describes a scalable method for the extraction of valuable proteins, such as marine gelatin and Hydrolyzed Gelatin Peptides (HGPs), but also non-collagenous proteins, in order to define future applicationsin the food, packaging, and cosmetic industries. The process starts from unsorted mixed Yellowfin tuna scraps after a patented industrial dehydration process that enhances the logistics in the valorisation of highly perishable biomasses. Materials and Methods Samples made up of raw crude heads, fins, skin, tails, and bones from Generale Conserve's (ASdoMAR®) mixed unsorted canned Yellowfin tuna processing side streams were dehydrated and milled using an industrial process patented by Themis S.p.A. [2] to stabilize them over time as a fish biomass with a low residual humidity. Besides, microbiological activity of samples pre and post dehydration was monitored to ensure the occurred stabilization. Before gelatin extraction, the starting material was analyzed in terms of proximate analysis to assess residual moisture (AOAC 950.46B), protein fraction (AOAC 981.10), ashes (942.05) and applying Hara-Radin method for the evaluation of lipid content [3]; moreover, amino acid composition was determined by HPLC [4,5]. The gelatin extraction can be divided into three main steps: pre-treatment, extraction, and drying. In addition to the extraction of gelatin, this extraction method also allows the recovery of non-collagenous proteins and HGPs. The following analyses were proposed for the characterization of the extracted gelatin and non-collagenous proteins: their composition in residual moisture, protein, ashes, and lipid contents (AOAC Official Method 935.46), and amino acidic composition [4,5]. The FT-IR analysis was applied to study the secondary structure of extracted gelatin and, as for its rheological parameters, a solution of 6.67% was prepared and the viscosity was measured at 40°C, while the gelling point was detected in a range of 5-40°C [6]. Results and Discussion The proximate analysis of the starting material reported low values of residual moisture (under 5%) which means that the industrial dehydration treatment was very effective. As expected for large-sized fish, the crude protein fraction analysis revealed a high percentage of total nitrogen, over 50%; ashes were found to constitute about 30% of the total composition, reflecting a high presence of bones in the biomass, which are considered a good gelatin source. Lipid fraction presented a 13% value and parallel research on its quality and omega-3 composition are currently ongoing. The study of the amino acidic profile revealed that Glycine (13.80%), Glutamic Acid (11.92%), and Alanine (7.33%) make up the majority of the components, Cysteine and Taurine (around 1% each) are the minor ones, while the collagen/gelatin-characteristic amino acids Hydroxyproline and Proline account for 10.57% together. The extraction process allows to recover both non-collagenous proteins and also collagenous ones (i.e., gelatin and HGPs) with yields equal to 14.6, 1.7% and 3.2%, respectively. Autumn School in Food Chemistry – Pavia September 2023 The extracted gelatin was analysed as follows: in terms of stability and shelf life, the low residual moisture value of 2% is considered a good result; crude protein fraction of 88% shows that gelatin has been well purified; implying that the defatting phase was effective (lipids below 1%), although demineralization still needs to be improved, since ashes were found to be 5%. The composition of amino acids was found to be similar to the one reported by Nurilmala et al. [7] for gelatin extracted from skin tuna, showing a high concentration of Glycine (24.57%) and about 9% for Arginine, Glutamic Acid, Proline, Hydroxyproline and Alanine, with ratio OH-Pro/Pro equal to 0.98. The FT-IR spectrum shows the characteristics peaks of collagen/gelatin: Amide A, B, I, II, and III. The viscosity and the gelling point were measured obtaining 5.0 mPa s and 14.6 °C respectively; these specific and characteristics results are lower than those reported in literature [8] for gelatin extracted from previously separated fish side streams, nevertheless further investigations using different concentrations are currently ongoing and several studies including different fields of application, such as foods, packaging and cosmetics, are in fieri above all considering the different range of the obtained gelatin and HGPs in terms of molecular weight . As for non-collagenous proteins, the analysis of the composition revealed that they are constituted by residual moisture (5.3%), proteins (67.0%), and ashes (24.0%); the amino acidic profile shows 14.59% for Glycine and 12.18% for Glutamic Acid, followed by about 8% for Aspartic Acid and Alanine, 7.25% for Arginine, while Proline (6.82%) and Hydroxyproline (4.32%) were considerably lower than what was found in gelatin. Conclusions The disposal of the side streams caused by the enormous growth in fishing production over the past few years is becoming an urgent environmental and economic issue to solve. In the literature, the extraction of gelatin starting from separated leftovers (skin and bones) has already been widely discussed. Differently, in this work, a scalable process to extract valuable proteins from unsorted mixed tuna scraps coming from canned tuna industry is presented, demonstrating that it is possible to benefit from the entire side stream, avoiding the onerous step of separation which could represent a significant advantage for industry. References 1) Https://Ecoefishent.Eu/. 2)Https://Patents.Google.Com/Patent/WO2015181769A1/Ja.Https://Patents.Google.Com/Patent/WO2015 181769A1/Ja 3) Hara, A.; Radin, N. S. Analytical Biochemistry. 1978, 420–426. 4) Lorenzo, J. M.; Purriños, L.; Temperán, S.; Bermúdez, R.; Tallón, S.; Franco, D. Poultry Science. 2011, 931- 940. 5) Domínguez, R.; Borrajo, P.; Lorenzo, J. M. Journal of Food Composition and Analysis. 2015, 61-67. 6) Gómez-Guillén, M. C.; Turnay, J.; Fernández-Dı ́az, M. D.; Ulmo, N.; Lizarbe, M. A.; Montero, P. Food Hydrocolloids. 2002, 25–34. 7) Nurilmala, M.; Hizbullah, H. H.; Karnia, E.; Kusumaningtyas, E.; Ochiai, Y. Marine Drugs. 2020, 98. 8) Mafazah, E. M.; Pranoto, Y.; Rohman, A. "Extracting of Yellowfin Tuna (Thunnus Albacares) Fish Skin Gelatin as Influenced by Alkaline Concentration and Extraction Times", IOP Conference Series Earth and Environmental Science, Yogyakarta, Indonesia 2018, p. 139.

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