쌀을 이용한 요거트 제조 기술개발 = Development of manufacturing process for rice-based yogurt

다국어 초록 (Multilingual Abstract)

Rice is a major food source that has been cultivated in Southeast Asia including Korea and forms the basis of dietary life, but rice consumption is continuously decreasing due to westernized dietary life. In order to solve this problem, the development of processed rice foods is being actively progressed to increase the processing rate of rice and to respond to the trend of convenience and westernized dietary changes. Rice processed foods are limited to rice flour products, simple processed products, and beverages using rice. On the other hand, lactic acid bacteria fermented foods are mostly milkbased dairy products and have problems due to lactose intolerance or allergies.
This study attempted to develop a lactic acid bacteria fermented beverage using rice in accordance with the global trend of vegan food using rice with the least allergy among grains. Therefore, for the fermentation of lactic acid bacteria, the selection and identification of bacteria suitable for fermentation of lactic acid bacteria, optimization of the medium, and establishment of the conditions for fermentation of lactic acid bacteria were established. In order to apply the lactic acid bacteria fermentation conditions to various rice varieties, the pretreatment process of rice was optimized and the lactic acid bacteria fermentation process was established to present basic data for industrialization of processed rice fermented foods.
Rice, the main raw material used for fermentation of lactic acid bacteria, was produced and sold by varieties (Non-waxy rice, waxy rice, black rice, and red rice) provided by Soybean Food. Among non-waxy rice, the content of resistant starch in Saegoami (25.7%) and Dodam (35.1%), which have a high amylose content, was found to be 0.54% and 8.67%, respectively, but after gelatinization, the content of resistant starch decreased sharply to less than 1%. It was confirmed that the function of resistance starch disappeared. Rice pretreatment was performed using liquefied enzymes (Termamyl, Fungamyl) and glycosylated enzymes (AMG) to establish optimal conditions. The a-amylase activity of the liquefaction enzymes Termamyl and Fungamyl was found to be 340.0 and 356.4 KU/mg, respectively, and the glucosidase activity of AMG was 805.9 U/mg. Of the two liquefaction enzymes, Fungamyl was selected as the liquefaction enzyme because Fungamyl had a higher reaction rate than Termamyl under the same conditions. In the case of Fungamyl treatment, the amount of reducing sugar produced was optimized at 0.025% (v/v) or higher, whereas in the case of AMG, the amount of reducing sugar produced tended to increase as the treatment amount increased. Since 96% saccharification of rice starch was achieved with 28.9 g/L, the optimum treatment amount of each enzyme was 0.025% Fungamyl and 0.25% AMG. As a result of the individual and mixed treatment of 0.025% Fungamyl and 0.25% AMG, the same effect as the mixed treatment was confirmed even with a single treatment of AMG, and the enzyme treatment for the pretreatment of rice was decided as AMG treatment. The properties of starch change of reactants by liquefaction and saccharification were confirmed using sugar analysis and molecular weight analysis methods.
