Physicochemical and textural properties of mozzarella cheese made with konjac glucomannan as a fat replacer

Accepted Manuscript Physicochemical and textural properties of mozzarella cheese made with konjac glucomannan as a fat replacer

Shuhong Dai, Fatang Jiang, Harold Corke, Nagendra P. Shah PII: DOI: Reference:

S0963-9969(18)30163-7 doi:10.1016/j.foodres.2018.02.069 FRIN 7432

To appear in:

Food Research International

Received date: Revised date: Accepted date:

3 January 2018 26 February 2018 27 February 2018

Please cite this article as: Shuhong Dai, Fatang Jiang, Harold Corke, Nagendra P. Shah , Physicochemical and textural properties of mozzarella cheese made with konjac glucomannan as a fat replacer. The address for the corresponding author was captured as affiliation for all authors. Please check if appropriate. Frin(2017), doi:10.1016/ j.foodres.2018.02.069

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ACCEPTED MANUSCRIPT Physicochemical and textural properties of Mozzarella cheese made with konjac glucomannan as a fat replacer Shuhong Daia, Fatang Jiangb, Harold Corkec, Nagendra P. Shaha,* a

Food and Nutritional Sciences, School of Biological Sciences, The University of

Glyn O. Philips Hydrocolloid Research Centre at HUT, Hubei University of

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Hong Kong, Pokfulam Road, Hong Kong

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Technology, Wuhan 430068, China

Department of Food Science and Engineering, School of Agriculture and Biology,

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Shanghai Jiao Tong University, Shanghai, China

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* Corresponding author: Prof. Nagendra P. Shah; Food and Nutritional Sciences, School of Biological Sciences, The University of Hong Kong, Pokfulam Road, Hong

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Kong; Tel.: +852 2299 0836; fax: +852 2299 9114; E-mail address: [email protected].

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Graphical abstract

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ACCEPTED MANUSCRIPT Highlights • Konjac glucomannan was utilized in Mozzarella cheese as a fat replacer • Mozzarella cheese with konjac had higher moisture and water activity • Mozzarella cheese with konjac showed whiter, more greenish and more yellowish

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color

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• Mozzarella cheese with konjac influenced firmness and stickiness textural

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parameters

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• Konjac can improve fat-reduced Mozzarella cheese

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ACCEPTED MANUSCRIPT ABSTRACT Konjac glucomannan (KGM) is a natural polysaccharide with several favorable nutritional characteristics, and exhibits functional properties as a potential fat-replacer in dairy products. In our study, composition, color and browning (L*, a* and b*

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before and after heating), and textural characteristics of low-fat and skimmed

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Mozzarella cheese with KGM (LFKGM and SKKGM) were compared with those of

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full-fat, low-fat and skimmed Mozzarella cheese controls (FFC, LFC and SKC) after 0, 7, 14, 21 and 28 days of storage at 4 °C. In general, LFKGM and SKKGM had

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similar composition to LFC and SKC, respectively, except that LFKGM had higher

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moisture than LFC and SKKGM had high aw than SKC. The LFKGM and SKKGM had higher L* (lightness) than LFC and SKC, respectively, and LFKGM had similar

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whiteness to FFC before and after heating. However, the browning factor was not

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affected by KGM addition. The a* values (greenness) of LFKGM and SKKGM were more negative than for LFC and SKC before and after heating. The b* values

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(yellowness) of LFKGM and SKKGM were higher than LFC and SKC, respectively.

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Grated SKKGM exhibited lower firmness than SKC, and LFKGM exhibited higher stickiness than LFC. The melted LFKGM and SKKGM had similar resistance and stretch quality to LFC and SKC when they were stretched, respectively. The changes in the lightness, moisture and firmness as affected by KGM addition in the cheeses were more close to those of full-fat cheese compared with the cheeses without KGM, indicating KGM would be a potential fat replacer to be used in Mozzarella cheese. Key words: polysaccharide; dairy; application; interaction 4

ACCEPTED MANUSCRIPT Abbreviations: KGM, konjac glucomannan; FFC, full-fat Mozzarella cheese control; LFC, low-fat Mozzarella cheese control; SKC, skimmed Mozzarella cheese control; LFKGM, low-fat Mozzarella cheese with KGM addition; SKKGM, skimmed

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Mozzarella cheese with KGM addition; BF, browning factor

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ACCEPTED MANUSCRIPT 1. Introduction Mozzarella cheese is a popular dairy product and its consumption has grown dramatically during the past decades (O'Reilly et al., 2002). Recent research indicates that consumption of regular-fat dairy foods is not associated with an increased risk of

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cardiovascular disease and is inversely associated with metabolic syndrome, weight

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gain and the risk of obesity (Astrup et al., 2016; Drehmer et al., 2016). However,

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excessive consumption of dairy products could still lead to weight gain and cause some health disorders (Nagai, Uyama, & Kaji, 2013; Raza et al., 2017). Therefore, the

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reduction in fat content of Mozzarella cheese is desirable for some consumers and

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fat-reduced dairy foods still have considerable market value. Fat performs many important functions within a food (McClements & Demetriades, 1998), removal of fat

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from Mozzarella cheese causes defects such as rubbery texture and undesirable color

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(Fife, McMahon, & Oberg, 1996). Besides the reduction of fat in dairy foods, ingredient addition to provide specific functional properties is also of interest to

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scientists and consumers (Palatnik et al., 2017).

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Using fat replacers in low-fat or fat-free products is a common strategy to improve the physicochemical properties of foods. The functionality of fat replacers based on carbohydrates is due to their ability to increase gel formation and viscosity, to provide taste and texture, and to increase water retention capacity (Dervisoglu & Yazici, 2006). Zisu & Shah (2005) studied the influence of two fat replacers (one was modified corn starch-based product, and another one was a blend of α-lactoglobulin, carrageenan and xanthan gum) on the textural characteristics of low-fat Mozzarella cheeses. The 6

ACCEPTED MANUSCRIPT addition of the two different fat replacers increased the moisture content in cheeses and softened the cheese texture. Inulin, as an important carbohydrate, was also used as a fat replacer in various cheeses. It was reported to improve textural properties of low-fat Mozzarella and Cheddar cheeses (Wadhwani, 2011), and was beneficial in

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manufacturing different kinds of fat-reduced cheeses (Karimi et al., 2015).

