Received: 11 December 2020  |  Revised: 18 July 2021  |  Accepted: 21 July 2021 DOI: 10.1002/fsn3.2507 O R I G I N A L R E S E A R C H Impact of type and level of stabilizers and fermentation period on the nutritional, microbiological, and sensory properties of short- set Yoghurt Chinazom Martina Eze1 | Kehinde Oludayo Aremu1 | Emmanuel Oladeji Alamu2  | Thomas Muoneme Okonkwo1 1Department of Food Science and Technology, University of Nigeria, Nsukka, Abstract Enugu State, Nigeria This study aimed to produce short set yoghurt with different stabilizers at different 2Food and Nutrition Sciences Laboratory, concentrations and determine the effects of the stabilizers and length of fermenta- International Institute of Tropical Agriculture (IITA), Southern Africa Hub, Chelstone, tion on the nutritional, microbiological, and sensory properties of short set yoghurt. Lusaka, Zambia Stabilized yoghurt samples were produced using 0%, 0.5%, and 1.0% concentrations Correspondence of carboxyl methylcellulose (CMC), corn starch, and gum acacia with different fer- Emmanuel Oladeji Alamu, International mentation periods (1–5 hr), respectively. Samples were analyzed for the proximate, Institute of Tropical Agriculture (IITA), 7th Floor, Grosvenor House, 125 High Street, physicochemical, microbial, and sensory properties using standard laboratory meth- Croydon CRO 9XP, UK. ods. Results showed that an increase in stabilizer concentration and fermentation Email: oalamu@cgiar.org time decreased the moisture content but increased the total solids, protein, fat, ash, sugars, pH level, and total titratable acidity. The viscosity of the yoghurt samples significantly (p < .05) increased with the addition of stabilizers (1.48 ± 0.03 cP to 275.57 ± 4.08 cP), with CMC having the highest increase (p < .05) and gum acacia the least. However, the lactic acid production reduced as the concentration of sta- bilizers increased but showed an increase with fermentation time. The total viable count (TVC) reduced significantly (p < .05) with an increase in the concentration of stabilizer and fermentation time. Hence, short set yoghurt samples containing CMC yielded highest protein (0.5%), fat (1.0%), and ash contents (1.0%). Yoghurt samples produced with a 1.0% concentration of gum acacia gave an optimum pH (0.5%), TTA, mouthfeel, appearance, flavor, and taste. In contrast, yoghurt produced with corn starch produced the most desirable overall acceptability, viscosity, total solids at 1.0%, and TVC (at 0.5%) concentration. K E Y W O R D S carboxyl methylcellulose, fermentation time, short set yoghurt, stabilizers, yoghurt properties This is an open access article under the terms of the Creat ive Common s Attri bution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. © 2021 The Authors. Food Science & Nutrition published by Wiley Periodicals LLC. Food Sci Nutr. 2021;00:1–16. www.foodscience-nutrition.com  |  1 2  |     EZE Et al. 1  | INTRODUC TION include vegetable or tree gums such a gum tragacanth and gum ar- abic (also known as gum acacia), agar, corn starch, gelatin, and pec- Yoghurt is one of the most popular fermented dairy products, and it tin. Cellulose compounds such as methylcellulose and CMC (sodium is a semisolid milk product and the best known of all fermented milk carboxyl methyl cellulose) are also used (Bakirci and Macit, 2017). products with increasing consumption worldwide (Shiby and Mishra, Much research has been carried out on yoghurt in terms of the final 2013). It is obtained by the souring of milk using a pure culture of product but not so much on the effect of these stabilizers locally a particular strain of Lactobacillus or a mixed culture of bulgaricus for yoghurt production during fermentation and on the nutritional and Streptococcus thermophilus and Lactobacillus bulgaricus in a 1:1 composition of the final product. Stabilizers used in yoghurt pro- ratio (Sansawal et al., 2017). It can be manufactured from fresh an- duction are many and varied. However, there is little information on imal liquid cow milk. In recent times, powdered cow milk is being how some of the stabilizers locally used in Nigeria influence the fer- used and vegetable milk (Soy milk) as a major base material (Obiora mentation rate of yoghurt and the nutritive value for which yoghurt et al., 2020). Lactic acid and the other compounds formed during is consumed. From previous research, it is noteworthy to say that the fermentation of milk make yoghurt a food product that is acidic fermentation increases the vitamin content of products, especially and creamy, appreciated for its taste and nutritional qualities, nota- some B-c omplex vitamins, due to microbial activities during fermen- bly for its calcium content (Widayat et al., 2020). Yoghurt is thus a tation, where synthesis and breakdown of substances occur (Nkhata very convenient food as compared to very fragile milk. Due to the et al., 2018). Yoghurt starter cultures utilize some vitamins present health benefits and taste, it constitutes an appreciable proportion in milk during fermentation for their growth. However, this incre- of total daily food consumption or even just as a refreshing beverage ment depends on the inoculation rate, the strain of yoghurt starter in several countries (Khan et al., 2011). It is regarded as a nutrition- cultures, and the fermentation conditions (Sharma et al., 2020). ally balanced food, containing almost all the nutrients present in milk Stabilizers or hydrophilic colloids bind water, prevent separation of and a more assimilable form (Olugbuyiro & Oseh, 2011). Yoghurt various ingredients, increase the viscosity, and inhibit the formation is a source of highly nutritive protein, energy from added cane of large crystals, which are attributes for consumer acceptability. sugar, milk fat and unfermented lactose, and vitamins (Ihekoronye It is, therefore, necessary to rebuild yoghurt with stabilizers and & Ngoddy, 1985). It is more nutritive than milk in terms of vitamin thickeners at such concentrations that will give the desired body to content, digestibility, and as a source of calcium and phosphorus the final product. This goal will be achieved by optimum selection of (Foissy, 1983). It is believed that yoghurt has valuable “therapeutic stabilizers with protective colloid properties, by assessing how the properties” and helps in curing gastrointestinal disorders (Bhattarai activities of the fermenting organisms will be enhanced or inhibited and Das, 2016). It also prevents and controls diarrhea, can modulate by the hydrocolloids used concerning vitamin synthesis by evaluat- the inflammatory response produced by carcinogens, and helps in ing the chemical, microbiological, nutritional, and sensory proper- reducing the inflammatory response through an increase in apopto- ties of yoghurt produced under controlled incubation fermented at sis. Yoghurt is a smooth viscous smooth gel with a distinct taste of 42℃ for 5 hr. The objectives of this work were to produce short sharp acid and green apple flavor (Chen et al., 2017). Some yoghurts set yoghurt with different stabilizers at different concentrations and have a heavy consistency like custard or milk pudding, whereas oth- determine the effects of these stabilizers and length of fermentation ers are purposely soft boiled and practically drinkable (Weerathilake on the nutritive value of short set yoghurt. et al., 2014). Firmness and smoothness are two of yoghurt's most important essential textural characteristics. The type of culture used is also an important factor in determining the microstructure and 2  | MATERIAL S AND METHODS the textural properties of yoghurt (Lee & Lucey, 2010). Yoghurt is classified primarily according to its chemical composition (full- fat, 2.1 | Raw materials reduced- fat, and low- fat), manufacturing method (set and stirred yoghurt), flavor type, and postincubation process. Yoghurt is made The raw materials used in producing the short set yoghurt sam- using the method of production before incubation, cooling, and ples were purchased at Ogige primary market in Nsukka Local straining, and a firm jelly-l ike texture characterizes it. Government Area of Enugu State, Nigeria. These include milk (peak), On the other hand, short set yoghurt is a type of yoghurt pro- granulated sugar, starter culture (Yoghurmet), and stabilizers (car- duced under controlled incubation and fermentation at 42℃ for 0 to boxyl methylcellulose, corn starch, and gum acacia). 