Production of Mihaliç cheese with increased vitamin B2 levels using propionic acid bacteria.

Abstract

The vitamin B12 production capacity of propionic acid bacteria (PABs) was evaluated; parameters affecting vitamin B12 production were selected and optimized, and the applicability of a microbial biofortification approach to a traditional product was investigated by applying it to Mihaliç cheese. In the study, 21 PAB isolates originating from Mihaliç cheese and a DSM strain originating from Swiss cheese, known to produce B12, were activated and their vitamin B12 production capacities were determined. It was determined that the isolates coded K32 and T11, Propionibacterium freudenreichii subsp. shermanii and Propionibacterium freudenreichii subsp. freudenreichii, produced the maximum amount of vitamin B12 at point level, with 1.71–0.04 mg / L and 1.57–0.14 mg / L of vitamin B12, respectively.Subsequently, factors affecting production such as pH, temperature, riboflavin, lactate, casein, salt, time, inoculum ratio, inoculum type, and aeration were screened using the Plackett-Burman design; as a result of the analyses, riboflavin and incubation time were transferred to the optimization stage as the most effective parameters. These two variables were transferred to the optimization stage using Response Surface Methodology with Central Composite Design (CCD), and the resulting model was found to be statistically significant (p<0.001). Following optimization, the optimum points for the parameters, determined as riboflavin at 57.58 mg / L and incubation time at 10.67 days, were applied to Mihaliç cheese production. In addition to the control group (K18), 1% of Propionibacterium freudenreichii subsp. shermanii isolate coded K32 was added to the experimental group (R18), and Mihaliç cheese was matured for 90 days.After 90 days, the final product showed vitamin B12 content of 0.52–0.01 µg / 100 g in the control group cheese and 1.82–0.03 µg / 100 g in the experimental group cheese. A 3.5-fold increase in vitamin B12 levels was observed in the experimental group compared to the control sample (p < 0.05). Physicochemical analyses of pH, titratable acidity, salt, dry matter, ash, protein, and fat were performed on the cheeses, along with total mesophilic bacteria, lactic acid bacteria, yeast-mold, and PAB counts; textural properties and sensory evaluation were also conducted. According to the results, optimizing production parameters to increase vitamin B12 content did not negatively affect technological quality; and the fermentation and ripening process maintained microbial and structural stability.