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Plants of the genus Amorphophallus have a long history for use as a food source in

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subtropical and tropical Asia (Chua et al., 2010). Konjac glucomannan (KGM) is a natural hetero-polysaccharide extracted from the tuber of Amorphophallus konjac

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(Nishinari, Williams, & Phillips, 1992). It is a linear random copolymer of

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β-1,4-linked D-mannose and D-glucose at a ratio of 1.6:1 (Kato & Matsuda, 1969). There is a low degree of acetyl groups at the C-6 position together with some limited

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branching at the C-3 position, which is responsible for its solubility in water (Davé &

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McCarthy, 1997). The soluble, stable and high molecular weight properties of KGM give its thickening properties, gelling behavior, water holding capacity and

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biocompatibility (Davé & McCarthy, 1997; Tomczyńska‐Mleko et al., 2014). KGM

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is dispersible in hot or cold water, and KGM solution exhibits very high viscosity with values higher than other colloids such as guar and locust bean gum (Davé & McCarthy, 1997; Vanderbeek et al., 2007). All these physicochemical properties of KGM give it the potential to be used as a fat replacer in Mozzarella cheese. KGM is also used as a food additive and dietary supplements or nutraceuticals (Chua et al., 2010). Some literature reported that KGM is beneficial to the health of human beings and it has an ability to cure some diseases (Walsh, Yaghoubian, & 7

ACCEPTED MANUSCRIPT Behforooz, 1984; Huang, Zhang, & Peng, 1990; Vuksan, Jenkins, & Spadafora, 1999). KGM has been reported to promote weight loss (Kraemer et al., 2007), and it has anti-hyperglycemic and hypercholesterolemia activities, anti-inflammatory activity and prebiotic activity (Fang & Wu, 2004; Chen et al., 2006; Chua et al., 2010). The

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nutraceutical properties of KGM also make it a potential fat replacer for use in dairy

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products.

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Very few studies reported the application of KGM as fat replacer in dairy products. da Silva et al. (2016) improved the rheological and textural properties of low-fat

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processed cheese by KGM addition. Our previous work (Dai, Corke, & Shah, 2016)

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also used KGM as a fat replacer in low-fat/skimmed yogurt, and the textural characteristics and structure of the yogurts was improved by KGM addition. However,

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the application of KGM as fat replacer in dairy products is still unexploited and there

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are no published studies investigating the KGM used as fat replacer in Mozzarella cheese. In this study, KGM was added to milk before the formation of cheese curd in

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the manufacturing of low-fat and skimmed Mozzarella cheeses. The aim was to

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evaluate the possibility of using KGM as fat replacer in Mozzarella cheese and to investigate the effects of KGM addition on physicochemical and textural properties of low-fat/skimmed Mozzarella cheese during storage. 2. Materials and methods 2.1. Materials Whole, low-fat and skimmed cow’s milk (fresh, with protein content of 3.6, 3.8, 3.6 g/100 mL and fat content of 5.1, 2.5, 0.7 g/100 mL, respectively) used in this study 8

ACCEPTED MANUSCRIPT were purchased from a local milk company (Farm Milk Company Limited, Hong Kong). Konjac purified flour (KJ30, KGM content ≥ 85%) was provided by Hubei Konson Konjac Gum Co., Ltd. (Wuhan, Hubei Province, China). 2.2. Cheese making

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Konjac purified flour was dissolved into sterile Milli-Q water at 22 °C to make

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0.5% KGM solution and which was stored at 4 °C overnight to allow complete

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hydration before use. Each experimental Mozzarella cheese was manufactured according to Ayyash & Shah (2011a) with some modification. The Mozzarella cheeses

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manufactured using full-fat, low-fat and skimmed milk without KGM addition were

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named as full-fat control cheese (FFC), low-fat control cheese (LFC) and skimmed control cheese (SKC), respectively, and the cheeses manufactured using low-fat and

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skimmed milk together with KGM were named as LFKGM and SKKGM. Milk (5 L

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for manufacture of Mozzarella cheese without KGM or 4 L for manufacture of Mozzarella cheese with KGM addition) was poured into a cheese vat (FT20-A,

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Armfield Limited, Ringwood, England), tempered to 40 °C and inoculated with

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STI-13 starter culture (Chr. Hansen, Bayswater, Victoria, Australia) consisting of Streptococcus thermophilus. The amount of the starter culture added was about 0.1 g starter culture per 1 L milk. After 45 min, chymosin (CHY-MAXTM PLUS, Chr. Hansen, Bayswater, Victoria, Australia) diluted (1:20) with distilled water was added at a rate of 10 mL per 25 L of milk. For manufacture of Mozzarella cheese with KGM, a mixture of 250 mL KGM solution and 1 L milk which was heated at 85 °C for 30 min under magnetic stirring to facilitate the interaction of KGM and milk, then, the 9

ACCEPTED MANUSCRIPT mixture was cooled to 40 °C and added into the cheese vat at the same time as chymosin addition. The final concentration of KGM added in milk was 0.25 g KGM per 1 L milk. The milk coagulated after 35 min. The curd was cut into 1 cm cubes and cooked at 40 °C with continuous agitation for 15 min. The whey was drained and curd

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was manually pressed to release additional whey. The curd was left in the cheese vat,

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cheddared and milled when the pH of the slabs reached 5.3 to 5.2. The curd was

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weighed and salted at a level of 1.5% (w/w), and then allowed to mellow for another 20 min in the cheese vat. The salted curd was then hand stretched for 7 min in hot

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water containing 4% salt at 85 °C, and then manually kneaded in blocks

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(approximately 100 g per block). Each block was vacuum-packed into barrier bags using a vacuum package machine (MS1160, Magic Seal, Kai Shi Electric Limited,