The contents of flavonoids and polyphenols, which are functional ingredients of rice by variety, were mainly found in black rice and red rice.
Changes in physiologically active substances according to the pretreatment of rice were analyzed in small amounts of flavonoids, polyphenol compounds, and antioxidants in non-waxy rice and waxy rice, but antioxidants were 3,255 and 141 mg% in mixed samples, respectively, in black rice and red rice, which are colored rice. Black rice showed the highest value of 3,473, 153 mg%, and saccharification at 3,418 and 157 mg%, respectively. This is the same pattern as polyphenol compounds, and when gelatinization to saccharification proceeds, black rice decreases and red rice increases.
Important lactic acid bacteria in rice yogurt fermentation are yogurt starters commercially available for spawning: Hanmiyogurtting Powder (HANMI), Lyofast SAB 440 B (SACCO), HANSEN ABT-4, HANSEN YC-381 4 types and 2 isolates (Isolate 1, 2) was used, and as a result of culturing for 36 hours using pretreated rice, the number of lactic acid bacteria in Isolate 1 directly separated from the yogurt starter (HANMI) was good at 3.10×10 8 and 3.95×10 8
CFU/mL, respectively. Was less than 10 6 CFU/mL. The selected lactic acid bacteria HANMI and Isolate 1 were purely separated from the fermented product and identified using API kit and 16S rRNA gene sequencing. As a result, HANMI was identified as L. plantarum and Isolate 1 as L. harbinensis. In the case of L. plantarum, a standard strain was distributed and used from KCTC. In order to optimize the basic medium for the two lactic acid bacteria, glucose, which is a saccharified product of rice, and yeast extract for food were selected as the basic composition. The optimum concentrations of carbon and nitrogen sources for the growth of two lactic acid bacteria species and production of metabolites (lactic acid and acetic acid) were identified as 2.5% (w/v) glucose and 1% (v/v) yeast extract, respectively, and the optimum temperature was 37 ◦ C, and the initial pH was determined to be 6.0. As a result of fermentation of lactic acid bacteria using individual lactic acid bacteria and mixed lactic acid bacteria under optimal medium composition and culture conditions, the number of viable bacteria and lactic acid/acetic acid maintained 10 8 CFU/mL, 8.8 g/L, 0.8 g/L or more, The mixed culture of L.
harbinensis and L. plantarum was excellent.
Lactobacillus fermentation was performed using 3% (w/v) rice and 1% yeast extract to meet the conditions similar to the optimal carbon source (2.5% glucose) for fermentation of lactic acid bacteria. Lactobacillus fermentation using glucose and lactobacillus fermentation pretreated with non-waxy rice were similar in growth and production of metabolites. In the case of lactic acid bacteria fermentation using black rice and red rice containing functional ingredients, when using black rice, metabolites were produced up to 1.9 times more than non-waxy rice and the number of viable bacteria was maintained. In particular, in the case of red rice, about 20 hours of lag time existed after the inoculation of lactic acid bacteria, and overall fermentation results between non-waxy rice and black rice were obtained. In the case of fermentation of lactic acid bacteria using rice, the best results were obtained when black rice was used.

