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Meyve suyu atıklarından elde edilen pektinin fonksiyonel probiyotik içecek üretiminde prebiyotik olarak değerlendirilmesi

Evaluation of pectin obtained from fruit juice waste as a prebiotic in the production of functional probiotic beverage

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  1. Tez No: 1009253
  2. Yazar: DİDEM KARA
  3. Danışmanlar: DOÇ. DR. DERYA KAHVECİ KARINCAOĞLU
  4. Tez Türü: Yüksek Lisans
  5. Konular: Gıda Mühendisliği, Food Engineering
  6. Anahtar Kelimeler: Belirtilmemiş.
  7. Yıl: 2023
  8. Dil: Türkçe
  9. Üniversite: İstanbul Teknik Üniversitesi
  10. Enstitü: Lisansüstü Eğitim Enstitüsü
  11. Ana Bilim Dalı: Gıda Mühendisliği Ana Bilim Dalı
  12. Bilim Dalı: Gıda Mühendisliği Bilim Dalı
  13. Sayfa Sayısı: Belirtilmemiş.

Özet

Fermente probiyotik meyve suyu içecekleri, insan sağlığı üzerindeki yararlı etkileri ile vegan, laktoz intoleransı olan, düşük kolesterollü diyetlere ihtiyaç duyan ve fonksiyonel içecek arayışında olan tüketiciler tarafından tercih edilmektedir. Bu içeceklerde probiyotiklerin hayatta kalmasını arttırmak için ürün prebiyotiklerle zenginleştirilmektedir. Lactobacillus türlerinin fermantasyon sırasında ürettiği ve biyoaktiviteye sahip postbiyotik bileşenlerden biri olan gama-aminobütirik asit (GABA), merkezi sinir sistemindeki başlıca inhibe edici nörotransmiterlerden biri olarak işlev gören, protein olmayan bir amino asittir. Son yıllarda yapılmış çalışmalar GABA'nın özellikle ruh sağlığı üzerine birçok yararlı etkisi olduğunu göstermektedir. Günümüzde, bağışıklık güçlendirici ve ruh hali iyileştirici içeceklere artan talebe yönelik fonksiyonel içeceklerin üretimi önemli bir hedef haline gelmiştir. Tüketici araştırmaları da özellikle ruh hali iyiliğini amaçlayan fonksiyonel gıdalara yönelik talebin arttığını ve bu trendin kalıcı olacağını göstermektedir. Bu doğrultuda tasarlanmış olan projenin temel amacı, GABA üreticisi olduğu tespit edilen laktik asit bakterisi kullanılarak fermente edilen, hem GABA'nın hem de meyve-sebzelerin sağlık yararlarını birleştiren probiyotik bir sebze suyu elde etmektir. Üründe probiyotik canlılığını arttırmak amacıyla da endüstriyel gıda atıklarından elde edilecek pektin ve hidrolizatlarının kullanılması, böylece üretimde sürdürülebilirlik hedeflerine uyum sağlanması amaçlanmaktadır. Bu kapsamda, öncelikle meyve suyu atığı olan nar posasından asidik ekstraksiyon ile %8,91 verimle pektin elde edilmiştir. Elde edilen nar pektini, ticari pektinaz ile enzim miktarı ve hidroliz süresine bağlı olarak 4 farklı koşulda hidrolize edilmiştir. Elde edilen pektin ve pektin hidrolizatlarının esterleşme derecesi tayini, galakturonik asit tayini, FTIR ve şeker analizleri yapılmıştır. Nar pektini ve hidrolizatlarının esterleşme derecesi (DE) %34,36 ila %49,90 arasında değişmiş olup düşük metoksil pektinler elde edilmiştir. FTIR analizinde ekstrakte edilen maddelerin pektin olduğunu kanıtlayan pektin moleküllerinden beklendiği gibi spektrumda tipik bantlar ortaya çıkmıştır. Tüm pektin ve hidrolizatlarında arabinoz, galaktoz, glikoz, mannoz ve ramnoz varlığı analiz edilmiş, glikoz ve ramnoz varlığı tespit edilmiştir. Ardından, elde edilen pektin ve hidrolizatların besiyeri ortamında Lactobacillus brevis canlılığına ve GABA üretimine etkisi incelenmiştir. Besiyerinde probiyotik canlılık ve GABA üretimi sonuçlarına göre, optimum hidroliz koşulları 400 µl pektinaz/g ile 2 saat hidroliz olan P4H2 hidrolizatı seçilmiştir. Literatürde, havuç suyunun Lactobacillus brevis ile fermentasyonunda prebiyotik olarak pektin hidrolizat ilavesinin, GABA üretimine ve probiyotik canlılığa etkisinin incelendiği bir çalışma mevcut değildir. Bu nedenle, prebiyotik olarak P4H2 pektin hidrolizatını farklı konsantrasyonlarda (%1, %2 ve %4) ihtiva eden havuç suyu Lactobacillus brevis suşu ile fermente edilerek çalışmaya özgün bir nitelik sağlanmıştır. Fermente meyve suyunda probiyotik canlılık test edilmiş ve GABA üretim potansiyeli kromatografik tayin yöntemleri ile izlenmiştir. Prebiyotik olarak P4H2 pektin hidrolizatını ihtiva eden fermente havuç sularının (45,24±4,52 mg/ml) GABA miktarının pektin hidrolizatını içermeyenlere (32,61±8,12 mg/ml) göre daha yüksek olduğu tespit edilmiştir. Ayrıca elde edilen prebiyotikli fermente havuç sularına toplam fenolik madde analizi ve toplam antioksidan aktivitesi analizi yapılmıştır. P4H2 hidrozilatı %4 konsantrasyonu ile toplam fenolik madde (93,35±7,88 GE mg/g) ve antioksidan aktivitesinde (CUPRAC: 303,45±9,51 TE mg/g; DPPH: 119,9±23,3 TE mg/g) en yüksek etkiyi göstermiştir. P4H2 hidrozilatı oranı azaldıkça fenolik madde miktarı ve antioksidan aktivitesi de önemli ölçüde azalmıştır. Çalışma sonucunda, pektin hidrolizatı ilaveli sebze suyunda Lactobacillus brevis fermantasyonu ile postbiyotik bir bileşen olan GABA üretimi araştırılmış, yeni bir fonksiyonel içecek geliştirilmiştir.