5 hr, improving textural properties and preventing wheying- off de- fects. In comparison, long set yoghurt is a type of yoghurt produced and fermented at 30℃ for 0 to 16 hr (Abayneh, 2020). 2.2 | Sample preparation Stabilizers and thickeners are essential in several manufactured products and dairy products such as chocolate dressing, milk drinks, 2.2.1 | Preparation of short set Yoghurt mix ice cream, and yoghurt. These substances prevent the separation of various ingredients, increase the viscosity, and inhibit the forma- Short set yoghurt was produced according to the method of Lee and tion of large crystals. Substances used as stabilizers and thickeners Lucey (2010) with slight modification. Dried milk sample (250 g) and EZE Et al.      |  3 granulated sugar (10 g) were weighed and made up to two liters (2 L) with clean water to produce an equivalence of fresh milk. The mix- W2 −W3 %Moisture Content = × 100 (1) ture was divided into six equal parts each for the six concentrations W2 −W1 of the three stabilizers (CMC, corn starch, and gum acacia) added into the yoghurt mix. The stabilizers were added at a concentration where W1 = Initial weight of the empty crucible; W2 = weight of cruci- of 0.5% and 1%, respectively. A control sample with a 0% stabilizer ble + weight of the sample before drying; W3 = weight of dish +weight was also produced. Each yoghurt mix was pasteurized at 80℃ for of the sample after drying. 60 s, cooled to 43℃, inoculated with 2% starter culture (Yoghurmet) consisting of Streptococcus thermophilus and Lactobacillus bulgaricus, and packed into plastic containers of two liters (2 L) of capacity and 2.4.2 | Determination of ash content allowed to ferment for 5 hr at 45℃ from where samples were with- drawn for analyses at intervals of 1 hr. The ash content of the freshly prepared yoghurt samples was deter- mined according to the standards of AOAC (2010). A preheated and cooled crucible was weighed (W1), and a 2 g sample was weighed 2.3 | Production of Stabilized Yoghurt Samples into the preheated cooled crucible (W2). The sample was charred to remove hydrogen and oxygen and facilitate the ashing procedure on Each yoghurt mix with its stabilizer proportion was pasteurized at a Bunsen flame inside a fume cupboard. The charred sample in the 80℃ for 60 s to hasten hydration and solubilization of the solid in- crucible was transferred into a preheated muffle furnace (Carbolite gredients and, more importantly, destroy organisms present in the AAF— 1100) at 550℃ for 2 hr until a white or light gray ash was ob- mix (Figure 1). tained (W3). It was cooled in a desiccator, weighed, and documented. This was followed by homogenization of the yoghurt sample The ash content of the samples was calculated using Equation (2) using stainless steel, Mariam@ 6 in 1 blender, German Model No: M2 (1800W power), which helped to homogenize all the ingredients, es- W3 −W1 %Ash content = × 100 (2) pecially the stabilizers and also helped to break down fat globules in W2 −W1 milk into smaller more consistently dispersed particles, which gave a smoother and creamier product. Cooling to inoculation temperature where W1 = weight of the empty crucible; W2 = weight of cruci- of 40– 43℃ allowed to achieve the cooling effect suitable for the ble + sample before ashing; W3 = weight of crucible + sample after culture. Inoculation with starter culture was done with 2% starter ashing. culture (Streptococcus thermophilus and Lactobacillus bulgaricus). After inoculation with a starter culture, each sample was divided into two portions of one liter each. Each one liter of the sample was 2.4.3 | Determination of fat content dispensed into ten incubation plastic bottles of 100 ml. The first set of 1 liter from each sample was used to produce a short set yoghurt The fat content of the yoghurt samples was determined using the by incubating at 40–4 3℃ for 5 hr. Soon after homogenizing with standard AOAC (2010) method. A Soxhlet extractor with a reflux a starter culture (Yoghurmet), samples were withdrawn from each condenser and a 500-m l round bottom flask was fixed. The yoghurt portion of 1 liter for analyses for the zero hour of incubation. After sample (2 g) was weighed into a labeled thimble, and petroleum that, samples were withdrawn at intervals of 1 hr within the fermen- ether (300 ml) was filled into the round bottom flask. The extrac- tation period of 5 hr for short set yoghurt. tion thimble was sealed with cotton wool. The Soxhlet apparatus, after assembling, was allowed to reflux for about 6 hr. The thimble was removed with care, and the petroleum ether drained into a con- 2.4 | Proximate analyses tainer for reuse. When the flask was free of ether, it was removed and dried at 70℃ for 1 hr in an oven (GALLEMKAMP Oven Model: 2.4.1 | Determination of moisture content REX—C 900). It was cooled in desiccators and then weighed. The fat content of the samples was calculated using Equation (3). The moisture content of the samples was determined according to the standard method of the Association of Official Analytical Weight of fat% fat content = × 100 (3) Weight of the sample Chemists (AOAC, 2010). The crucibles were washed thoroughly and dried in the oven (GALLEMKAMP Oven Model: REX—C 900) at 100℃ for 1 hr. The hot- dried crucibles were cooled in a desiccator 2.4.4 | Determination of crude fiber (Laboware Plastic Vacuum, Desiccator, Capacity: 150 mm) and then noted down (W1). The sample (2 g) was weighed (CAMRY Electronic The crude fiber was determined using the method described by Weighing Balance) into the crucible (W2) and dried at 70℃ until a AOAC (2010). About 5 ml of the sample (w3) was digested with constant weight was obtained (W3). The moisture content of the 200 ml of 0.22 N H2SO4; it was filtered and washed severally samples was calculated as given in Equation (1). and transferred into another conical flask. The mixture was then 4  |     EZE Et al. F I G U R E 1   The flow chart for the production of stabilized short set yoghurt. Source: Adapted (Early, 1998) dissolved in 200 ml of 1.25% NaOH solution, boiled for 30 min, cold 2.5 | Digestion of the sample filtered, and washed with boiling water. The residue was dried in an oven (GALLEMKAMP Oven Model: REX— C900) at 105℃ for 2 hr, The sample (2 g) was weighed into a Kjeldahl digestion flask followed cooled in a desiccator, and weighed. It was incinerated at 550℃ for by the addition of anhydrous barium sulfate (BaSO4) and hydrated 2 hr in a muffle furnace (Carbolite AAF—1 ,100), cooled again in a copper (II) tetraoxosulfate (VI) as a catalyst. About 25 ml of concen- desiccator, and weighed. The percentage of crude fiber was calcu- trated tetraoxosulfate (VI) acid (H2SO4) was added with a few boil- lated, as shown in Equation 4. ing chips. The flask with its content was heated in a fume chamber until a clear solution was obtained. The solution was cooled to room W2 −W1 (4) temperature, after which it was transferred into a 250- ml volumetric %Crude Fiber = × 100 W3 flask and made up to the level with distilled water. where W1 = weight of the sample before incineration; W2 = weight of the sample after incineration; W3 = weight of the original sample. 2.6 | Distillation The distillation unit was cleaned, and the apparatus was set up. A 2.4.5 | Determination of protein content 100- ml conical flask (receiving flask) containing 5 ml of 2% Boric acid (H3BO4) was placed under the condenser with the addition The protein content of the samples was determined accord- of 2 drops of methyl red indicator. A digest of 5 ml was pipetted ing to the standard procedure of AOAC (2010) using the Kjeldahl into the apparatus through the small funnel, washed down dis- method. tilled water, and followed by the addition of 5 ml of 60% sodium EZE Et al.      |  5 hydroxide (NaOH) solution. The digestion flask was heated until 100 ml of distillate (ammonium sulfate) was collected in the receiv- Qty of NaOH (ml)Titratable acidity (%) = × 0.009 × 100 (7) Qty of Sample (g) ing flask. 2.8.3 | Determination of total solids 2.7 | Titration The total solid content of the freshly prepared yoghurt with dif- The solution in the receiving flask was titrated with about 0.04 M ferent concentrations of stabilizers was determined using AOAC hydrochloric acid (HCl) to get a pink color. The same procedure was (2010). The sample (5 g) was dried to a constant weight in a hot air carried out on the blank. oven (Gallenkamp) at 130℃. The total solid content was obtained as a percentage (%) of total solids. Vs − Vb × Nacid %Nitrogen = × 0.0401 × 100 (5) W Weight of Dried Solid TotalSolid (%) = × 100 (8) Weight of Sample where Vs = volume (ml) of acid required to titrate the sample; Vb = vol- ume (ml) of acid required to titrate blank; Nacid = Normality of acid 2.8.4 | Determination of apparent viscosity (0.1 N); W = weight of the sample in gram. The viscosity of short set yoghurt samples was determined as de- scribed by Ayernor and Ocloo (2007) with the use of Universal 2.7.1 | Determination of Carbohydrate Torsion Viscometer (Gallenkamp Technico Compenstat, Gallenkamp Co. Ltd, England). 10% (w/v) of each of the yoghurt samples was Carbohydrate (by difference) was determined using method de- used. The determination was done using an 11/16- inch pendulum scribed by AOAC (2010). It was calculated by getting the sum of the (no. 4) of standard wire gauge. The values obtained were converted other proximate parameters and subtracting it from 100 as nitrogen- to centipoises (cP) and recorded. free extract (NFE) as follows: %Carbohydrate (NFE) = 100 − (M + P + F + A) (6) 2.9 | Microbial analyses where M = Moisture content; P = Protein; F = Fat; A = Ash. Microbiological analyses were carried out as described by Prescott et al., (2005). A serial dilution of the sample was done. The sample was placed at ambient temperature. The total viable count was performed 2.8 | Physicochemical analyses at intervals of 1 hr within the fermentation period of 5 hr for short set yoghurt and 4- hr interval fermentation period of 24 hr for a long set 2.8.1 | Determination of pH yoghurt. Total viable count (TVC) and mold count were determined by the pour plate method on nutrient agar and Sabouraud dextrose A pocket- sized pH meter (Hanna Instrument, Woonsocket, China, agar (SDA), respectively, as described by Prescott et al., (2005). R10895) was used to determine the pH of the samples, according to AOAC (2010) method. Approximately 10% (w/v) of each of the yoghurt samples was mixed with CO2-f ree distilled water, and the 2.10 | Sensory evaluation mixture was shaken vigorously. The pH meter was calibrated using a buffer solution of 4.0 and 7.0. After 10 min of calibration, the pH According to Ihekoronye and Ngoddy (1985), the sensory evalua- meter electrode was dipped into a prepared suspension of the sam- tion was carried out using a 20- man semitrained panelist. The pan- ples for the pH measurement. elists were instructed to indicate their preference for the samples. According to Iwe (2002), a nine- point Hedonic scale, where 9 was the highest score and 1 was the lowest score for each characteristic 2.8.2 | Determination of titratable acidity such as color, flavor, mouthfeel, and overall acceptability, was used. The total titratable acidity was determined using the AOAC (2010) method. The sample (5 ml) at 25℃ was measured into a flask and di- 2.10.1 | Data analyses and experimental design luted twice its volume with distilled water. Phenolphthalein indicator (2 ml) was added to each yoghurt sample and titrated with 0.1 mol/L The study was designed in a split- plot design using Design Expert® NaOH to the first permanent pink color. The acidity was reported version 11.1.2.0. The analysis of variance (ANOVA) was conducted as the percentage of lactic acid by weight, as shown in Equation (7). on the obtained data using GEN STAT RELEASE 10.3 DE. The least 6  |     EZE Et al. significant difference (LSD) was used to compare the treatment (Table 2). The fat content ranged from 2.65 ± 0.01% to 3.42 ± 0.05%. means. Statistical significance was accepted at p < .05 (Steel & Short set yoghurt samples stabilized with corn starch recorded the Torrie, 1980). highest values (3.11%– 3.42%), while those containing gum acacia recorded the least fat content (2.65%– 3.13%). This was attribut- able to the residual oil in corn starch. The effect of concentrations 3  | RESULTS AND DISCUSSION of the stabilizers on the fat content was significant (p < .05). The result obtained agreed with the findings of Tamime and Robinson 3.1 | Effects of stabilizers and fermentation time on (2007), who reported that fat contents ranging from 2.60% to 3.24% the proximate composition of short set Yoghurt are for yoghurt types regarded as low fat and should contain less than 3.5% fat, while full-f at yoghurt contains more than 3.5%. The Table 2 shows the effect of stabilizers and length of fermentation on short set yoghurt samples produced could, therefore, be categorized the proximate composition of the short set yoghurt. as low-f at yoghurt. The fermentation time had a significant effect (p < .05) on the fat content of the stabilized short set yoghurt sam- ples. An increase in the fat content was observed with an increase 3.1.1 | Moisture content in fermentation time, and the values ranged from 2.65 ± 0.01% to 3.42 ± 0.05%. Yadav et al., (2007) studied the effect of milk fat con- The decrease in the moisture contents (from 88.54 ± 0.02% to tent on the acid development during fermentation and rheological 82.26 ± 0.03% ) of all short set yoghurt samples formulated with properties of plain yoghurt. The authors indicated increasing fat carboxyl methyl cellulose (CMC), corn starch, and gum acacia were content as fermentation proceeded. The interaction effect of sta- statistically significant (p < .05) as compared to the control. bilizers and concentrations of the stabilized short set yoghurt's fat The decrease in moisture content from the start could be at- content indicated that the stabilizers' behaviors varied at different tributed to the fact that the fermentation increases the proportion concentrations. Therefore, the differences between stabilizers were of dry matter in the food. The concentration of vitamins, minerals, magnified by concentrations used. At 0% concentrations, the sta- and protein appears to increase when measured on a dry weight bilizers had similar fat contents. However, at a 0.5% concentration, basis (Olasupo & Okorie, 2019). All CMC-s tabilized short set yoghurt corn starch (C1) had a higher fat content than CMC, while gum acacia samples had lower moisture content at 0.5% and 1.0% concentration had the least fat content. Also, at 1.0% concentration, corn starch compared to samples stabilized with gum acacia, which had higher (C2) had a higher concentration than CMC, with gum acacia being the moisture content at similar concentrations. The lower moisture con- least, but the differences were wider at 1.0% concentration than at tent could be attributed to the ability of CMC to increase the viscosity 0.5% concentration. This agrees with Angor, 2016, who stated that of the sample, which makes it exhibit functional properties of thick- CMC used as an edible coating film reduced fat absorption and im- ening, stabilization in agreement with Davidson (1980). On the other proved moisture retention in starchy products and poultry products. hand, gum acacia has higher water solubility (up to 50% w/v) and rel- It could also be attributable to the fact that corn starch has more atively low viscosity than other exudate gums (Dragnet, 2000). The calories than CMC and gum acacia, which is more of edible fiber, interaction effect of the different stabilizers and their concentrations thus the low- fat content. on the moisture content of short set yoghurt samples revealed signif- icant differences (p < .05). This is indicative of the fact that the be- havior of the stabilizers was not the same at different concentrations. 