국문 초록 (Abstract)

유산균 발효에 사용되는 주원료인 쌀은 품종별(멥쌀, 찹쌀, 흑미, 홍국미)로 생산·판매되는 쌀가루를 대두식품㈜에서 제공받아 사용하였다. 멥쌀 중에서 아밀로스 함량이 높은 새고아미(25.7%)와 도담쌀(35.1%)의 저항전분 함량은 각각 0.54%와 8.67%로 확인되었으나 호화 후에는 1% 미만으로 저항전분 함량이 급격하게 낮아지는 사실이 확인되어 저항전분의 기능이 사라지게 되었다. 쌀 전처리는 액화효소(Termamyl, Fungamyl)와 당화효소(AMG)를 사용하여 최적화 조건을 확립하였다. 액화효소인 Termamyl과 Fungamyl의 -amylase 효소활성은 각각 340.0과 356.4 KU/mg으로 확인되었고, AMG의 glucosidase 활성은 805.9 U/mg로 확인되었다. 2가지 액화효소 중 Fungamyl이 동일한 조건에서 Termamyl보다 반응속도가 높아 액화효소로 Fungamyl을 선정하였다. Fungamyl 처리량은 0.025% (v/v) 이상에서 환원당 생성량이 최적화된 반면, AMG의 경우에는 처리량이 증가함에 따라 환원당 생성량이 증가하는 경향을 보이고 0.25% (v/v) 처리시 환원당 생성량이 28.9 g/L로 쌀 전분의 당화가 96% 이루어지므로 각 효소의 최적 처리량은 Fungamyl 0.025% 및 AMG 0.25%로 적용하였다. 0.025% Fungamyl과 0.25% AMG를 개별 및 혼합처리한 결과, AMG의 단일처리 만으로도 혼합처리와 동일한 효과를 확인하여 쌀의 전처리를 위한 효소처리는 AMG 단일처리로 결정하였다. 액화 및 당화효소에 의한 반응물의 전분 변화는 당 분석과 분자량 분석방법을 이용하여 확인하였다.
품종별 쌀이 지니고 있는 기능성 성분인 플라보노이드와 폴리페놀의 함량은 주로 흑미와 홍국미에서 확인되었다. 쌀 전처리에 따른 생리활성물질 변화는 멥쌀과 찹쌀에서 플라보노이드, 폴리페놀화합물, 항산화물 모두 적은 양이 분석되었으나, 유색미인 흑미와 홍국미에서는 항산화물이 혼합시료에서 각각 3,255, 141 mg%, 호화에서는 각각 3,473, 153 mg%, 당화에서 3,418, 157 mg%로 흑미가 가장 높은 값을 나타내었고, 이는 폴리페놀화합물과 동일한 패턴으로 호화에서 당화로 진행될 때 흑미는 감소하고 홍국미는 증가하는 경향을 보였다.
쌀 요거트 발효에서 중요한 유산균은 종균용으로 시판되는 요구르트 스타터인 한미요구르팅 파우더(HANMI), Lyofast SAB 440 B (SACCO), HANSEN ABT-4, HANSEN YC-381 4종과 분리균주 2종 (Isolate 1, 2)을 이용하였으며, 전처리를 거친 쌀에 접종하여 36시간 배양한 결과, 요구르트 스타터(HANMI)와 직접 분리한 Isolate 1에서 유산균수가 각각 3.10×108, 3.95×108 CFU/mL로 좋았으며 나머지의 경우에서는 106 CFU/mL 미만이었다. 선정된 유산균인 HANMI와 Isolate 1를 발효물에서 주요 유산균을 순수 분리하여 API kit와 16S rRNA gene sequencing을 이용하여 동정한 결과, HANMI는 L. plantarum, Isolate 1은 L. harbinensis로 동정되었다. L. plantarum의 경우 KCTC에서 표준균주를 분양받아 사용하였다. 2가지 유산균에 대하여 기본적인 배지를 최적화하기 위하여 쌀의 당화산물인 포도당과 식품용 이스트엑기스를 기본조성으로 선택하였다. 2가지 유산균종의 생육과 대사산물(젖산 및 초산) 생산에 미치는 탄소원과 질소원의 최적 농도는 각각 2.5% (w/v) 포도당과 1% (v/v) 이스트엑기스로 확인되었으며 최적 온도는 37◦C, 그리고 초기 pH는 6.0으로 결정되었다. 최적 배지조성과 배양조건에서 개별 유산균과 혼합 유산균을 이용하여 발효를 진행한 결과, 생균수와 젖산/초산이 108 CFU/mL 및 8.8 g/L, 0.8 g/L 이상을 유지하는데 L. harbinensis와 L. plantarum의 혼합 배양이 우수하였다.
유산균 발효를 위한 최적 탄소원(2.5% glucose)과 유사한 조건을 맞추기 위하여 3% (w/v) 쌀과 1% yeast extract를 이용하여 유산균 발효를 진행하였다. 포도당을 이용한 유산균 발효와 멥쌀을 전처리한 유산균 발효는 생육과 대사산물의 생산이 유사하게 확인되었다. 기능성 성분을 함유한 흑미와 홍국미를 이용한 유산균 발효의 경우에는 흑미를 이용한 경우 대사산물의 생성이 멥쌀에 비하여 최대 1.9배 더 많이 생성되었으며 생균수도 유지되었다. 특이하게 홍국미의 경우에는 유산균 접종 이후에 약 20여시간의 Lag time이 존재하였으며 전반적으로 멥쌀과 흑미의 중간정도의 발효결과를 얻을 수 있었다. 쌀을 이용한 유산균 발효는 흑미를 이용하는 경우가 가장 좋은 결과를 얻을 수 있었다.

목차 (Table of Contents)

• Ⅰ. 서론 ··············································································································· 1
• Ⅱ. 재료 및 방법 ··································································································· 5
• Ⅲ. 결론 ··············································································································· 16