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Dongguan, China), and then stored at 4 °C. Properties of experimental cheese were

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tested at 0, 7, 14, 21 and 28 days of storage. 2.3. Cheese composition

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The protein content was determined by the Kjeldahl method, and fat content by the

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Babcock method (AOAC International, 1999). The moisture content was determined using a moisture analyzer (HB43-S, Mettler Toledo, Switzerland). The water activity (aw) was measured using a water activity meter (4TEV, AquaLab, Decagon Devices, Pullman, USA). Actual cheese yield was determined by dividing the weight of cheese by the total weight of the materials (milk/milk and KGM solution) used to make cheese multiplied by 100. The pH was measured according to Ayyash & Shah (2011b) with some 10

ACCEPTED MANUSCRIPT modification. Grated cheese (10 g) was macerated with 20 mL of distilled water, and the pH of the slurry was measured using a pH meter (Model 250A; Science International Corporation, Hong Kong). 2.4. Color and browning analysis

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The color and browning of cheese were determined according to Barbano, Yun, &

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Kindstedt (1994) with some modification. An aliquot of the ground cheese was

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weighed (15 g) into an aluminum pan (7.5 cm in diameter and 1.5 cm high), and then the pans containing the cheese samples were spread out in a preheated forced-air oven

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at 100 °C for 1 h. A Minolta Chroma Meter (CR-300 Series, Minolta, Osaka, Japan)

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was used to measure the color of cheeses before heating. A standard white plate provided with the meter was used to calibrate the Chroma Meter. The color of the

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cheeses after heating and cooling to 22 °C was also measured; this represented

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browning of cheeses. The L* (lightness), a* (red/greenness) and b* (yellow/blueness) values were taken on four different places for each sample.

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A browning factor (BF), defined as the ratio of L* value before and after heating,

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was used to describe the browning property as: BF (%) = L* value before heating/L* value after heating × 100. 2.5. Texture analysis Texture analysis of cheese was performed using a TA.XT Plus model texture analyzer (Stable Micro Systems, Godalming, Surrey, UK) with a 5 kg load cell at 22 °C. The values of texture attributes were analyzed from the resulting graphs using the Exponent version 6.1.4.0 equipment software. 11

ACCEPTED MANUSCRIPT 2.5.1. Softness of grated cheese A TTC Spreadability Rig (HDP/SR, a set of precisely matched male and female perspex cones) and a Heavy Duty Platform (HDP/90, a standard base for fitting many attachments for the Texture Analyser) was used for this experiment. The grated cheese

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was placed into the female cone and pressed it down to eliminate air pockets. The

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excess sample was scraped off with a knife, to leave a flat test area. Before testing, the

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male cone probe was calibrated against the female cone, making the starting point to be at the same height for each test (25.0 mm) above the female cone. The test speed

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and distance were set to 3.0 mm/s and 20 mm, respectively.

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2.5.2. Assessment of the stretch quality of melted cheese A Cheese Extensibility Rig (A/CE, a probe set used for testing stretching quality of

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melted cheese, consisting of a vessel and double-sided fork probe), a PT100

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Temperature Probe and a Flexible Clamping Arm were used for this experiment. The cheese samples were removed from the refrigerator just before testing and cut into

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small cubes. Cheese cubes (40 g) were filled evenly around the fork without

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relocating the fork within the pot. The sample pot with fork and cheese were heated for 60 s in a microwave oven to create a molten mass. Once the cheese sample was melted, the cheese retainer was inserted firmly into the sample pot. The sample pot assembly was then pushed into the slotted base and the PT100 probe was inserted into the cheese carefully, where it did not make contact with the fork. Once the temperature reached 55 °C the test commenced. The test speed and distance were set to 20 mm/s and 270 mm, respectively. 12

ACCEPTED MANUSCRIPT 2.6. Statistical analysis Each cheese sample was made in triplicate and analysis was carried out in each of the triplicate replicates. Data were reported as mean ± standard deviation for triplicate replicates of each sample. One-way analysis of variance (ANOVA) and Tukey’s test

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were performed at 95% confidence intervals (P < 0.05) using IBM SPSS Statistics

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20.0 (Armonk, NY). ANOVA and Tukey’s test were applied for different cheeses at

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each storage time and also for each cheese at different storage time. 3. Results and discussion

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3.1. Composition

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Reduction of weight loss due to water evaporation is the main advantage claimed for vacuum packaging of cheese, but not all weight loss may be prevented (Nunez et

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al., 1986). When cheese is packaged under vacuum using a high barrier plastic

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material, the cheese presents a wet surface due to water migration to the surface (Pantaleao, Pintado, & Poças, 2007). Thus, the compositional changes of cheese by

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vacuum packaging during storage was necessary in this study.

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3.1.1. Fat content

The LFC had a higher fat content than LFKGM during storage, while SKC had similar fat content to SKKGM (Table 1). Heat treatment of milk before cheese making causes some defects in cheese manufacture, however, the defects with respect to moisture retention and texture were reported to be minor (Singh & Waungana, 2001). The effect of heat treatment of milk on the changes in properties of the cheeses was not considered in this paper. LFKGM and SKKGM had higher actual cheese yield 13

ACCEPTED MANUSCRIPT (9.92 and 7.93%, respectively) than LFC and SKC (9.25 and 6.80%, respectively). The fat content is defined as the ratio of fat weight to cheese weight, the cheese weight is related to cheese yield. As the fat content in milk to make cheese is constant, the higher cheese yield of LFKGM led to slightly lower fat content than LFC.

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However, for SKC and SKKGM, because the fat content was quite low, the yield

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difference was not enough to affect the fat content in the cheese. There were no

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significant changes in the fat content of all the cheeses during storage within a cheese sample. The unchanged fat content of low-moisture Mozzarella cheese during storage

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was also reported by Ayyash & Shah (2011a).