3.1.3 | Protein content The interaction effect showed that at 0% concentration, the different stabilizers behaved the same way, having similar proportions of mois- The different stabilizers had a significant effect (p < .05) on the pro- ture. However, a higher concentration of different stabilizers magni- tein content of the short set yoghurt samples. The protein values fied the differences between the stabilizers in the moisture content ranged from 3.06 ± 0.02% to 3.71 ± 0.02% (Table 2). The effect of of short set yoghurt. At 0.5% concentration, the yoghurt sample CMC, corn starch, and gum acacia on the protein content revealed containing gum acacia had higher moisture content followed by corn that CMC gave the highest protein content (3.25 to 3.71%) followed starch, while yoghurt containing CMC had the least moisture content. by gum acacia (3.28 to 3.62%), with corn starch having the lowest At 1.0% concentration, a similar trend was observed, but the differ- protein value (3.06 to 3.43%). This was in contrast with the findings ences were high. This is attributable to the high water holding capac- of Alakali et al., (2007). Gum acacia gave high protein content close ity of the stabilizer (CMC), which exhibited a higher water retention to that of CMC. This could be attributed to the fact that gum acacia ability compared to the other stabilizers (Angor, 2016). has a covalent association with protein moieties rich in hydroxypro- line, serine, and proline (Dragnet, 2000). Concentrations of the dif- ferent stabilizers also had a significant effect (p < .05) on the protein 3.1.2 | Fat content content of the stabilized short set yoghurt samples. The protein content significantly (p < .05) decreased with an in- The stabilizers’ effect on the fat content of short set yoghurt sam- crease in the concentration of the stabilizers. This result obtained ples indicated that significant (p < .05) differences were observed corroborated with the study carried out by Alakali et al. (2007), which EZE Et al.      |  7 TA B L E 1   Ingredient mixers for the production of short set yoghurt samples Stabilizer concentrations Starter cultureb  Sugar Sample code Stabilizer g (%) Liquid milka  (ml) (g) (g) A Control (No stabilizer) 0.00 2000 10 10 B1 CMC 10 g (0.5%) 2000 10 10 B2 20 g (1.0%) 2000 10 10 C1 Corn Starch 10 g (0.5%) 2000 10 10 C2 20 g (1.0%) 2000 10 10 D1 Gum Acacia 10 g (0.5%) 2000 10 10 D2 20 g (1.0%) 2000 10 10 aLiquid milk produced by dissolving 250 g powdered milk + 10 g of sugar and made up to 2 L with water. bStarter culture = Yoghurmet. reported that a higher concentration of stabilizers reduces the nutri- (p < .05) for the short set yoghurt samples. The ash content was tional quality of yoghurt samples by causing a reduction in the pro- highest at 1.0% concentration and lowest at 0% concentration due tein content of yoghurt due to the dilution effect. Protein contents to the higher quantity of stabilizer at 1.0% concentrations. It was significantly (p < .05) increased with an increase in fermentation observed that percent ash content generally increased with an in- time. This trend could be traced to the concentration of proteins in crease in fermentation time. This increase was found to be highly the yoghurt samples due to moisture loss, which caused an increase significant (p < .05) due to the concentration effect resulting from in other components. The range of values obtained was lower than moisture loss. There was increase in ash content from 0.34 ± 0.02% that reported by Bibiana et al. (2014), who found out that the protein to 0.79 ± 0.01% as fermentation time progressed (Table 1). The val- contents of other brands of yoghurt sold in Owerri, Imo state, were ues obtained could be compared with the range given by Mbaeyi within the range of 3.76% to 5.08%. The interaction effect between and Awaziem (2007), who reported yoghurt to contain ash from the different stabilizers and their concentrations was found to be 0.49% to 0.98%. The interaction effects between the three differ- significant (p < .05) as well. This indicated that the effects of the ent stabilizers and their concentrations were found to be significant stabilizers were not the same at different concentrations. It was ob- (p < .05) (Table 2). This was in contrast with the reported work of served that the 0% concentration of all the stabilizers had similar Alakali et al., (2007). This significant interaction suggested that effects on the protein contents since the absence of the stabilizers the differences in ash content due to stabilizers varied at different did not create a dilution effect. At 0.5%, short set yoghurt samples concentrations. containing CMC had higher protein content than those stabilized with corn starch. Also, at 1.0% concentration, a similar trend was observed. However, the differences in protein content were wider 3.2 | Effects of stabilizers and fermentation and smaller at 1% concentration due to more significant moisture time on the physicochemical properties of short loss and increased dilution effect at 1.0% concentration. Fermented set Yoghurt milk products are good high- quality protein sources with high bio- logical value (Canadian Dairy Commission, 2007). Therefore, it was Tables 3 and 4 show the effect of stabilizers (type and concentration) observed that the dilution effect caused differences in the type and and length of fermentation on the physicochemical properties of the quantity of stabilizers used. It was observed that gum acacia gave short set yoghurt samples. high protein content at 1.0% concentration due to higher reaction rates; a higher quantity of protein was observed at the higher tem- perature of incubation for short set yoghurt. 3.2.1 | pH The short set yoghurt samples containing 0%, 0.5%, and 1.0% of 3.1.4 | Ash contents CMC, corn starch, and gum acacia indicated that the differences between effects of stabilizers were not significant between CMC The addition of CMC, corn starch, and gum acacia, each at levels of and corn starch but differed significantly (p < .05) from gum acacia. 0%, 0.5%, and 1.0% concentrations, caused significant differences Although titratable acidity production in yoghurt containing corn (p < .05) between the ash contents of short set yoghurt samples to starch was higher than yoghurt containing CMC (Table 4), the pH which different stabilizers were added. CMC recorded the highest of the yoghurt samples containing CMC and corn starch was simi- percent ash content, presumably due to the high sodium content lar, suggesting that the pH of yoghurt containing corn starch had of CMC (Benyounes, 2012). The effect of different concentrations a higher buffering capacity. This was unlike the yoghurt containing (0%, 0.5%, and 1.0%) of the stabilizers was found to be significant gum acacia, which appeared to be less buffered. However, CMC gave 8  |     EZE Et al. TA B L E 2   Effect of fermentation period (hours) on the proximate properties of short set Yoghurt Fermentation period (hr) Proximate parameters Sample 0 1 2 3 4 5 Moisture A 88.54 ± 0.02a 87.59 ± 0.33b 86.81 ± 0.53b 84.33 ± 1.13c 82.63 ± 0.02d 82.46 ± 0.03d B1 84.77 ± 0.02 a 84.65 ± 0.02b 84.29 ± 0.02c 84.12 ± 0.03d 83.78 ± 0.02e 83.40 ± 0.03f B2 84.41 ± 0.02 a 84.29 ± 0.02b 84.09 ± 0.02c 83.85 ± 0.02d 83.26 ± 0.03e 82.78 ± 0.03f C1 85.63 ± 0.02 a 85.20 ± 0.02b 85.10 ± 0.03c 84.85 ± 0.02d 84.59 ± 0.02e 83.62 ± 0.03f C2 85.43 ± 0.02 a 84.75 ± 0.02b 84.21 ± 0.02c 84.15 ± 0.03d 82.79 ± 0.02e 82.26 ± 0.03f D1 86.97 ± 0.02 a 86.44 ± 0.02b 85.76 ± 0.03c 85.51 ± 0.03d 84.73 ± 0.03e 84.27 ± 0.03f D2 86.43 ± 0.02 a 86.41 ± 0.03a 86.33 ± 0.03b 85.24 ± 0.03 84.59 ± 0.09d 84.23 ± 0.03e Fat A 3.12 ± 0.04c 3.13 ± 0.02c 3.15 ± 0.02c 3.25 ± 0.01b 3.28 ± 0.02ab 3.33 ± 0.05a B1 2.93 ± 0.01 c 2.92 ± 0.04c 2.94 ± 0.03c 3.07 ± 0.05b 3.11 ± 0.02b 3.23 ± 0.02a B de2 2.98 ± 0.02 2.97 ± 0.02 de 3.04 ± 0.06d 3.16 ± 0.04c 3.26 ± 0.02b 3.35 ± 0.05a C1 3.11 ± 0.02 b 3.12 ± 0.02b 3.15 ± 0.02b 3.21 ± 0.04a 3.25 ± 0.02a 3.25 ± 0.01a C2 3.18 ± 0.02 d 3.20 ± 0.01 cd 3.24 ± 0.04 cd 3.25 ± 0.05c 3.34 ± 0.02b 3.42 ± 0.05a