Description

Production of Mihaliç cheese with increased vitamin B2 levels using propionic acid bacteria. Technical Area This invention relates to the fields of food technology and food biotechnology, particularly milk and dairy products. The invention relates to cheese production methods within the scope of product technology. The invention concerns propionic acid. Increasing vitamin B12 biosynthesis through the use of bacteria and this approach in Mihaliç It is based on integrating cheese into the production process. State of the Art Vitamin B12 is an essential micronutrient for human health, and it is produced in red blood cells in the body. vital processes such as cell formation, DNA synthesis and nervous system functions It plays a role. Because the human body cannot synthesize it, vitamin B12 is part of the diet. It must be obtained through this method. Animal-derived sources are used to meet daily requirements. Foods constitute the basic sources of nutrition. The chemical synthesis of vitamin B12 is a multi-step process, technically complex and economically challenging. It is costly. Therefore, industrial-scale production is mainly carried out using microorganisms. This is carried out through propionic acid bacteria. Specifically, Propionibacterium freudenreichii species are biologically active sources of vitamin B12. They are widely used because of their ability to synthesize various forms. Milk and dairy products are an important source of vitamin B12 due to its high bioavailability. While being a source of nutrition, cheeses produced using traditional methods Vitamin B12 content is limited. A large portion of the starter cultures used in cheese production... It is mostly composed of lactic acid bacteria, and these microorganisms produce vitamin B12. Their synthesis capacity is limited. Therefore, vitamin B12 synthesis is affected in traditional cheese production. The amount generally remains at low levels. Propionic acid bacteria are particularly effective in the ripening of Swiss-type cheeses, improving aroma and porosity. It plays an important role in terms of formation and structural development. However, these bacteria... Controlled and targeted enhancement of vitamin B12 production potentials using cheese matrix. Studies aimed at increasing its intake are limited. In current practices, vitamin B12... Increasing the content is mostly done by adding vitamins to the final product (fortification). This approach is being implemented. This approach leads to the alteration of the product's natural structure and negatively impacts consumer perception. the perception of "additives," the stability of the vitamin B12 form, and cost issues. This gives rise to debate. 1 35 In the literature, propionic acid bacteria are used to produce vitamins in liquid culture media or model systems. Although there are studies on B12 production, optimizing this production is not possible. integrating it into a traditional cheese production process and technologically and microbiologically improving the cheese and a technical solution for implementation while preserving sensory characteristics It is not found. In addition, vitamin B12 is found in local and traditional products such as Mihaliç cheese. A defined production method for increasing the level of 40 through microbial biosynthesis. It is not available. Therefore, it is a natural fermentation process that is compatible with the traditional Mihaliç cheese production process. a new and improvable approach to increasing vitamin B12 levels, based on these principles A technical solution is needed. 45 Purpose of the Invention The aim of this discovery is to investigate the biosynthesis potential of propionic acid bacteria for vitamin B12. By utilizing a method compatible with the traditional Mihaliç cheese production process, based on natural fermentation, and 50 is to develop a method that maintains technological quality. In this context, the invention relates to the external application of vitamin B12 to traditional Mihaliç cheese, which has a limited vitamin B12 content. In situ biological enrichment via microorganisms without vitamin supplementation. It aims to provide this. Another objective of the invention is to control the propionic acid bacteria. While its consumption helps increase vitamin B12 synthesis, the cheese also has its own unique structure and taste. 55. The goal is to preserve the aroma and microbiological stability. Furthermore, the invention involves determining the fermentation parameters that affect vitamin B12 production and By integrating these parameters into the cheese production process, repeatable and industrial-grade results can be achieved. It aims to offer a production method that is feasible on a large scale. Accordingly, the invention aims to increase the functional value of a traditional dairy product, for the consumer. It can be achieved through an enrichment approach preferred by 60 and adds value. The aim is to obtain a high-quality, new Mihaliç cheese product. Description of the invention a) Microorganism Selection 2 65 Within the scope of the invention, high B12 content among propionic acid bacteria originating from Mihaliç cheese. Strains with the capacity to produce vitamins are selected. Those found to have maximum production are chosen. Propionibacterium freudenreichii subsp. shermanii (e.g., isolate coded K32) was used. b) B12 Optimization of Production Parameters Parameters affecting vitamin B12 production by propionic acid bacteria were screened and optimized. 70 was obtained. As a result of optimization: Riboflavin concentration: 57.58 mg / L and incubation. The duration has been set at 10.67 days. c) Mihaliç Cheese Production Mihaliç cheese is produced as follows: • The milk is thermized and cooled to fermentation temperature. 75 • In experimental production, milk was treated differently from the control group: o 1% selected propionic acid bacteria, The optimum amount of riboflavin (57.58 mg / L) is added. • Coagulation, blanching, swollen, pressing, and brining (initially 14%, then 18%) The processes are carried out in accordance with traditional Mihaliç cheese production methods. 80 • The cheeses are aged in 18% salt brine. • Maturation period starts 11 days after the final brine treatment. It begins and is carried out for at least 60–90 days. d) Final Product Specifications Mihaliç cheese produced using this method has the following properties: 85 • The amount of vitamin B12 increases 3.5 times compared to traditional production, • Quality parameters such as pH, salt, dry matter, protein and fat are maintained. • Sensory characteristics: Mihaliç cheese retains its characteristic texture, taste, and aroma. • The study investigated in situ vitamin B12 content of Mihaliç cheese under optimized conditions. It is a suitable product for biofortification and does not negatively affect technological quality. It shows that a functional cheese can be produced without affecting 90% of vitamin B12. The values ​​are given in Table 1. 3 • Table 1 Vitamin B12 Determination Values ​​in Cheese (µg / 100 g cheese) Vitamin B12 (µg / 100g cheese) Example code for Day 1, Day 30, Day 60, Day 90. Ca Da Ba Aa K18 0.990.01 1.110.01 0.860.04 0.520.01 BCb Cb Bb Ab R18 2.14 ± 0.06 2.20 ± 0.02 2.04 ± 0.07 1.82 ± 0.03 • A,D (→) show a difference in the same row during the maturation period (p<0.05). a, b, (↓) indicate the difference between examples in the same column. 95 • (K18: Control sample matured in 18% salt brine; R18: in 18% salt brine) (sample with added riboflavin and culture that has been matured) How the invention can be applied to industry. The invention can be implemented within the existing Mihaliç cheese production infrastructure without requiring additional equipment. With a value of 100, it enables the production of a value-added dairy product with enhanced functional value. 4

Claims

1. Mihaliç cheese with increased vitamin B12 levels using propionic acid bacteria. production method.

2. According to Claim 1, the bacterium used is Propionibacterium freudenreichii subsp. It is Shermanii (K32).

3. According to Claim 1, the riboflavin supplementation should be 57.58 mg / L.

4. According to claim 1, the inoculum ratio should be 1%.

5. According to Claim 1, the cheese must be aged for at least 90 days, starting from the 11th day. A production method characterized by its maturation process.

6. Mihaliç cheese with increased vitamin B12 content, obtained according to Claim 1.