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3.1.2. Protein content

For the Mozzarella cheese made from milk with lower fat content, the protein

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content in the cheese was higher (Table 1). This is in accordance with the results of

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Rudan et al. (1999), who observed higher protein content as the fat decreased in fat-reduced Mozzarella cheeses. The same authors explained that the results may be

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observed because the moisture did not replace the fat on an equal basis. The protein

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contents of LFKGM and SKKGM were similar with those of LFC and SKC, respectively. This suggested that the KGM addition did not affect the protein content of the Mozzarella cheeses. The protein content of all the cheeses did not change during storage. This is in accordance with the results of Ayyash & Shah (2011a) that the protein content of low-moisture Mozzarella cheese did not change during storage. 3.1.3. Moisture The SKC and SKKGM had higher moisture content than other cheeses (Table 1). 14

ACCEPTED MANUSCRIPT This may be because of higher protein content in SKC and SKKGM that made more water absorbed in the protein matrix (Mistry & Anderson, 1993). McMahon & Oberg (1998) also reported a higher moisture in the fat-reduced Mozzarella cheese than the full-fat control cheese. LFKGM had higher moisture than LFC for the first 7 days; it

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indicated that the KGM addition increased the moisture in low-fat Mozzarella cheese.

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This may be because of the high viscosity of KGM that slightly increased the water

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retention capacity of the cheese. However, LFKGM and LFC showed no significant difference in moisture between them after 7 days of storage, and SKKGM also had

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similar moisture to SKC during storage. Oberg et al. (2015) reported that low-fat

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Mozzarella cheese with xanthan gum addition had similar moisture to the low-fat control cheese, and they thought it was because only a small amount (2%, w/w) or

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less slurry was added in the low-fat Mozzarella. Thus, the small amount of KGM

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(2.5%, w/w) in our study may lead to the unchanged moisture in skimmed Mozzarella cheese by KGM addition. According to our previous work (Dai et al., 2017),

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incompatibility between the polysaccharide and milk protein could occur when KGM

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is mixed with milk above a certain level, and it may affect the curd formation during cheese manufacture. Thus, the 0.5% KGM solution was added to milk at a level of 5%, which was reported to be a stable system for making cheese (Dai et al., 2017), and heat treatment (85 °C for 30 min) was introduced to promote the interaction between KGM and milk before cheese making. The reason why KGM increased moisture in low-fat Mozzarella cheese but did not change the moisture in skimmed Mozzarella cheese is unknown; it may be related to the complicated interaction of KGM, casein 15

ACCEPTED MANUSCRIPT and fat in cheese during cheese manufacturing and storage. The moisture content of FFC, SKC and SKKGM remained stable during storage, and that of LFC and LFKGM had no significant difference between day 0 and 7, 7 and 14, 14 and 21, 21 and 28. This result is in accordance with that of Ayyash & Shah (2011a) and

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McMahon & Oberg (1998) that the moisture of Mozzarella cheeses did not change

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during storage.

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3.1.4. Water activity

The processing, product quality and stability of a food are not only related to the

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amount of water but also to the state of the water in the food system. Thus, water

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activity (aw) is an important parameter within a food, as it describes the level of the water in bound state and is depended on the water interaction with other components

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(Duggan et al., 2008). The KGM addition in low-fat and skimmed Mozzarella cheese

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increased the aw. Therefore, the LFKGM had higher aw than LFC only at day 0, and SKKGM had higher aw than SKC at day 7, 14 and 21 (Table 1). Duggan et al. (2008)

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also reported that the addition of starch in an imitation cheese (imitation cheese was

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made with dry casein instead of milk) increased aw slightly. When water activity increases, less energy is needed for the water to escape from the binding site to the protein matrix in the cheese, subsequently, the cheese melts more easily (Ma et al., 2013). 3.1.5. pH changes The pH of all the cheeses was higher than 5.2 (Table 1), which was the pH of the cheese curd at salting. The increase in pH between salting and storage time in 16

ACCEPTED MANUSCRIPT Mozzarella cheese has also been observed in some studies (Feeney, Fox, & Guinee, 2001; Guinee et al., 2002; Sheehan & Guinee, 2004). The increased pH may be caused by the loss of lactic acid, changes in soluble calcium and phosphate in the stretch water and re-solubilization of micellar calcium phosphate of the curd during

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plasticization (Sheehan & Guinee, 2004). Generally, the pH did not differ

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significantly among the cheeses. A similar result was reported by Rudan et al. (1999)

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that the pH of Mozzarella cheese was not affected by fat reduction. Rudan, Barbano, & Kindstedt (1998) also reported that the pH of Mozzarella cheese with fat replacer

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(Salatrim®) did not differ with the control cheese. Generally, the pH of the cheeses

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remained stable during storage. This trend was different from the results reported by Barbano, Yun, & Kindstedt (1994) for low-moisture Mozzarella cheese and by

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Sheehan & Guinee (2004) for fat-reduced Mozzarella cheese that the pH value

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decreased over time. In addition, it was also different from the results reported by Guo, Gilmore, & Kindstedt (1997) and Guinee et al. (2002) that the pH value of Mozzarella

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cheese increased over time. The different trend of pH change during storage may be

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related to the differences in the cheese manufacture procedure, composition of cheese, buffering capacity and thermal inactivation of starter culture (Sheehan & Guinee, 2004).

3.2. Color and browning 3.2.1. Lightness The L* value is an estimation of food lightness and goes from 0 to 100 (darkest black to brightest white, respectively), higher L* value represents a whiter sample 17

ACCEPTED MANUSCRIPT (Zare et al., 2011). The whiteness of Mozzarella cheese can be affected by fat and protein matrix in cheeses and by the change in serum phase confined in the matrix during heating (Metzger et al., 2000). The L* value of ground cheese samples before (L*) and after (browning L*) heating is shown in Figure 1 (A and B, respectively).