D1 2.67 ± 0.02 cd 2.65 ± 0.01 e 2.65 ± 0.02d 2.70 ± 0.01bc 2.73 ± 0.02b 2.85 ± 0.06a D2 2.70 ± 0.06 c 2.71 ± 0.02c 2.70 ± 0.01c 2.85 ± 0.02b 2.92 ± 0.11b 3.13 ± 0.01a Protein A 3.37 ± 0.03d 3.36 ± 0.04d 3.38 ± 0.02 cd 3.43 ± 0.02c 3.59 ± 0.02b 3.67 ± 0.06a B1 3.35 ± 0.04 c 3.32 ± 0.02bc 3.33 ± 0.03b 3.64 ± 0.03a 3.68 ± 0.01b 3.71 ± 0.02a B2 3.31 ± 0.02 c 3.28 ± 0.04c 3.34 ± 0.04c 3.43 ± 0.02b 3.46 ± 0.01a 3.53 ± 0.05a C c c b b a1 3.13 ± 0.03 3.18 ± 0.02 3.26 ± 0.05 3.31 ± 0.02 3.43 ± 0.04 3.47 ± 0.03 a C2 3.06 ± 0.02 d 3.12 ± 0.04c 3.12 ± 0.03c 3.28 ± 0.02b 3.34 ± 0.04a 3.34 ± 0.03a D1 3.33 ± 0.04 d 3.32 ± 0.03d 3.33 ± 0.04d 3.43 ± 0.02b 3.56 ± 0.03b 3.62 ± 0.04a D2 3.29 ± 0.02 3.28 ± 0.05 c 3.29 ± 0.02c 3.34 ± 0.05bc 3.39 ± 0.03ab 3.44 ± 0 0.03a Ash A 0.34 ± 0.02e 0.54 ± 0.02d 0.56 ± 0.02d 0.61 ± 0.01c 0.65 ± 0.02b 0.69 ± 0.02a B1 0.65 ± 0.05 bc 0.58 ± 0.03c 0.63 ± 0.04bc 0.70 ± 0.01ab 0.75 ± 0.05a 0.76 ± 0.03a B2 0.61 ± 0.02 b 0.62 ± 0.03b 0.65 ± 0.04b 0.76 ± 0.04a 0.78 ± 0.02a 0.79 ± 0.01a C1 0.56 ± 0.04 b 0.57 ± 0.05b 0.57 ± 0.02a 0.66 ± 0.03a 0.69 ± 0.01a 0.71 ± 0.04a C2 0.56 ± 0.03 a 0.59 ± 0.08a 0.59 ± 0.02a 0.68 ± 0.01a 0.66 ± 0.01a 0.69 ± 0.02a D1 0.47 ± 0.05 c 0.53 ± 0.03bc 0.54 ± 0.03b 0.63 ± 0.01a 0.68 ± 0.01a 0.66 ± 0.04a D2 0.50 ± 0.02 b 0.52 ± 0.04b 0.52 ± 0.04b 0.64 ± 0.05a 0.69 ± 0.01a 0.69 ± 0.00a Note: Mean ± SD of triplicate readings. Values with a different superscript in the same column are significantly different. Keys: Fermentation period: 0–5 hr; Sample A = Short set yoghurt without any stabilizer; Sample B1 = Short set yoghurt with 0.5% CMC; Sample B2 = Short set yoghurt with 1.0% CMC; Sample C1 = Short set yoghurt with 0.5% Corn starch; Sample C2 = Short set yoghurt with 1.0% Corn starch; Sample D1 = Short set yoghurt with 0.5% Gum acacia; Sample D2 = Short set yoghurt with 1.0% Gum acacia. the highest pH value when compared to the other stabilizers. This 1.0% concentrations. The highest pH was produced at 0.5% concen- could be attributed to the fact that CMC is a stabilizer that is more tration by CMC and corn starch. The lowest pH was produced at soluble in alkali conditions and insoluble in acidic conditions and 0.5% concentration by gum acacia. This low pH value of gum acacia has an optimum pH range of 6.0–8 .5 (1 in 100 solutions) (Dragnet, could be as a result that gum acacia as a stabilizer has an optimum 2000). It could also be attributable to a lower level of fermentation pH range of 4.5 (William and Phillips, 2009), which lowered the pH of CMC. It gave a higher pH value than corn starch and gum aca- of the yoghurt prepared with this stabilizer. It could also be attrib- cia, which was somehow more fermented. The differences caused uted to the stability of gum acacia in acid conditions and high solubil- by the concentration of the stabilizers on the pH value of short set ity (Eqbal & Abdullah, 2013) or that gum acacia was also fermented yoghurts were statistically significant (p < .05). This means that the along with lactose and possesses lower buffering capacity. The de- pH generally decreased with an increase in the concentration of the crease in pH as the concentration rises may be attributable to the stabilizers, but the nature of the decrease was significantly (p < .05) continued fermentation of the lactic acid bacteria and the acidity ef- different for different stabilizers. The pH of the yoghurt with no fect of the added stabilizers (Ibrahim & Khalifa, 2015). Fermentation stabilizer 0% (control) differed significantly from those of 0.5% and time had a significant (p < .05) effect on the pH value obtained for EZE Et al.      |  9 TA B L E 3   Physicochemical properties of Short Set Yoghurt Samples at different fermentation period Physicochemical parameters Sample 0 1 2 3 4 5 pH A 6.19 ± 0.03a 6.18 ± 0.03a 6.18 ± 0.01a 5.09 ± 0.03b 4.52 ± 0.02c 4.18 ± 0.02d B1 6.27 ± 0.01 a 6.26 ± 0.01a 6.25 ± 0.02a 5.16 ± 0.04b 4.85 ± 0.04c 4.15 ± 0.04d B2 6.25 ± 0.02 a 6.25 ± 0.02a 6.23 ± 0.02a 5.23 ± 0.01b 4.97 ± 0.03c 4.20 ± 0.01d C1 6.18 ± 0.01 a 6.17 ± 0.01a 6.17 ± 0.03a 5.06 ± 0.05b 4.84 ± 0.04c 4.45 ± 0.03d C2 6.17 ± 0.03 a 6.16 ± 0.02a 6.13 ± 0.03a 5.09 ± 0.02b 4.93 ± 0.03c 4.45 ± 0.03d D1 5.99 ± 0.04 a 6.96 ± 0.02a 5.84 ± 0.02b 5.76 ± 0.04c 4.96 ± 0.02d 4.10 ± 0.03e D2 5.89 ± 0.02 d 5.87 ± 0.05ab 5.84 ± 0.02bc 5.81 ± 0.02c 4.49 ± 0.02d 4.14 ± 0.04e TTA A 0.29 ± 0.01d 0.30 ± 0.01 cd 0.31 ± 0.00d 0.63 ± 0.00b 0.75 ± 0.00a 0.75 ± 0.00a B e1 0.29 ± 0.01 0.30 ± 0.00 de 0.31 ± 0.00d 0.52 ± 0.00c 0.55 ± 0.02b 0.69 ± 0.00a B2 0.48 ± 0.00 e 0.48 ± 0.00e 0.54 ± 0.00d 0.56 ± 0.00c 0.59 ± 0.00b 0.69 ± 0.00a C1 0.36 ± 0.00 d 0.39 ± 0.00c 0.39 ± 0.00c 0.59 ± 0.00b 0.60 ± 0.01b 0.70 ± 0.04a C2 0.28 ± 0.01 f 0.30 ± 0.01e 0.39 ± 0.00d 0.52 ± 0.00c 0.59 ± 0.00b 0.70 ± 0.01 a D1 0.55 ± 0.00 e 0.57 ± 0.03e 0.61 ± 0.00d 0.65 ± 0.00c 0.75 ± 0.00b 0.84 ± 0.00a D2 0.34 ± 0.00 e 0.35 ± 0.00e 0.36 ± 0.00d 0.69 ± 0.00c 0.79 ± 0.00b 0.89 ± 0.02a Viscosity A 1.48 ± 0.03e 8.17 ± 0.05d 8.87 ± 0.32d 103.92 ± 1.52c 197.89 ± 1.06b 119.62 ± 0.70a B e e d c1 58.31 ± 1.79 96.66 ± 1.36 98.74 ± 1.43 154.30 ± 3.21 166.27 ± 2.97 b 264.00 ± 2.39a B2 97.74 ± 0.49 e 105.06 ± 0.84d 106.41 ± 0.90d 161.76 ± 1.04c 169.88 ± 1.58b 275.57 ± 4.08a C1 2.45 ± 0.04 e 784.00 ± 0.01d 8.45 ± 0.05d 120.62 ± 0.48c 128.45 ± 0.78b 239.51 ± 1.81a C2 3.02 ± 0.04 d 8.10 ± 0.03c 8.55 ± 0.03c 130.60 ± 0.77b 130.65 ± 0.82b 248.47 ± 1.26a D1 1.49 ± 0.03 e 4.85 ± 0.25d 5.99 ± 0.13d 90.82 ± 1.42c 106.26 ± 1.54b 120.68±429a D2 2.5 ± 0.03 e 5.19 ± 0.02d 6.18 ± 0.03d 101.36 ± 1.28c 111.00 ± 1.20b 126.49 ± 2.93a Total Solid A 11.64 ± 0.02d 12.41 ± 0.35c 13.91 ± 0.53c 15.67 ± 1.13b 17.37 ± 0.02a 17.54 ± 0.03a B1 15.23 ± 0.02 f 15.35 ± 0.02e 15.70 ± 0.02d 15.88 ± 0.03c 16.22 ± 0.02a 16.60 ± 0.03a B2 15.59 ± 0.02 f 15.71 ± 0.02e 15.90 ± 0.02d 16.36 ± 0.03c 16. 74 ± 0.03b 17.22 ± 0.03a C1 14.37 ± 0.02 d 14.80 ± 0.02c 14.89 ± 0.03c 15.15 ± 0.02bc 15.41 ± 0.02b 16.05 ± 0.58a C2 14.57 ± 0.02 f 15.25 ± 0.02e 15.79 ± 0.02d 15.85 ± 0.03c 17.44 ± 0.04b 17.74 ± 0.03a D1 13.03 ± 0.02 f 13.56 ± 0.02e 14.24 ± 0.03d 14.49 ± 0.03c 15.27 ± 0.03b 15.74 ± 0.03a D d2 13.57 ± 0.02 13.59 ± 0.03 d 13.67 ± 0.03d 14.76 ± 0.03c 15.40 ± 0.09b 15.70 ± 0.13a Notes: Mean ± SD of triplicate readings. Values with different superscripts in the same column are significantly different. Keys: Fermentation period: 0– 5 hr; Sample A = Short set yoghurt without any stabilizer; Sample B1 = Short set yoghurt with 0.5% CMC; Sample B2 = Short set yoghurt with 1.0% CMC; Sample C1 = Short set yoghurt with 0.5% Corn starch; Sample C2 = Short set yoghurt with 1.0% Corn starch; Sample D1 = Short set yoghurt with 0.5% Gum acacia; Sample D2 = Short set yoghurt with 1.0% Gum acacia. short set yoghurt samples. The pH values decreased (6.27 ± 0.01 to 3.2.2 | Total titratable acidity (TTA) 4.10 ± 0.03) as fermentation time increased. This decrease in pH as fermentation progressed could be attributed to the increased Titratable acidity values of the short set yoghurt samples containing and sustained metabolic activity of acid- producing microorganisms CMC, corn starch, and gum acacia at 0%, 0.5%, and 1.0% concen- (Gassem & Abu- Tarboush, 2000), resulting in the continued produc- trations showed significant differences (p < .05) between the sta- tion of lactic acid with consequent depression of pH. The result also bilizers, with gum acacia giving the highest values for TTA (Table 3). revealed a significant (p < .05) difference in the interaction effect The differences caused by concentrations of stabilizers on the short between the different stabilizers and their concentrations. The in- set were statistically significant (p < .05). It was observed that the teraction effect between stabilizers and concentration for the short addition of CMC at the level of 1.0% concentration depressed the set yoghurt samples at 0.5% and 1.0% concentrations did not mag- production of lactic acid. This was clearly shown in Table 4 by the nify much difference between the stabilizers. Due to higher reaction production rate of lactic acid in the presence of a high level of CMC. rates of yoghurt produced with the short set method, the