Özet (Çeviri)

The effect of the gastrointestinal microbiota on the health of humans and animals is important. Probiotics, which are known as live microorganisms that have beneficial effects on the health of the host when taken in sufficient amounts, gain importance in this sense. Today, consumers have included probiotics in their diets with this awareness and even take them as microbial nutritional supplements. Traditionally, probiotics have been used in yogurt and other fermented milk products. However, recently alternative raw materials have been studied to determine whether they are suitable substrates for the production of non-dairy probiotics. Lactic acid bacteria, which are most preferred as probiotics, are a group of grampositive, cocci or bacillus bacteria that secrete lactic acid during the fermentation of carbohydrates, are non-sporeless, catalase-negative, non-aerobic but aerotolerant. Lactic acid bacteria isolated from traditional fermented foods have received great attention due to their potential bioactivity and can produce various bioactive compounds (postbiotics) during fermentation. Known as an indigestible food ingredient that stimulates the growth and activity of health-promoting colon bacteria, prebiotics are important for the survival of probiotics. Prebiotics are chemically stable structures or indigestible carbohydrates such as fibers and oligosaccharides that modulate the gut microbiota. Prebiotics are easy, inexpensive, and safe to include in the diet. Among the prebiotic carbohydrates, fructans, FOS and inulin have dominated the prebiotic market to date. Maximum utilization of fruit and vegetable wastes in the production of prebiotics are promising approaches that have a positive impact on the environment, the food industry, and consumer health. Pomegranate peel, which constitutes approximately 40-50% of the total fruit weight, has an excellent added value potential as it is a rich source of polyphenols and other bioactive compounds. In addition, pomegranate peel extracts have the potential to be used as prebiotics due to their ability to increase the proliferation of lactic acid bacteria. Pectin is a complex polysaccharide widely distributed in the cell walls and middle lamina of higher plants. One of the most promising applications of pectin is the production of pectic oligosaccharides (POS) with short monosaccharide chains, and these oligosaccharides are proposed as excellent candidates for next generation prebiotics. The galacturonic acid ratio and the degree of esterification of pectin are very important for the biochemical, physical and functional properties of pectin. In addition to pectin, the prebiotic effects of pectin hydrolysates obtained by enzymatic or chemical hydrolysis are also investigated. In general, classical solvent extraction is used to obtain pectin. In the production of pectin hydrolysates, it is aimed to break down pectin using enzymes known as pectinolytic enzymes or pectinase. Gamma-aminobutyric acid (GABA), one of the postbiotic components with bioactivity produced by Lactobacillus species during fermentation, is a non-protein amino acid that functions as one of the major inhibitory neurotransmitters in the central nervous system. Recent studies show that GABA has many beneficial effects on cardiovascular diseases, insomnia, depression, chronic alcoholism and diabetes, improving conditions such as stress and anxiety, and especially mental health. GABA stands out as a postbiotic in this period when the growth in immune-boosting and mood-enhancing foods accelerates with increasing consumer demand. There are two approaches to increase GABA efficiency; these are to develop higher yielding strains and to optimize the fermentation conditions. Fermentation with LAB with high GABA production ability is a good choice for GABA yield. Consumers consume food not only to satisfy hunger and provide essential nutrients, but also to prevent diet-related diseases and improve physical and mental well-being. For this reason, functional foods are developed in order to meet the food demands of consumers for their lifestyle and needs. Fermented probiotic fruit juice drinks are preferred by consumers who are vegan, lactose intolerant, need low cholesterol diets and are looking for functional beverages with their beneficial effects on human health. To increase the survival of probiotics in these beverages, the product is enriched with prebiotics. Today, the production of functional beverages has become an important target for the increasing demand for immune-boosting and mood-enhancing beverages. Consumer research also shows that the demand for functional foods, especially aimed at mood wellness, is increasing and this trend will be permanent. The main aim of the project, which was designed in this direction, is to produce a probiotic vegetable juice that combines the health benefits of both GABA and fruits and vegetables, fermented using lactic acid bacteria found to be a GABA producer. In order to increase the viability of