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The color L* values and browning L* values of FFC and LFKGM did not differ, and

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were the highest among the cheeses during storage. SKKGM had lower color L*

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value and browning L* values than LFC, but higher values than SKC during storage. The results indicated that fat reduction decreased the whiteness of the Mozzarella

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cheeses before and after heating. It is because fat can contribute to the whiteness of

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dairy products by scattering light (Quiñones, Barbano, & Philips, 1998; Rudan et al., 1998b). In addition, the results also illustrated that the addition of KGM in low-fat

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and skimmed Mozzarella cheeses enhanced the whiteness of the cheeses and gave the

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low-fat Mozzarella cheese a similar whiteness to full-fat cheese before and after heating. It may be because KGM promoted the aggregation of casein and peptides in

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colloidal suspension as white particles in cheese and subsequently increased the

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amount of light reflected from the cheese (Metzger et al., 2000). And the white color of KGM flour may also be responsible for the increased whiteness in KGM added cheeses (Dai, Corke, & Shah, 2016). The L* value of all the cheeses decreased slightly during 28 days of storage. The decreased L* value with time may have been caused by time-dependent changes in the serum phase of cheeses and degradation of the casein matrix in Mozzarella cheese (Rudan, Barbano, & Kindstedt, 1998; Metzger et al., 2000; Metzger et al., 2001). 18

ACCEPTED MANUSCRIPT BF represents how dark the cooked cheese is compared to its uncooked status, and larger values of BF represent higher changes in the lightness of the cheese after heating (Wang & Sun, 2003). Generally, the lightness of Mozzarella cheese was expected to decreased after heating, therefore, BF of the cheese should exceed 100%

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(Wang & Sun, 2003). BF of FFC, LFC and LFKGM exhibited no significant

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differences among each other, and were higher than those of SKC and SKKGM at

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day 7, 14 and 28 (Figure 1, C). BF of SKC and SKKGM exhibited no significant differences between each other at each tested day. The results illustrated that KGM

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cheeses remained stable during storage.

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addition did not affect the BF of the Mozzarella cheeses. On the whole, BF of the

3.2.2. Greenish and yellowish color

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The a* values of all the cheeses were negative before heating during the storage

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period (Figure 2), indicating a greenish characteristic in the cheeses. The absolute values of color a* and browning a* of LFKGM and SKKGM were higher than LFC

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and SKC, respectively. As fat replacer would cause a dense microstructure of casein

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matrix in Mozzarella cheese, the greenish characteristic of Mozzarella cheeses by the KGM addition may be because that the complicated interactions between KGM, casein and fat may lead to a dense structure of the cheeses with KGM, which was the main substance that contributed to light scattering in the cheeses (Zisu & Shah, 2005). The greenish color (-a* value) of all the cheeses decreased during 28 days of storage. The b* values of all the cheeses were positive before heating during the storage period (Figure 3), indicating yellowish characteristics of the cheeses. Generally, the 19

ACCEPTED MANUSCRIPT b* values of LFKGM and SKKGM were higher than those of LFC and SKC, respectively. It indicated that the addition of KGM increased yellowness of the Mozzarella cheeses. The increased yellowish color of the cheese by KGM addition may be caused by the Maillard reaction of milk protein and KGM when heated at

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85°C for 30 min (Dai, Corke, & Shah 2016). The addition of KGM was also reported

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to increase lightness (L*) and yellow color (b*) of surimi gels (Park, 1996). The color

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b* values of all the cheeses remained stable during storage. 3.3. Texture

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3.3.1. Softness of grated cheese

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Mozzarella cheese is expected to have unique functionalities in both un-melted and melted states (Imm et al., 2003). For un-melted Mozzarella cheese, its shred state and

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overall texture are important functionalities (Kindstedt, 1991), and it is very

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frequently used in the grated form (Pluta, Ziarno, & Kruk, 2005). For Mozzarella cheese that is sold in grated form in the package, its textural properties are more

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important when it is subjected to extrusion. The textural study on the grated cheeses

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could give instrumental assessment of the internal changes in cheeses. However, most studies on the texture of Mozzarella cheese are focused on the texture of cheese blocks, no research has reported the texture of grated Mozzarella cheese. Firmness represents the maximum force during the probe penetration to shear in texture test. The firmness of SKKGM was lower than for SKC after 14 days of storage, while the firmness of LFKGM was similar to that of LFC during storage (Table 2). The decreased firmness of skimmed Mozzarella cheese by KGM addition 20

ACCEPTED MANUSCRIPT may be because KGM could play the role of a lubricant, it may break up the protein matrix to provide a softer texture of the cheese (Romeih et al., 2002; Tang and Wang, 2007). While for low-fat Mozzarella, because the higher fat content and lower protein content, the lubricant role of KGM was not obvious, thus, it showed no significant

PT

difference in the firmness of LFC and LFKGM. Addition of fat replacer (Salatrim®)

RI

decreased the hardness of fat-reduced Mozzarella cheese as also reported by Rudan,

SC

Barbano, & Kindstedt (1998). The firmness of FFC, LFC and LFKGM remained stable while that SKC and SKKGM decreased at day 28 compared with that at day 0.

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The decreased firmness of Mozzarella cheeses during storage has also been reported

MA

by Guinee, Feeney, & Fox (2001) and Zisu & Shah (2007), and it was thought to be related to the hydration of the protein matrix and proteolytic changes. While the

D

unchanged firmness of FFC, LFC and LFKGM may be related to the higher fat

PT E

content and lower protein content that slowed down the proteolysis in the cheese matrix.

CE

The “work of shear” represents the total amount of force required to perform the

AC

shearing process during the texture test. LFKGM and SKKGM had similar “work of shear” to LFC and SKC, respectively (Table 2). Mulvaney, Barbano, & Yun (1997) reported that work of shear was related to the extensive elastic network in cheeses. Thus, the elastic network of the cheeses was not affected by KGM addition, it may be because of the small amount of KGM added in cheeses. The work of shear of the cheeses went up and down during storage. Totally, those of SKC and SKKGM decreased while those of FFC and LFKGM remained stable after 28 days of storage 21

ACCEPTED MANUSCRIPT compared with those at day 0. The fluctuation of the work of shear in cheeses may be because of the complicated molecular interactions (such as electrostatic and hydrophobic interactions etc.) between protein, fat, and polysaccharide during cheese ripening.