final pH It was shown that at 0% concentration, the production rate of lactic produced in 5 hr was below 4.5. acid was 0.12% per hour, but this depressed to 0.09% per hour when 10  |     EZE Et al. TA B L E 4   Rate of fermentation of short set yoghurt samples Short Set yoghurt Conc. of MC (%/ Viscosity TTA (%/ TVC (cfu/ LAB (cfu/ Vit. B3 (SSY)_Stabilizers stabilizer (%) hr) (cP/hr) hr) pH (unit/hr) ml/hr) ml/hr) mg/ml/hr SSY_CMC 0 1.36 28.13 0.12 0.46 0.65 0.36 0.08 0.5 0.28 36.94 0.09 0.45 1.76 0.6 0.02 1.0 0.33 32.54 0.04 0.43 0.13 0.34 0.01 SSY_Corn Starch 0 1.36 28.13 0.12 0.46 0.65 0.36 0.08 0.5 0.35 47.41 0.07 0.39 0.58 0.32 0.01 1.0 0.62 20.48 0.09 0.37 0.53 0.44 0.01 SSY_Gum Acacia 0 1.36 28.13 0.12 0.46 0.65 0.36 0.08 0.5 0.54 28.14 0.06 0.36 0.77 1.28 0.02 1.0 0.50 29.52 0.13 0.37 0.90 0.27 0.02 Note: Mean ± SD of triplicate readings. Values with a different superscript in the same column are significantly different. Keys: MC, Moisture content; TVC, Total viable count; LAB, Lactic acid bacteria; TTA, Total titratable acidity; Vit. B3, Niacin content. CMC increased to 0.5%. When CMC increased to 1% concentration, viscous properties. It is possible that whereas the lactic acid bacteria the lactic acid production depressed further to 0.04% per hour. were not fermenting CMC, they could ferment corn starch and gum CMC is an anionic, water-s oluble polymer capable of forming a acacia to some extent. very viscous solution. CMC is insoluble in acidic conditions and more Hence, the higher concentration of corn starch and gum aca- soluble in alkali conditions, and the solubility is pH-d ependent (Ergun cia resulted in a higher rate change. The fermentation time had a et al., 2016). Therefore, the low acid production could be attributed significant effect (p < .05) on the total titratable acid of short set to its formation of highly viscous systems, which caused diffusion yoghurt samples. It was observed that TTA improved as fermenta- resistance that reduced mobility of reactants and the consequence tion time progressed. The TTA values for short set yoghurt ranged was the reduction of the rate at which the reacting species came from 0.29 ± 0.01% to 0.89 ± 0.02%. This result agreed with those together for fermentation to take place (Alakali et al., 2007). It could previously reported for other Labnehs (a strained yoghurt used for also be observed from Table 4 that significant interaction between sandwiches in an Arab country) (Benkerroum and Tamime, 2004). It stabilizers and concentrations of the different stabilizers suggests also agreed with the findings of Ahmad (1994), who reported that that the different concentrations of the stabilizers magnified the the total titratable acidity ranged from 0.87% to 1.13%. The interac- differences in the niacin content between the stabilizers. At 1.0% tion effect of stabilizers and concentrations was significant (p < .05), concentration, the rate of elaboration of niacin reduced further and suggesting that the effects caused by different stabilizers used were maintained the differences observed at 0.5% concentration. Gum different at different concentrations. acacia did not impede the production of titratable acidity. Instead, the higher concentration of gum acacia resulted in a higher lactic acid production rate (Table 4). It was seen that at 0.5% concentra- 3.2.3 | Viscosity tions, the rate of lactic acid production by gum acacia was 0.01% per hour, while at 1.0% concentration, it increased to 0.05% per hour. The viscosity values obtained for the short set yoghurt samples sta- Gum acacia is a stabilizer that functions as an emulsifying agent in bilized with CMC, corn starch, and gum acacia at 0, 0.5, and 1.0% milk products by producing a firmer texture (Roeper, 2014). Gum concentrations showed that significant differences (p < .05) were acacia has high water solubility (up to 50%. w/v) and relatively low found between the different stabilizers (Table 3). The differences in viscosity than other exudate gums. This polymer's highly branched viscosities have been attributed to the chemical and physical charac- molecular structure and relatively low molecular weight are respon- teristics of the stabilizers used. CMC can form high viscous colloidal sible for these properties (Dragnet, 2000). The low viscosity of gum solutions with water, insoluble in ethanol and slightly hygroscopic acacia allowed greater freedom of mobility of reactants, which en- (Dragnet, 2000). Corn starch can disperse and suspend other ingre- abled reacting species to come together for fermentation to take dients or particulate matter, thereby forming gels and provides the place. The titratable acidity of the short set yoghurt samples sta- body with food products (Erickson, 2006). On the other hand, gum bilized with corn starch at the level of 0%, 0.5%, and 1.0% concen- acacia dissolves easily in water (up to 50%), and the resulting solution trations had a less inhibitory effect on the production of titratable does not interact easily with other chemical compounds (ITC, 2008). acidity compared to CMC at similar concentrations. It was seen from Therefore, a comparison of the viscosity of gum acacia with sodium Table 4 that the rate of change at 0.5% concentration was 0.07% carboxyl methylcellulose, a common thickening agent, showed that and 0.09% per hour in the short set. Therefore, the result shows even at a concentration above 30%, gum acacia solution has a lower that corn starch in concentrations beyond 0.5% and 1.0% could be viscosity than 1.0% sodium carboxyl methylcellulose at low shear wastage unless used in combination with other stabilizers with lower rates. Also, while gum acacia is Newtonian in behavior with viscosity EZE Et al.      |  11 being shear rate- independent, sodium carboxyl methylcellulose dis- This could be attributed to the coagulation of the protein and played non- Newtonian shear thinning characteristics (Williams & carbohydrate during fermentation (Amankwah et al., 2009) as pH Phillips, 2009). decreased. However, due to the higher reaction rate caused by the The viscosity of gum acacia decreased in the presence of elec- elevated incubation temperature in the short set yoghurt, higher vis- trolytes due to charge screening and at low pH when the carboxyl cosity was achieved in the short set yoghurt within 5 hr. Interaction groups become undissociated (Williams & Phillips, 2009). At 1.0% effects between stabilizers and their concentration were found sig- concentration, CMC recorded the highest viscosity compared to nificant (p < .05). The behaviors of the stabilizers at different con- other stabilizers. Viscosity increased significantly (p < .05) with an centrations were different for different stabilizers. Thus, an increase increase in the concentration of each stabilizer, with CMC having in the concentrations of stabilizers magnified the differences in ef- the greatest effect. CMC increased from 58.33 ± 1.79 cP at 0% con- fect between stabilizers. centration to 152.74 ± 3.21 cP at 1.0% concentration, corn starch increased from 58.33 ± 1.79 cP to 88.23 ± 0.05 cP, and gum acacia increased from 58.33 ± 1.79 cP to 58.76 ± 0.08 cP. The rate of in- 3.3 | Effects of stabilizers and fermentation time crease in viscosity of short set yoghurt, as shown in Table 3, revealed on the microbial qualities of short set Yoghurt that the increase in viscosity caused by CMC and corn starch peaked at 0.5% concentration. Therefore, further addition of CMC and corn 3.3.1 | Total viable count starch would not increase the viscosity of short set yoghurt further. Gum acacia showed a linear relationship, and this implies that as the The total viable count of the short set yoghurt stabilized with CMC, concentration of the stabilizer increases, viscosity increases. It was corn starch, and gum acacia showed significant (p < .05) differences observed that there was an increase in viscosity of short set yoghurt among the stabilizers. CMC and gum acacia gave the highest values as the fermentation time increased. of TVC and lactic acid bacteria counts compared to corn starch for short set yoghurt. Total viable count