Şerbetçi otu atığından selüloz elde edilmesi ve yenilebilir film formülasyonlarında kullanımının incelenmesi
Extraction of cellulose from spent hops and investigation of its use in edible film formulations
Yazar: ZEYNEP AKGÜN
Danışman: DR. ÖĞR. ÜYESİ CELALE KIRKIN GÖZÜKIRMIZI
İstanbul Teknik Üniversitesi / Lisansüstü Eğitim Enstitüsü / Gıda Mühendisliği Ana Bilim Dalı / Gıda Mühendisliği Bilim Dalı
Derece: Yüksek Lisans, 2024
Konu: Gıda Mühendisliği = Food Engineering

Özet:
Son yıllarda hızla artan nüfusla birlikte tüketimin artışı ve bununla birlikte gıda üretim ve atık miktarında önemli bir artış olmuştur. Bira üretimi sonucunda da önemli miktarda atık oluşmaktadır. Bira üretimi sırasında oluşan atıklar miktar olarak sırasıyla arpa, malt, şerbetçi otu, atık su, maya ve şıra atığıdır. Ayrıca kullanılan şerbetçi otunun yaklaşık %85'i yan ürün atığı oluşturmaktadır. Karbonhidratlar, proteinler, lifler, vitaminler, mineraller ve fenolik bileşikler açısından zengin olan bira atıklarının büyük miktarı değerli kaynaklara dönüştürülebilmektedir. 
Atıklardaki artışla birlikte çevre kirliliği ve ekonomik kayıpların önüne geçebilmek adına atıkların değerlendirilmesi konusunda çalışmalar geliştirilmiştir. Atıklar kimya, tıp, kozmetik, gıda gibi farklı alanlarda değerlendirilebilmektedir. Ambalaj materyali olarak kullanılan yenilebilir film ve kaplamalarda da son zamanlarda atıklardan elde edilen birçok yan ürün kullanılmaya başlanmıştır.
Yenilebilir film ve kaplamalar, gıdanın yüzeyine ya da gıda bileşenleri arasına yapıştırılan, ince tabakalı, gıdayla birlikte tüketilebilen polisakkaritler, proteinler, lipitler veya bunların birleşimi ile oluşturulan maddelerdir. Ayrıca bu film ve kaplamalar antioksidanlar, antimikrobiyal maddeler, probiyotik gibi biyoaktif bileşiklerin taşıyıcıları olarak kullanılmaktadır. Üretimden tüketime kadar olan süre içerisinde gıdaların kalite kayıplarını önlemek, mikrobiyal koruma ile gıda güvenliğini sağlamak, raf ömrünü uzatmak, duyusal özellikleri geliştirmek ve besin değerlerini korumak için alternatif ambalaj yöntemi olarak yenilebilir film ve kaplamalar geliştirilmiştir.
Bu çalışma, şerbetçi otu atığından selüloz ekstraksiyonu, selülozun modifikasyonu ile karboksimetil selüloz (CMC) elde edilmesi, filmlerin hazırlanması ve karakterizasyonu aşamalarını içermektedir. İlk aşamada şerbetçi otu atığı öğütülerek selüloz elde etmek için alkali ortamda geleneksel ve prob ultrason (20 kHz,  200 W) destekli iki farklı yöntem ile ekstrakte edilmiştir. Prob ultrason 10 dakika ve 30 dakika olacak şekilde uygulanmıştır. Prob ultrason uygulamasının ekstraksiyon verimi üzerinde etkisi olmadığı görüldüğü için geleneksel yöntem ile ekstraksiyona devam edilmiştir. İkinci aşamada elde edilen selüloza bazik ortamda sodyum mono klor asetat ile yapılan karboksimetilasyonu ile suda çözünürlük sağlanmıştır ve  CMC elde edilmiştir. Elde edilen selüloz ve CMC SEM analizi ile karakterize edilmiştir. Selüloz ve CMC görüntüleri incelendiğinde, selüloz yüzeyinin daha gözeneksiz ve çatlaksız ve yoğun olduğu, CMC'nin ise daha gözenekli ve kopuk yapıda ayrı polimerlere dönüştüğü görülmüştür.