PT

The stickiness represents the maximum negative force that the probe proceeds to

RI

withdraw from the sample. A stickier sample will require a greater force to remove the

SC

probe. FFC had higher stickiness (absolute value, similarly hereinafter) than the other cheeses (Table 2), this may be because of the more elastic network of cheese with

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lower fat content that made the lower fusion of the grated cheese in this particular test.

MA

The stickiness of LFKGM was higher than LFC, while that of SKKGM was similar to that of SKC. It may be because of the high viscosity and the lubricant role of KGM

D

that increased the stickiness of low-fat Mozzarella cheese. The stickiness of SKC,

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LFKGM and SKKGM increased after 28 days storage, this was in accordance with Kindstedt et al. (1995) and Chen, Wolle, & Sommer (2009) that the low-moisture,

CE

part-skim Mozzarella cheese became more adhesive and sticky with age.

AC

The “work of adhesion” represents the negative force region on the texture curve that the probe proceeds to withdraw from the sample and indicates the adhesive characteristics of the sample. Work of adhesion of FFC, LFC and LFKGM was negative while those of SKC and SKKGM were positive and close to zero during storage (Table 2). The adhesion of LFKGM and SKKGM was similar to that of LFC and SKC, respectively, except that of LFKGM was more negative than LFC at day 0 and 21. It indicated that KGM addition increased the adhesiveness of the low-fat 22

ACCEPTED MANUSCRIPT cheese, and it may be also caused by the high viscosity and lubricant role of KGM in cheese.

FFC, LFC and LFKGM showed increased work of adhesion, while SKC and

SKGM showed stable adhesion during 28 days of storage. Zisu & Shah (2007) reported similar results that the work of adhesion was low in low-fat Mozzarella

PT

cheeses and increased with storage time.

RI

3.3.2. Stretch quality of melted cheese

SC

For melted Mozzarella cheese, the stretching property is an important functionality. The force at 5 s is taken as the resistance to extension and indicates the toughness of

NU

the melted cheeses. The tougher the melted cheese, the higher is this force value. The

MA

resistance of LFKGM and SKKGM was similar to that of LFC and SKC, respectively (Table 3). Thus, the KGM addition did not change the toughness of the melted

D

Mozzarella cheeses to be stretched. The KGM addition did not affect the resistance in

PT E

cheeses, possibly because of the complicated interactions between KGM, casein and fat may lead to a dense structure of the cheeses with KGM (Zisu & Shah, 2005).

CE

Generally, the resistance of the cheeses decreased during storage, indicating the less

AC

tough texture of the melted cheeses to be stretched. Zisu & Shah (2007) reported that low-fat Mozzarella cheese became more pliable in stretch appearance over storage time. The more pliable stretch texture of the cheeses with time may be caused by the complicated interactions of the substances in cheese, such as proteolysis and lipolysis. Stretch quality represents the gradient of stretch region in the test plot. The greater the gradient over this region the tougher is the stretching quality. Similar to the results of resistance, LFKGM and SKKGM had similar stretch quality to LFC and SKC, 23

ACCEPTED MANUSCRIPT respectively (Table 3). In addition, the stretch quality of the cheeses also decreased during storage. In order to have a better understanding of fat-reduced Mozzarella cheese with KGM addition, sensory evaluation of the cheeses is necessary. However, as this paper

PT

is focused on the physicochemical and textural properties of the cheeses, the sensory

RI

evaluation was not performed. Further studies about the sensory performance of the

SC

cheeses is needed to describe the acceptance of the Mozzarella cheese with KGM addition, and the sensory evaluation may involve in progressive profiling/ temporal

NU

dominance of sensations (Esmerino et al., 2017), rapid sensory profiling methods

MA

(Fonseca et al., 2016), check-all-that-apply questions (Oliveira et al., 2017) or descriptive analysis using a trained panel (Gaze et al., 2017).

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4. Conclusions

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Our study suggested that KGM addition mainly affected the color and browning of the cheeses, and slightly increased moisture, aw of the cheeses, and texture of the

CE

grated cheeses was also affected by KGM with lower firmness and higher stickiness.

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And the changes of the lightness, moisture and firmness as affected by KGM addition in the cheeses were more close to those of full-fat cheese compared with the cheeses without KGM, indicating KGM could improve some characteristics of the fat-reduced Mozzarella cheeses. Thus, KGM could be a potential fat replacer to be used in fat-reduced Mozzarella cheeses. References AOAC International. (1999). Official Methods of Analysis. Vol. II. 5th edition. 24

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cultures.

ACCEPTED MANUSCRIPT Table 1. Composition of cheeses during 28 days storage at 4 °C1. Storage Time (Day)

