بهینه‌سازی تولید دوغ حاوی آنزیم ترانس گلوتامیناز با استفاده از کازئینات سدیم

Introduction: “Doogh” is a native beverage in Iran and has in important share in beverage industry (Zomorodi and Kianfar 2018). It can be made by yoghurt dilution or direct acidification of milk. Traditionally, doogh is made by mixing yogurt, water, and a little salt as well as some aqueous extracts of local herbs. However, Doogh, like other acidic dairy beverages has a serious problem because of its low pH, which causes phase separation, and leads to casein accumulation and gives a product with an undesirable, non-uniform appearance (Sanli et al., 2011). On the other hand, heat treatments such as pasteurization have a key importance in the production of dairy products because this is a common step in the processing of milk products. The heat treatment exposes reactive groups on the protein, which were previously inaccessible, and this, in turn, affects the rheological properties of the products (Joudaki et al., 2013). It has been reported that polysaccharide hydrocolloids addition could be prevent phase separation in doogh during storage. But creation undesirable taste in the higher concentration is a seriouse problem (Tsevdou et al., 2013). The enhancement of food texture with enzymes has gained increasing importance as an alternative to conventional processing strategies for protein containing foods. Microbial transglutaminase or glutaminyl-peptide-amine γ-glutamyl transferase (mTGase, EC 2.3.2.13) is a calcium independent enzyme. It forms covalent crosslinks of inter- or intra-molecular ε-(γ-glutamine)-lysine isopeptidic bonds by catalyzing an acyl transfer reaction between a γ-carboxyamide group in protein-bound glutamine residues (acyl donor) and an ε-amino group in a protein-bound lysine residue (acyl acceptor) (Lorenzen et al., 2002). Microbial transglutaminase represents an interesting tool for texture modification of protein-containing foods. Since 1998, this enzyme with “No. GRN 000095” was approved by the FDA as “Generally Recognized as Safe (GRAS)” and there are confirmed documents for MTGase in the US, Japan and Europe as a safe ingredient in different types of processed foods (Ardelean et al., 2012). Acid gels, such as yogurt, made from enzyme-treated milk are characterized by reduced syneresis and improved viscosity. In this study, effect of sodium caseinate on functionality of transglutaminase was assayed too.   Concerning milk proteins, casein monomers can be easily cross-linked by mTGase, whereas whey proteins only polymerize after heat treatment or in the presence of reducing agents. The literature on mTGase-induced cross-linking on casein micelles, caseins or caseinates, whey proteins or milk, has been reviewed recently (Barbaros et al., 2012). The aim of this study was to investigate the effect of transglutaminase enzyme and sodium caseinate to improve quality and stability characteristics of Doogh.
Material and methods: This study was carried out to improve the quality and stability properties of Doogh using sodium caseinate (at 0, 0.5 and 1%) and microbial transglutaminase (at 0, 5, 10 u/g protein). For this purpose, after pasteurization and homogenization of raw milk (0.6 % fat), sodium caseinate and microbial transglutaminase added at 45 and 50^oC, respectively. Heating of treated milk was performed after 2 h (72^oC- 15 s) to inactive enzyme. Fermentation was carried out by adding starter culture (yogurt starter culture consist of Lactobacillus bulgaricus and Streptococcus thermophiles) and incubation was performed at 42^oC until 12 h. Finally, prepared yogurt mixed with potable water at 1:1 ratio and 0.65% salt added to mixture. The pasteurized treated samples were assayed for physicochemical and sensory properties. Acidity was measured through titration with sodium hydroxide as Dornic degree and viscosity was assayed at ambient temperature by a rotational Brokfield viscometer. Phase separation of the different formulations was carried out in 250 mL glass tubes for 30 days at 5 ^oC storage. Evaluation of the sensory attributes of the produced Dooghs with various formulations in terms of taste, consistency, appearance, smell and color was carried out by 30 untrained panelists and the average of scores as overall acceptability were used for statistical analyses. In the second phase, based on the obtained results, optimized sample was selected using response surface methodology in central composite design and particle size distribution (determined by dynamic light scattering) and zeta potential tests were measured.
Results and discussion: The results showed that the use of transglutaminase increased significantly effect on acidity, while the effect of sodium caseinate was not significant. Transglutaminase and sodium caseinate had a significantly increase effect on sensory properties and viscosity, while phase separation decreased significantly. Optimization of formula was performed based on tranglutaminase, sodium caseinate and acidity in range, phase separation in minimize, apparent viscosity and overall acceptability in maximize. The importance of all of factors was selected the same. And optimum content of transglutaminase and sodium caseinate to have high stability product were calculated 4.30U/g Protein and 0.71% respectively through optimization with desirability 0.928. In optimum situation phase separation and apparent viscosity were obtained 2.15% and 29.10 cP, respectively. Optimum overall acceptability was acquired 4.25. Comparison of optimum and control samples showed that the optimum sample had higher viscosity (6.01 cP in control and 29.10 cP in optimized sample) and overall acceptability (1.55 in control and 4.25 in optimized sample) with at least phase separation (2.95% in control and 2.15% in optimized sample). Applying transglutaminase and sodium caseinate also reduced particle size (10.05 nm in control and 3.55 nm in optimized sample) and zeta potential (11.4 mV in control and -21.2 mV nm in optimized sample).
Conclusion: Generally, according to the obtained results, it can be said that applying MTG and sodium caseinate, is possible to produce the stable Doogh with desired properties in terms of physicochemical and sensory properties and long storage duration.