Üçüncü aşamada, elde edilen CMC ile agar, jelatin ve gliserol kullanılarak 9 farklı formülasyonda film çözeltisi oluşturulmuştur ve bunlardan üçü ile film elde edilebilmiştir. Ayrıca, film çözeltisi oluşturmak için manyetik ısıtıcı ve orbital karıştırıcı olmak üzere iki farklı yöntem kullanılmıştır. Orbital karıştırıcı kullanılarak hazırlanan 7 farklı filmden sadece bir tanesi ile film oluşturulabilmişken, manyetik ısıtıcı ile oluşturulan iki farklı formülasyondaki çözeltinin ikisi ile film oluşturulabilmiştir. Bu sebeple çalışmaya manyetik ısıtıcı ile devam edilmiştir. Manyetik ısıtıcı kullanılarak elde edilen 0,15CMC:0,85Agar ve 1CMC:0,75Agar:0,25Jelatin formülasyonlarındaki yenilebilir filmler renk, opaklık, nem miktarı ve kalınlık analizleri ile karakterize edilmiştir. Elde edilen filmlerin renk, opaklık, nem miktarı ve kalınlık değerleri karşılaştırılmıştır. Nem miktarı ve kalınlık açısından elde edilen filmler arasında istatistiksel açıdan bir fark bulunmamıştır (p>0,05). Renk ve opaklık açısından ise filmler arasında istatistiksel açıdan önemli bir fark bulunmuştur (p<0,05). FİLM 2'nin opaklık değeri FİLM 1'den daha yüksek bulunmuştur. Filmlerde opaklık parametresine bakıldığında görsel olarak FİLM 1 tercih edilebilirken, güneş ışığının etkisinin önüne geçebilmek için ise FİLM 2 film tercih edilebilir. 
Sonuç olarak şerbetçi otu atığının değerlendirilerek selüloz elde edelebileceği ve böylelikle farklı alanlarda ham madde olarak kullanılabileceği ve şerbetçi otu atığının yenilebilir filmler için alternatif malzeme olarak kullanılabileceği konusunda umut vadetmektedir.

Abstract:
In recent years, there has been a significant increase in the amount of food production and waste with the increase in consumption with the rapidly growing population. These wastes cause both environmental pollution and economic losses. These wastes generated during production have started to be utilized with different methods. In this way, both environmental pollution can be prevented and economic losses are reduced. Especially recently, the reuse of agricultural wastes has been a subject of great interest. Industrial beer production also generates a significant amount of waste. These wastes are organic and can be easily recycled.  The wastes generated during beer production are barley, malt, hops, wastewater, yeast and wort waste. A large amount of beer waste, which is rich in carbohydrates, proteins, fibers, vitamins, minerals and phenolic compounds, can be converted into valuable resources. Approximately 85% of the hops used during beer production constitute by-product waste. Spent hops have a high content of protein, phenolic compounds and fiber. From 1 hL of beer production, 0.3 kg of hop waste is obtained. Since spent hops have a bitter taste, their direct use in feed is limited due to their low added value.
Since most of the plastics and polymers used as packaging materials, as well as production waste, are not recyclable and do not easily dissolve in nature, they cause great damage to the environment and cause economic losses. Materials such as plastic, glass and metal used as packaging materials create waste problems. Synthetic polymers used as food packaging materials are not sustainable and cause environmental problems. For this reason, the trend towards biodegradable polymers as packaging materials has increased and many by-products obtained from polysaccharides, lipids, proteins, microorganisms and agricultural wastes have started to be used instead of synthetic polymers. As a result, studies on edible films and coating have increased.
Interest in edible films and coatings as alternative packaging materials is increasing day by day. Edible films and coatings are thin-layer, food-consumable polysaccharides, proteins, lipids or combinations thereof that are adhered to the surface of food or between food components. In addition, these films and coatings are used as carriers of bioactive compounds such as antioxidants, antimicrobial agents, probiotics. Since the chemical structure of these three materials differs from each other, they give different properties to the film in film production. The hydrocolloid that is used has a major impact on the characteristics of edible films. Transparent, uniform edible films with moderate mechanical qualities are formed using polysaccharides. Nevertheless, the water solubility and low water vapor permeability of these films restrict their use. When compared to edible films made of polysaccharides, protein-based films can exhibit remarkable mechanical and gas barrier qualities; nevertheless, their limited water vapor resistance restricts their use. Generally, lipids are used to help reduce water transfer, polysaccharides to control the passage of gases and proteins to ensure that the films are mechanically durable. Edible films and coatings have been developed as an alternative packaging method to prevent quality losses, ensure food safety with microbial protection, extend shelf life, improve sensory properties and preserve nutritional values from production to consumption. These films have applications in many food categories such as meat and meat products, fruits and vegetables, dairy, seafood and bakery products and are used as a secondary packaging between the food and the primary packaging material.