Sample2 0

7

14

21

28

FFC

31.62±0.34a,A

31.82±0.25a,A

32.46±0.04a,AB

33.13±0.26a,B

33.14±0.28a,B

LFC

23.29±0.34b,AB

22.83±0.27b,B

24.47±0.03b,A

24.13±0.30b,A

24.30±0.28b,A

SKC

3.99±0.01c,A

4.33±0.29c,A

4.33±0.29c,A

4.33±0.29c,A

4.66±0.28c,A

LFKGM

22.48±0.02d,A

22.15±0.28b,A

22.31±0.27d,A

22.63±0.27d,A

22.82±0.27d,A

SKKGM

4.16±0.29c,A

4.16±0.28c,A

4.16±0.29c,A

4.16±0.29c,A

4.49±0.00c,A

FFC

20.31±0.61a,A

20.55±0.26a,A

20.50±0.38a,A

20.88±0.15a,A

20.21±0.23a,A

LFC

24.65±0.58b,A

26.72±0.30b,B

26.35±0.46b,AB

26.81±0.25b,B

25.61±0.17b,A

SKC

40.14±0.56c,AB

41.67±0.31c,A

41.87±0.61c,AB

40.68±0.34c,AB

40.40±0.34c,B

LFKGM

25.10±0.27b,A

25.41±0.25d,A

25.51±0.26b,A

25.59±0.23d,A

25.26±0.13b,A

SKKGM

39.83±0.12c,A

40.05±0.28e,AB

40.40±0.22c,AB

40.83±0.13c,B

39.73±0.52c,AB

FFC

41.90±0.41ab,A

41.70±0.53a,A

41.83±0.61a,A

43.13±0.35a,A

41.78±0.49a,A

LFC

40.77±0.27b,A

41.00±0.85a,AB

41.80±0.48a,AB

41.81±1.03a,AB

42.49±0.42ac,B

SKC

46.49±0.41c,A

47.07±0.39b,A

47.14±0.32b,A

47.60±0.34b,A

47.65±0.25b,A

LFKGM

42.41±0.38a,A

44.26±0.72c,AB

43.91±1.11ab,AB

43.77±0.31a,B

43.82±0.21c,B

SKKGM

47.52±0.69c,A

47.95±0.27b,A

47.00±0.44b,A

47.07±0.16b,A

47.27±0.33b,A

FFC

0.9619±0.0022a,A

0.9665±0.0012ab,A

0.9643±0.0020ab,A

0.9668±0.0014a,A

LFC

0.9545±0.0017b,A

0.9545±0.0018ab,AB

0.9623±0.0018ab,B

0.9631±0.0022ab,B

0.9636±0.0014a,B

SKC

0.9594±0.0017ab,AB

0.9559±0.0021a,A

0.9617±0.0015a,AB

0.9629±0.0013a,AB

0.9635±0.0013a,B

LFKGM

0.9636±0.0012a,A

0.9644±0.0023b,AB

0.9667±0.0017ab,AB

0.9674±0.0012ab,AB

0.9675±0.0009a,B

SKKGM

0.9645±0.0013a,A

0.9656±0.0015b,A

0.9676±0.0011b,A

0.9681±0.0012b,A

0.9676±0.0012a,A

5.70±0.05a,A

5.65±0.08a,A

5.53±0.01a,A

5.62±0.06ac,A

5.66±0.04a,A

5.57±0.05ab,ABC

5.48±0.05b,AC

5.41±0.01b,A

5.67±0.01a,B

5.50±0.02bd,C

SKC

5.46±0.04b,AC

5.44±0.01b,A

5.52±0.01a,B

5.51±0.03b,BC

5.56±0.01ad,B

LFKGM

5.41±0.02b,A

5.51±0.02b,BC

5.58±0.03a,C

5.55±0.03bc,BC

5.47±0.02bc,AB

SKKGM

5.39±0.02b,A

5.49±0.01b,B

5.56±0.03a,B

5.50±0.01b,B

5.43±0.02c,A

FFC

AC

LFC

CE

pH

SC

NU

D

0.9634±0.0009ab,A

PT E

aw

MA

Moisture (%)

RI

Protein Content (%)

PT

Fat Content (%)

1

Values are expressed as mean ± SD, n = 3. FFC = full-fat Mozzarella cheese control; LFC = low-fat Mozzarella cheese control; SKC = skimmed Mozzarella cheese control; LFKGM = low-fat Mozzarella cheese with KGM; SKKGM = skimmed Mozzarella cheese with KGM. a-d means for cheese sample at the same storage time without common letters are significantly different (P < 0.05). A-C means for the same cheese sample stored for different days without common letters are significantly different (P < 0.05). 2

35

ACCEPTED MANUSCRIPT Table 2. Textural parameters of grated cheeses1 Storage Time (Day)

Sample2 0

7

14

21

28

FFC

3967.2±392.7a,AC

4056.3±335.7ac,AC

5765.8±277.6a,B

5296.7±380.3ab,AB

3181.4±155.8a,C

LFC

5158.8±591.9ab,AB

5530.3±180.9b,A

6010.2±257.7a,AB

4850.4±517.1abc,AB

6636.2±232.8b,B

SKC

6101.8±166.5b,A

4302.2±170.2ac,B

5404.5±89.2a,C

4581.1±221.1a,B

3503.9±216.2a,D

LFKGM

4856.0±430.9ab,AB

5157.2±442.8ab,AB

5535.3±272.8a,AB

5659.1±111.6b,A

6077.4±58.0b,B

SKKGM

5609.8±322.9b,A

3625.5±260.3c,B

3808.6±167.2b,B

3373.6±167.2b,B

2296.0±108.3c,C

5881.1±730.8ab,AB

3319.0±390.8a,C

PT

Firmness(g)

Work of Shear (g.s) 4169.7±402.2a,AC

4247.4±673.1ab,AC

6958.4±496.8a,B

LFC

4005.5±700.8a,AB

4078.9±174.4a,A

4723.0±114.9ac,B

3847.2±467.4abc,AB

5875.8±301.4b,C

SKC

3776.5±387.4a,AB

2300.3±73.8b,AC

3314.9±295.4b,AB

2897.1±124.3ac,B

2281.2±163.5a,C

LFKGM

4349.2±491.2a,A

4085.9±557.1ab,A

4254.7±535.8bc,A

5078.4±153.6b,A

5171.2±17.1b,A

SKKGM

3428.6±35.0a,A

2348.7±356.3b,B

2309.4±117.2b,B

2397.9±268.0c,B

1401.6±52.2c,C

FFC

-868.2±34.2a,A

-382.3±9.8a,B

-147.9±25.5a,C

-136.64±36.3a,C

-793.7±92.9a,A

LFC

-19.2±2.9b,A

-36.6±7.7b,A

-27.2±2.4b,A

-18.0±6.2b,A

-44.2±12.3bc,A

SKC

-0.83±0.13b,A

-0.11±0.01c,A

-0.18±0.09b,A

-1.56±0.62b,AB

-2.76±1.10b,B

LFKGM

-67.7±2.9c,A

-73.2±2.6d,A

-83.2±7.6c,A

-97.4±16.7a,AB

-158.2±47.1c,B

SKKGM

-2.01±0.94b,A

-1.91±0.78c,A

-3.16±0.89b,AB

-4.37±0.79b,AB

-6.07±0.47b,B

-181.5±37.8a,AB

-195.6±33.3a,AB

-287.5±47.9a,B

SC

NU

-85.4±7.3a,A

-175.8±18.3a,AB

LFC

-1.22±0.29b,A

-9.13±1.50bc,B

-10.2±2.3b,AB

-7.84±1.39b,B

-21.7±3.2b,C

SKC

1.34±0.43b,A

3.56±0.42b,B

1.35±0.13b,A

1.01±0.09b,A

0.80±0.02b,A

LFKGM

-16.4±2.6c,A

-19.8±1.3c,A

-24.9±4.4b,AC

-69.0±14.7c,B

-40.4±4.7b,C

SKKGM

0.93±0.03b,A

0.88±0.08bc,A

0.74±0.20b,AB

0.50±0.03b,BC

0.38±0.09b,C

PT E

FFC

CE

1

D

Work of Adhesion (g.s)