decreased significantly (p < .05) with increased concentration. TVC values for the short set yoghurt 3.2.4 | Total solids samples ranged from 1.304 ± 2.00 to 8.30 ± 3.00 × 104 (Table 5). The rate of multiplication of microorganisms indicated that at 0.5% The values presented in Table 4 showed information on the total concentration of CMC, the rate of TVC was lower when compared solids of short set yoghurt samples stabilized with 0%, 0.5%, and to 1.0% concentration of the same stabilizer. This suggests that 1.0% concentrations of CMC, corn starch, and gum acacia. There this concentration of stabilizer (1.0%) provided the optimum con- was a significant (p < .05) difference in the total solids. Total solids ditions for the growth of the microorganisms. For corn starch and were higher in the samples with stabilizers than the control yoghurt gum acacia, the rate of total viable count production decreased with sample as the concentration increased. This trend was consistent an increase in the concentration of the stabilizers, apparently be- with the report of Mehanna et al., (2013), in which the total solids, cause they did not provide optimum conditions for the growth of the protein, and fat contents were found to be higher in stabilized yo- microorganisms. ghurt samples. The increment in the total solids was said to have resulted from the stabilizers incorporated in the samples. Samples stabilized with corn starch gave the highest total solids content 3.3.2 | Lactic acid bacteria count (17.74 ± 0.03%) followed by CMC (17.22 ± 0.03%), while gum aca- cia had the least total solids contents (15.74 ± 0.03%). This could LAB values for short set yoghurt samples at 0% concentration were be because each polymer chain in a dilute solution of corn starch 7.8 × 105 cfu/ml, 3.72 × 105 CFU/ml at 0.5% concentration, and is hydrated and extended, therefore exhibiting stable consistencies 2.96 × 105 CFU/ml at 1.0% concentrations. The LAB count values (Edali et al., 2001). The total solid content increased with an increase obtained ranged from 1.12 × 104 ± 3.00 to 8.40 × 104 ± 4.24 CFU / in fermentation time. This result was in agreement with the findings ml. However, the decrease in the activity of the lactic acid bacteria of Sahid et al., (2002), who reported the total solids content with caused an increase in the pH of the yoghurt samples. The rate of a range of 13.80% to 18.30%. This result showed that total solids LAB multiplication for the short set yoghurt (Table 5) indicated that accumulate as fermentation progresses because moisture was lost. at 0.5% concentration of CMC, multiplication of LAB was faster The interaction effect between stabilizer and concentrations on the when compared to 1.0% concentration. The rate of multiplication total solids contents was significant (p < .05). The increase could be of LAB for corn starch and gum acacia showed a decrease with an attributed to the accumulation of solid matter during fermentation. increase in the concentration of the stabilizer. This could be be- CMC incorporated in the short set yoghurt samples significantly had cause the optimum conditions for LAB proliferation were provided the highest total solids compared to other stabilizers, while 1.0% at a 0.5% concentration of CMC compared to other concentra- concentration recorded the highest total solids compared to other tions. From Table 5, it was observed that LAB proliferation for the concentrations. Therefore, the differences in the effect of the stabi- different stabilizers at 0.5% concentration was slow compared to lizers were magnified by the concentrations used. the rate of proliferation of LAB at 1.0% concentration; this could 12  |     EZE Et al. TA B L E 5   Microbial properties of short set yoghurt samples Fermentation period (hr) Microbial parameters Sample 0 1 2 3 4 5 TVC A 2.00 ± 2.65d 7.30 ± 2.00c 1.04 ± 2.65b 1.17 ± 2.56a 1.20 ± 2.56a 1.05 ± 2.65b B1 1.30 ± 2.00 f 2.30 ± 2.00e 3.50 ± 2.65d 4.10 ± 2.65c 9.30 ± 3.46b 1.02 ± 3.00a B2 1.60 ± 3.00 d 3.00 ± 3.00b 2.40 ± 3.00c 2.30 ± 3.00c 5.30 ± 3.00a 5.20 ± 3.00a C1 1.80 ± 3.00 b 1.90 ± 3.00b 2.00 ± 3.00b 2.20 x ± 3.00b 4.60 ± 2.00a 4.20 ± 2.00a C2 1.30 ± 3.46 c 2.10 ± 3.00b 2.20 ± 3.00b 2.40 ± 3.00c 3.90 ± 3.00a 4.00 ± 3.00a D 2.10 ± 1.73e 3.50 ± 3.00d1 4.70 ± 3.46 c 7.90 ± 3.46b 1.07 ± 2.12a 8.30 ± 3.00a D2 1.80 ± 1.73 e 2.36 ± 3.05d 4.50 ± 3.46c 4.90 ± 3.00c 5.50 ± 3.46a 5.60 ± 3.00b LAB A 2.60 ± 2.65f 7.60 ± 2.65d 1.21 ± 2.65a 1.12 ± 3.00b 9.60 ± 2.56c 3.70 ± 4.36e B 2.0 ± 3.00d 3.40 x ± 3.00 4.00 ± 3.00b 4.30 ± 3.00b 5.30 ± 2.00a 5.00 ± 3.00a1 c B 2.30 ± 3.00d 3.40 x ± 3.00c 3.57 ± 3.51bc 3.80 ± 2.00bc2 4.50 ± 3.00 a 4.00 ± 2.00ab C1 1.80 ± 3.00 b 2.00 ± 3.00b 2.00 ± 3.00b 2.20 ± 3.00b 3.40 ± 2.00a 3.20 ± 1.00a C d2 1.60 ± 3.00 1.90x±3.00cd 2.30 ± 3.00 b 2.30 ± 3.00b 3.70 ± 3.00a 3.60 ± 3.00a D1 2.10 ± 3.00 e 2.50 ± 3.00e 3.50 ± 3.46d 5.50 ± 3.46c 6.30 ± 3.00b 8.40 ± 4.24a D2 1.80 ± 3.00 b 2.30 ± 3.00b 2.90 ± 3.46a 3.00 ± 3.46a 3.30 ± 3.00a 3.10 ± 3.00a Note: Mean ± SD of triplicate readings (×104 CFU/ml). Values with different superscripts in the same column are significantly different. Keys: Fermentation period: 0- 5h; Sample A = Short set yoghurt without any stabilizer; Sample B1 = Short set yoghurt with 0.5% CMC; Sample B2 = Short set yoghurt with 1.0% CMC; Sample C1 = Short set yoghurt with 0.5% Corn starch; Sample C2 = Short set yoghurt with 1.0% Corn starch; Sample D1 = Short set yoghurt with 0.5% Gum acacia; Sample D2 = Short set yoghurt with 1.0% Gum acacia. be attributed to the conditions of fermentation, which did not The mean score for flavor ranged from neither liked nor dis- favor the rapid LAB growth. Fermentation time led to a significant liked (5.25 ± 0.55) to liked moderately (7.15 ± 0.49). Samples con- (p < .05) increase in the total viable count of short set yoghurt. taining 1.0% gum acacia had the highest score for flavor. Pecivova This was consistent with other reported works by Gassem and Abu- et al., (2013) reported that the addition of gum acacia could enhance Tarboush (2000), which reported LAB evolution in yoghurt, show- the quality and flavor of pizza flans. Notably, there was significant ing an increase in number versus fermentation time. The significant (p < .05) improvement in flavor for samples stabilized with corn interaction effect between stabilizers and the concentrations of starch followed by those containing gum acacia as concentration the different stabilizers suggested that the different stabilizers' be- increased. However, this trend was reversed in samples containing havior was different at different concentrations. This implies that CMC with an increase in concentration. The flavor of short set yo- higher concentrations magnified the differences between stabiliz- ghurt produced with corn starch was most preferred. This could be ers in the total viable count and lactic acid bacteria count in the due to starch hydrolysis, which leads to the release of sweetening yoghurt samples. properties of the starch (Erickson, 2006). There was marked signifi- cant (p < .05) improvement in the sensory score in each of the stabi- lized yoghurt samples at 1.0% concentrations. For short set yoghurt, 3.4 | Effects of stabilizers and fermentation time all the sensory parameters (especially flavor) improved with an in- on the sensory attributes of short set Yoghurt crease in the concentration of the stabilizers and as fermentation time progressed (3–5 hr). Therefore, the differences between the The sensory scores of short set yoghurt samples stabilized with stabilizers could be attributed to the acidity level developed during CMC, corn starch, and gum acacia at 0%, 0.5%, and 1.0% concentra- fermentation (Alakali et al., 2007). tions are presented in Table 6. There were significant (p < .05) dif- ferences in the sensory parameters of the yoghurt samples from the result. 3.4.2 | Mouthfeel The control yoghurt had higher mouthfeel scores when the fermen- 3.4.1 | Flavor tation was left for 3 hr compared to other treated short set yoghurt samples. As the fermentation progressed to 4 hr, gum acacia con- Table 6 showed the sensory scores of the short set yoghurt samples taining yoghurt samples were liked moderately (7.00 ± 0.00) at 1.0% stabilized with CMC, corn starch, and gum acacia. concentration. By the time the fermentation had reached 5 hr, only EZE Et al.      |  13 TA B L E 6   Sensory attributes of Short Set Yoghurt Samples Fermentation period (hr) Sensory parameters Sample 3 4 5 Color A 6.75 ± 0.44ab 7.00 ± 0.00a 7.35 ± 0.59a B1 6.45 ± 0.51 c 6.80 ± 0.41a 7.30 ± 0.47a B2 6.20 ± 0.41 b 6.60 ± 0.59b 6.55 ± 0.61a C1 5.60 ± 0.59 c 6.00 ± 0.34b 6.65 ± 0.49a C2 6.10 ± 0.31 c 6.75 ± 0.44b 7.30 ± 0.47a D1 6.10 ± 0.31 b 6.35 ± 0.49ab 6.60 ± 0.50a D2 6.25 ± 0.44 c 6.60 ± 0.68b 7.45 ± 0.51a Flavor A 6.00 ± 0.00c 6.40 ± 0.50b 6.80 ± 0.41a B1 6.20 ± 0.41 c 6.45 ± 0.51b 6.80 ± 0.41a B2 5.60 ± 0.50 a 5.65 ± 0.49a 5.25 ± 0.72b C 5.25 ± 0.55b1 5.80 ± 0.62 ab 6.20 ± 0.41a C2 5.85 ± 0.49 c 6.55 ± 0.69b 7.10 ± 0.31a D1 5.30 ± 0.92 b 6.20 ± 0.69a 6.40 ± 0.50a D2 6.15 ± 0.37 c 7.00 ± 0.46ab 7.15 ± 0.49a Taste A 6.15 ± 0.37c 6.30 ± 0.47b 6.95 ± 0.22a B1 5.80 ± 0.41 c 6.20 ± 0.41b 6.70 ± 0.47a B2 5.35 ± 0.49 a 5.05 ± 0.61b 5.35 ± 0.81a C1 5.15 ± 0.49 a 6.10 ± 0.64b 6.85 ± 0.59a C2 5.80 ± 0.41 c 6.55 ± 0.51b 7.05 ± 0.51a D1 4.60 ± 1.47 c 6.00 ± 0.97b 6.45 ± 0.61a D2 5.95 ± 0.39 b 6.90 ± 0.72a 7.10 ± 0.55a Mouthfeel A 6.25 ± 0.44b 6.55 ± 0.51b 6.90 ± 0.31a B1 6.30 ± 0.57 c 6.55 ± 0.51b 6.85 ± 0.37a B 5.40 ± 0.50b2 5.40 ± 0.68 b 5.65 ± 0.49a C1 5.35 ± 0.59 b 6.00 ± 0.32a 6.30 ± 0.47a C2 6.00 ± 0.46 b 6.80 ± 0.41ab 7.00 ± 0.46a D 4.80 ± 1.39c 6.10 ± 0.85b1 6.20 ± 0.69 a D2 6.10 ± 0.64 b 7.00 ± 0.00a 7.40 ± 0.50a Overall Acceptability A 6.05 ± 0.22c 6.80 ± 0.52b 7.35 ± 0.49a B1 6.00 ± 0.31 b 6.60 ± 0.50b 7.05 ± 0.39a B 5.90 ± 0.31b 5.55 ± 0.83c2 6.05 ± 0.69 a C1 5.20 ± 0.41 c 6.35 ± 0.61b 6.95 ± 0.22a C2 6.10 ± 0.55 c 7.30 ± 0.47b 7.90 ± 0.31a D1 5.40 ± 1.09 c 6.75 ± 0.44b 6.90 ± 0.31a D2 6.50 ± 0.51 b 7.25 ± 0.44a 7.65 ± 0.49a Note: Mean ± SD of triplicate readings. Values with different superscripts in the same column are significantly different. those samples stabilized with 1.0% gum acacia were found to have time progressed from 4 to 5 hr, compared to the samples without higher mouthfeel scores than the control yoghurt; nevertheless, the any stabilizer. A similar trend in corn starch was observed when gum improvement observed was not significant (p > .05). acacia was added to the short set yoghurt samples. All samples stabilized with CMC were found to score lower scores for mouthfeel than the control yoghurt samples as concentra- tions increased. With corn starch added to the samples, the mouth- 3.4.3 | Color feel scores significantly improved as the concentrations changed from 0.5% to 1.0%. A notable improvement was achieved in the The increased concentration of CMC was observed to have ad- mouthfeel of corn starch containing samples as the fermentation versely affected the appearance (color) of short set yoghurt samples 14  |     EZE Et al. significantly (p < .05) in comparison with the short set yoghurt with- The interaction effect between stabilizers and concentrations out a stabilizer. As the fermentation time increased (4 to 5 hr), the on the sensory parameters showed that the differences between decrease in the sensory color scores so observed was no longer sig- the stabilizers were different at different concentrations or that the nificant (p > .05). differences between stabilizers on the color, taste, flavor, mouth- Although samples in which corn starch was added were found feel, and overall acceptability of short set yoghurt were magnified at to have higher sensory scores for color as the fermentation time in- higher concentrations. creased (3–5 hr), the yoghurt samples without a stabilizer (control) still had higher sensory scores compared to the treated yoghurt samples. Only samples formulated with gum acacia at the 1.0% level 4  | CONCLUSION were scored higher in appearance than any other yoghurt samples (treated with stabilizers and plain yoghurt). The results obtained in this work indicated that the addition of sta- bilizers and the use of different fermentation times improved the proximate, physicochemical, microbial, and sensory properties of 3.4.4 | Taste short set yoghurts. It was also observed that 1.0% concentration of CMC and corn starch was optimal for short set yoghurt samples Compared to control samples, all short set yoghurt samples formu- fermented at 40– 43℃ beyond which their usage becomes wastage lated with CMC had lower taste scores and increased concentrations when compared to gum acacia which appeared to require a higher (from 0.5% to 1.0%). The decrease in the taste sensory scores in the concentration of more than 1.0% in order to equilibrate with 0.5%– CMC- containing yoghurt samples was only found to be significant 1.0% CMC. Due to its significant impact on the desirable qualities— (p < .05) when the fermentation time was 3 hr. total solids, moisture, overall acceptability, and good keeping quality There was an improvement in the taste sensory scores for all sam- on the produced yoghurt samples compared to other stabilizers ples formulated with corn starch as fermentation progressed (3–5 hr). (Gum acacia and CMC), the use of corn starch is therefore recom- All samples containing corn starch had increased taste scores as the mended for the production of short set yoghurts. However, further concentrations changed from 0.5% to 1.0%. There was an improvement improvement may be carried out on the sensory attributes and the in taste sensory scores observed in all samples formulated with gum acidity of short set yoghurt. Also, there is a need to improve gum acacia, which was significant (p < .05) with the increase in the fermen- acacia usage by combining it with other stabilizers to improve its tation time (3– 5 hr). Samples containing 1.0% gum acacia (7.10 ± 0.55) rheological properties since it produces good textural properties in were liked moderately. In comparison, gum acacia containing samples dairy products. at 0.5% level (4.60 ± 1.47) while the fermentation lasted for 3 hr was found to have the least taste sensory scores. Further addition of gum CONFLIC T OF INTERE S T acacia could increase dryness resulting in an adverse effect on the tex- The authors do not declare any conflict of interest before, during or ture and taste of the samples (Pecivova et al., 2013). after this study. AUTHOR CONTRIBUTION 3.4.5 | Overall acceptability Chinazom Martina Eze: Conceptualization (equal); Data cura- tion (equal); Investigation (equal); Methodology (equal); Project The control yoghurt samples (plain) had higher sensory scores for administration (equal); Resources (equal); Writing- original draft overall acceptability compared to the samples containing CMC, as (equal); Writing- review & editing (equal). Kehinde Oludayo Aremu: concentrations increased from 0.5% to 1.0%. Compared to the con- Conceptualization (equal); Data curation (equal); Formal analy- trol (short set yoghurt without stabilizer), there was an increase in sis (equal); Investigation (equal); Methodology (equal); Writing- the sensory scores for overall acceptability in yoghurt samples for- original draft (equal); Writing- review & editing (equal). Emmanuel mulated with corn starch as concentrations increased. Fermentation Oladeji Alamu: Conceptualization (equal); Data curation (equal); was found to influence the sensory scores of samples in which corn Investigation (equal); Resources (equal); Validation (equal); starch was added because of higher values obtained as fermentation Visualization (equal); Writing-r eview & editing (equal). Thomas progressed (from 3 to 5 hr), but this was not significant (p > .05). The Muoneme Okonkwo: Conceptualization (equal); Data curation highest sensory scores or overall acceptability was found in samples (equal); Funding acquisition (equal); Project administration (equal); formulated with corn starch at a 1.0% level. Samples in which gum Validation (equal); Visualization (equal); Writing- review & editing acacia was incorporated showed a similar trend as obtained in sam- (equal). ples containing corn starch. 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