This study includes the extraction of cellulose from spent hops, modification of cellulose to obtain carboxymethyl cellulose (CMC), preparation and characterization of edible films . In the first step, spent hops was ground and extracted in alkaline medium to obtain cellulose by two different methods: conventional and probe ultrasound assisted. Probe ultrasound was applied for 10 minutes and 30 minutes. While cellulose was obtained with a yield of % 35.78 with the traditional method, cellulose was obtained with a yield of %39.19 when probe ultrasound was used for 10 minutes and %36.22 when used for 30 minutes. Since it was observed that the probe ultrasound application had no effect on the extraction efficiency, the extraction was continued with the conventional method. In the second step, water solubility was achieved by carboxymethylation of the obtained cellulose with sodium mono chlorine acetate in basic medium and CMC was obtained. The modification of cellulose is both the more efficient use of carboxymethyl cellulose and its solubility is higher than cellulose and it is more widely used in edible films. The obtained cellulose and CMC were characterized by SEM analysis. When the images of cellulose and CMC were analyzed, it was observed that the cellulose surface was more porous, crack-free and dense, while CMC was transformed into separate polymers with a more porous and broken structure. In addition, the CMC morphological image obtained in the study is similar to the CMC images obtained from different materials in different studies.
In the third stage, 9 different formulations of film solutions were prepared using agar, gelatin and glycerol with the CMC obtained and three of them were able to obtain films. In addition, two different methods, magnetic heater and orbital stirrer, were used to form the film solution. Films could be formed with only one of the 7 different films prepared using the orbital heater. However, films could be formed with two of the two different formulations of the solution prepared with the magnetic heater. For this reason, the study was continued with the magnetic heater. The edible films in 0.15CMC:0.85Agar (FİLM 1) and 1CMC:0.75Agar:0.25Gelatin (FİLM 2) formulations obtained using magnetic heater.
In the fourth step, the films obtained were characterized by color, opacity, moisture content and thickness analysis. The color, opacity, moisture content and thickness values of the obtained films were compared. The color values of the films were measured by spectrophotometer according to the CIE L* a* b* color space. According to the results, it was observed that the L* and b* values of FİLM 1 were higher, thus it was lighter in color and more yellow compared to FİLM 2. The opacity values of the films were determined by measuring the absorbance values of the films between 400-800 nm wavelengths with UV-Vis spectrophotometer. Opacity values were calculated by dividing the absorbance values at 600 nm wavelength by the thickness of the films. FILM 1 opacity was calculated as 10.92 and FILM 2 opacity as 42.51 so the opacity of FİLM 2 was found higher than    FİLM 1. The light transmittance properties of the packaging materials used are important to protect food products from the harmful effects of light and to increase consumer acceptance. In these films, low opacity values are preferred for visual quality, while high opacity films are preferred to prevent bitterness due to the effect of sunlight. High opacity means that the films do not transmit light, that is, they reflect it. To determine the moisture content of the films, sections of the films were placed in an oven at 105 °C for 2 hours until constant weight. The moisture content of FİLM 1 was 39.18%, while the moisture content of FİLM 2 was 33.86%. There was no statistically significant difference between the two films (p>0.05). Water content in foods controls the growth of microorganisms as well as affecting food stability, appearance, texture and taste. Edible films with low moisture content prevent food damage and extend shelf life. High moisture content edible films facilitate microbiological spoilage. It is therefore important that the moisture content of the film is low. Film thicknesses were measured with a digital micrometer. FİLM 2 edible film measured 0.012 mm, FİLM 1 edible film measured 0.017 mm. There is no statistical difference between the thickness of the formed edible films (p>0.05). The thickness of the edible film affects the permeability rate of water vapor, gas and other volatile compounds. As the thickness increases, the water vapor and gas permeability also increases. As a result, there was no statistically significant difference between the films obtained in terms of moisture content and thickness (p>0.05). A statistically significant difference was found between the films in terms of color and opacity (p<0.05). 
Consequently, it is promising that cellulose can be obtained by valorizing spent hops and thus can be used as raw material in different fields and spent hops can be used as an alternative material for edible films. It is thought that FILM 1, which is relatively light in color, may be visually preferable, while FILM 2, which is dark in color, will provide better protection against food oxidation.