MA

Stickness (g)

RI

FFC

AC

Values are expressed as mean ± SD, n = 3. FFC = full-fat Mozzarella cheese control; LFC = low-fat Mozzarella cheese control; SKC = skimmed Mozzarella cheese control; LFKGM = low-fat Mozzarella cheese with KGM; SKKGM = skimmed Mozzarella cheese with KGM. a-c means for cheese sample at the same storage time without common letters are significantly different (P < 0.05). A-D means for the same cheese sample stored for different days without common letters are significantly different (P < 0.05). 2

36

ACCEPTED MANUSCRIPT Table 3. Textural parameters of melted cheeses1 Storage Time (Day)

Sample2 0

7

14

21

28

FFC

23.41±3.97a,AB

31.94±3.08a,A

17.92±2.92a,B

13.69±2.57a,B

18.70±2.30a,B

LFC

19.31±0.06a,A

66.85±14.33ab,A

40.57±8.88a,A

52.75±8.62bc,A

71.60±17.97ab,A

SKC

73.20±13.72a,ABC

78.07±7.90b,A

62.25±7.58ab,AB

39.27±7.11abc,BC

32.29±7.91ab,C

LFKGM

19.54±0.81a,A

40.01±3.05a,B

38.10±8.04a,AB

27.51±6.11ab,AB

31.97±4.16ab,AB

SKKGM

138.57±7.81b,A

76.42±13.01ab,B

76.11±10.36b,B

53.74±5.61c,B

37.35±1.98b,B

0.03±0.00a,B

0.01±0.00a,C

PT

Resistance (g)

FFC

0.11±0.02a,A

0.06±0.00a,A

0.04±0.03a,ABC

LFC

0.01±0.00b,A

0.35±0.08ab,B

0.20±0.01b,B

SKC

0.41±0.01c,A

0.23±0.03b,B

0.07±0.01a,C

LFKGM

0.07±0.02ab,A

0.18±0.03b,B

SKKGM

1.05±0.04d,A

0.17±0.02b,B

RI

Stretch quality (g/s)

0.27±0.05b,B

0.08±0.01b,C

0.10±0.01b,C

0.16±0.02bc,B

0.17±0.01c,B

0.03±0.01b,C

0.10±0.01ac,BC

0.08±0.01b,C

0.06±0.01b,C

SC

0.27±0.05bc,B

1

NU

Values are expressed as mean ± SD, n = 3. FFC = full-fat Mozzarella cheese control; LFC = low-fat Mozzarella cheese control; SKC = skimmed Mozzarella cheese control; LFKGM = low-fat Mozzarella cheese with KGM; SKKGM = skimmed Mozzarella cheese with KGM. a-d means for cheese sample at the same storage time without common letters are significantly different (P < 0.05). A-C means for the same cheese sample stored for different days without common letters are significantly different (P < 0.05).

AC

CE

PT E

D

MA

2

37

ACCEPTED MANUSCRIPT Figure 1. Color and browning (L*) and browning factor (BF) of cheeses during 28 days storage at 4 °C. FFC = full-fat Mozzarella cheese control; LFC = low-fat Mozzarella cheese control; SKC = skimmed Mozzarella cheese control; LFKGM = low-fat Mozzarella cheese with KGM; SKKGM = skimmed Mozzarella cheese with

PT

KGM. A is the color L* of cheeses; B is the browning L* of cheeses; C is the BF of

A-D

means for the same cheese sample stored for

SC

are significantly different (P < 0.05).

RI

cheeses. a-d means for cheese sample at the same storage time without common letters

different days without common letters are significantly different (P < 0.05).

NU

Figure 2. Color and browning (a*) of cheeses during 28 days storage at 4 °C. FFC =

MA

full-fat Mozzarella cheese control; LFC = low-fat Mozzarella cheese control; SKC = skimmed Mozzarella cheese control; LFKGM = low-fat Mozzarella cheese with

D

KGM; SKKGM = skimmed Mozzarella cheese with KGM. A is the color a* of a-e

PT E

cheeses; B is the browning a* of cheeses.

means for cheese sample at the same

storage time without common letters are significantly different (P < 0.05).

A-E

means

CE

for the same cheese sample stored for different days without common letters are

AC

significantly different (P < 0.05). Figure 3. Color and browning (b*) of cheeses during 28 days storage at 4 °C. FFC = full-fat Mozzarella cheese control; LFC = low-fat Mozzarella cheese control; SKC = skimmed Mozzarella cheese control; LFKGM = low-fat Mozzarella cheese with KGM; SKKGM = skimmed Mozzarella cheese with KGM. A is the color b* of cheeses; B is the browning b* of cheeses.

a-e

means for cheese sample at the same

storage time without common letters are significantly different (P < 0.05). 38

A-C

means

ACCEPTED MANUSCRIPT for the same cheese sample stored for different days without common letters are

AC

CE

PT E

D

MA

NU

SC

RI

PT

significantly different (P < 0.05).

39

ACCEPTED MANUSCRIPT

AC

CE

PT E

D

MA

NU

SC

RI

PT

Figure 1

40

ACCEPTED MANUSCRIPT

AC

CE

PT E

D

MA

NU

SC

RI

PT

Figure 2

41

ACCEPTED MANUSCRIPT

AC

CE

PT E

D

MA

NU

SC

RI

PT

Figure 3

42