probiotics in the product, it is aimed to use pectin and hydrolysates obtained from industrial food wastes, thus complying with the sustainability targets in production. In this context, first of all, pomegranate pulp, which is fruit juice waste, was obtained from a fruit juice business. The pomegranate pulp was dried and ground in a lyophilizer and its approximate composition (moisture content, oil, ash and protein content) was found. Ground pomegranate peel was extracted by acidic extraction method in 1% citric acid solution in a shaking water bath at 80ºC for 1 hour. The mixture was then cooled to room temperature and centrifuged at 4000 rpm for 25 minutes. Pectin was precipitated by adding ethanol to the liquid phase and keeping it at 4°C for 1 hour. The resulting pectin was frozen, then ground after drying in a lyophilizer. Yield calculation was carried out according to the amount of dried pectin and pectin was obtained with a yield of 8.91%. The obtained pomegranate pectin was hydrolyzed with commercial pectinase under 4 different conditions depending on the amount of enzyme and the hydrolysis time. For hydrolysis, 20 mg pectin/ml distilled water was prepared and pectinase enzyme was added to the solutions in a 50°C water bath at 4 pH. The reaction was started in a 50°C water bath. The reaction mixture was kept in 100°C water for 10 minutes to stop the reaction. The solutions, which came to room temperature, were centrifuged at 4000 rpm for 25 minutes and pectin was precipitated with ethyl alcohol. Re-freezing and drying in a lyophilizer were performed. The determination of the degree of esterification of the obtained pectin and pectin hydrolysates, the determination of galacturonic acid, FTIR and sugar analyzes were carried out. The degree of esterification (DE) of pomegranate pectin and its hydrolysates ranged from 34.36% to 49.90%, and low methoxyl pectins were obtained. Typical bands appeared in the spectrum, as expected from pectin molecules, which proved that the extracts were pectin in the FTIR analysis. The presence of arabinose, galactose, glucose, mannose and rhamnose in all pectin and its hydrolysates were analyzed and the presence of glucose and rhamnose was determined. Then, the effects of the obtained pectin and hydrolysates on Lactobacillus brevis viability and GABA production in the medium were investigated. In order to analyze the viability and GABA production potential of the strains, MRS broth medium with 2% monosodium glutamate was prepared. The pH of the medium was adjusted to 4.5 with 0.5 M citric acid solution before sterilization. Microorganism concentration was adjusted to 0.5 McF (108 cfu/mL) using sterile saline solution and inoculated at 1% rate into the prepared medium. All media were then incubated at 37°C for 72 hours at 100 rpm. Viability analysis was performed using the cast plate method using MRS Agar medium. The potential of the microorganism to produce GABA was realized by high performance liquid chromatography. According to the results of probiotic viability and GABA production in the medium, P4H2 hydrolyzate, which hydrolyzes for 2 hours with 400 µl pectinase/g, was chosen as the optimum hydrolysis conditions. Carrot juice containing pectin hydrosylate selected according to the viability and GABA analysis results was fermented with Lactobacillus brevis. Viability and GABA analyzes were performed. There is no study in the literature examining the effect of pectin hydrolyzate addition as a prebiotic on the fermentation of carrot juice with Lactobacillus brevis on GABA production and probiotic viability. For this reason, carrot juice containing P4H2 pectin hydrolyzate in different concentrations (1%, 2% and 4%) as a prebiotic was fermented with Lactobacillus brevis strain to provide a unique quality to the study. Probiotic viability was tested in fermented vegetable juice and GABA production potential was monitored by chromatographic determination methods. It was determined that fermented carrot juices containing P4H2 pectin hydrolyzate as a prebiotic had a higher GABA amount. In addition, the total phenolic content analysis of the obtained prebiotic fermented carrot juices was made according to the Folin-Ciocalteu method. The gallic acid equivalent (GAE) was given as mg/g using the gallic acid calibration curve for the total phenolic content. Two different methods, DPPH and CUPRAC, were used for the determination of total antioxidant activity. Trolox was used as standard in both methods. Looking at the results, P4H2 hydrosylate showed the highest effect on total phenolic substance and antioxidant activity with 4% concentration. As the ratio of P4H2 hydrosylate decreased, the amount of phenolic substance and antioxidant activity decreased significantly. As a result of the study, the production of GABA, a postbiotic component, by Lactobacillus brevis fermentation in pectin hydrolyzate added vegetable juice was investigated and a new functional beverage was developed.

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