Use of kefir and buttermilk to produce an innovative quark cheese

Abstract

Quark cheese is a fermented soft fresh cheese categorised under acid-rennet coagulated cheeses. In this study, alternative raw materials such as kefir and yayik buttermilk were used to produce Quark cheese in comparison with the cheese produced by the acidification of skim milk with mesophilic lactic culture. Samples were kept individually under 35 °C and 100 °C for coagulum formation. Obtained cheeses, were evaluated in terms of some physicochemical, microbiological and sensorial properties in addition to the volatile and peptide profiles. Quark produced from kefir and buttermilk was determined to have preferred properties directly affect the cheese characteristics.

Introduction

Fresh cheeses are non-ripened cheeses produced by either coagulation of acidified milk at pH 4.6–4.8 or by the combination of acid-heat application, and small amount of rennet enzyme is used in both alternatives (Litopoulou-Tzanetaki 2007). Acidification is usually provided with mesophilic lactic acid bacteria (LAB) mainly of Lactococcus lactis subsp. lactis, Lactococcus lactis subsp. cremoris and Lactococcus lactis subsp. diacetylactis.

Quark cheese is categorised under acid or acid/rennet-curd fresh cheeses. It is a soft white cheese with a slight acidic flavor and a moderate sour taste, made from pasteurized skim milk with starter culture and a bit rennet addition. It can be made from skim or full fat milk (Litopoulou-Tzanetaki 2007).

Kefir is a fermented dairy beverage which is produced by the activity of kefir grains, including a diverse microorganisms such as LAB, acetic acid bacteria and yeasts (Satir and Guzel-Seydim 2016). It has some functional roles to minimise the adverse impacts of lactose intolerance, immunity variation, stability of intestinal flora, as well as antimicrobial, anticarcinogenic and antioxidant activities (McCue and Shetty 2005; de Moreno de LeBlanc et al. et al. 2006). It has positive influence on the intestinal mucosa permeability, furthermore support the latitude of allergenic reactions patients (Wróblewska et al. 2009).

Yayik buttermilk is the aqueous phase left when full fat fresh or strained yoghurt is churned to butter traditionally instead of cream (Sagdıç et al. 2002). Yayik buttermilk is obtained as a nutritious liquid contains all the water-soluble components of yoghurt, including serum proteins, lactose, and minerals (Sodini et al. 2006). Similar to kefir, yayik buttermilk also includes lactic acid bacteria derived from yoghurt.

In this study, yayik buttermilk and kefir were used as alternative raw materials in the production of Quark cheese instead of cultured skim milk, due to their functional properties enhancing physical, aromatic and sensory properties of Quark cheese. Kefir and yayik buttermilk were used without the supplementation of starter culture during the production of Quark cheese, which were accepted having a natural starter bacteria. Yayik buttermilk was preferred as raw material in the production of Quark cheese because of its emulsifying ability (Sodini et al. 2006) and supportive effects on flavor (Sodini et al. 2006). Kefir was also used as alternative to milk because of its functional properties and organoleptic properties (Wszolek et al. 2007). To the best of our knowledge, there is not much information available in literature, regarding characteristics of Quark-type cheeses (Gámbaro et al. 2017; Miloradovic et al. 2018) or usage of different raw materials during Quark production. Therefore, we aimed to develop an innovative Quark-type cheese by using alternative raw materials such as kefir and yayik buttermilk which are rich in nutritional ingredients, having diverse microorganisms and more functional roles compared to milk. The objectives of the present research were to evaluate the physicochemical, sensory and microbiological properties, as well as the volatile and peptide profiles of newly designed Quark cheeses in comparison with the Quark produced from milk with mesophilic starter culture.

Materials and methods

Productions of kefir, yayik buttermilk, and skim-milk

Raw bovine milk was obtained from Department of Dairy Technology, Ankara University (Ankara, Turkey). The whole amount of milk was divided into two portions and the first part was pasteurised at 85 °C for 15 min and were cooled to 43 °C for yoghurt production. The other part was heated to 50–55 °C to remove the fat by a seperator (Subitas, Turkey). The fat content of the milk was measured as 0.5% following the fat seperation. This milk was again divided into two portions equally and both were pasteurised at 85 °C for 15 min and were cooled to 25 °C and 28 °C for kefir and cultured skim-milk productions, respectively.

For the production of yoghurt, full-fat (3.6%) pasteurised milk was inoculated with yoghurt starter culture containing Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus (DVS CH1, Chr. Hansen, Horsholm, Denmark) at a ratio of 2% (w/v) and incubated at 43 °C. The fermentation was ceased at pH 4.6. Following this stage, yoghurt were stored at 4 °C for 12 h. The yoghurt was mixed with water in a ratio of 2:1 for buttermilk production. Buttermilk was obtained by churning of yoghurt at 16–18 °C for 2 h followed by the seperation of the cream from the top.

To produce kefir, low-fat (0.5%) pasteurised milk was inoculated with 1% (w/v) kefir grains and incubated at 25 °C without agitation until pH decreases to 4.5 which took approximately 18 h. At the end of the fermentation, grains were seperated from product.

For the third raw material, mesophilic homofermentative culture (R-703, Chr-Hansen, Denmark) was supplemented into the other part of the low-fat pasteurised milk at a level of 0.01 g/L and incubated at 28 °C for 18 h with the aim of culture activation. Each raw material was stored at 4 °C overnight until they were used for Quark cheese production.

Production of quark cheese

Previously prepared kefir (K), yayik buttermilk (YBM), and cultured skim-milk (SM) were individually used for the production of Quark cheeses. The pH of K, YBM and SM are 4.31, 4.33 and 4.26, respectively. Moreover, the total solid contents are 17.33, 14,95 and 19.44% for the same raw materials, respectively. 0.01% (v/v) rennet was added to each raw material and all were divided into two for evaluating the effects of heat treatments at 35 °C and 100 °C. Samples were kept under 35 °C and 100 °C, for 16 h and 2 h, respectively for coagulum formation and each of them were rapidly cooled to 4 °C before removal of whey in cheese cloths. Filtration of the samples were carried out until the water droplets formation ends (Fig. 1). This stage was carried out for 16 h and 22 h for the samples of heat treated at 100 °C and 35 °C, respectively. Two replicates of each type of Quark cheese were produced on two successive weeks. The cheeses from each treatment were analysed on days 1 and 14 during storage.

Fig. 1.

Production of quark cheeses. a Drainage and the removal of serum phase, b Drained sample

Sample codes and the production conditions were given at the following;

SM35: Skim milk inoculated with mesophilic culture at 35 °C

SM100: Skim milk inoculated with mesophilic culture at 100 °C

K35: Kefir at 35 °C

K100: Kefir at 100 °C

YBM35: Yayik buttermilk at 35 °C

YBM100: Yayik buttermilk at 35 °C

Physicochemical analyses

The pH of Quark cheeses was determined by the insertion of a pH-meter electrode (Mettler Toledo MP 225, Columbus, OH) into the slurry sample obtained by maceration of a 10 g sample with 10 mL of distilled water. Titratable acidity was measured according to AOAC (1995). The total solid was analyzed by the oven drying method at 102 °C (IDF 1982), and fat content was determined by the Gerber-Van Gulik method. Total nitrogen was measured by applying the Kjeldahl method on samples prepared as described by Kuchroo and Fox (1982). The water-holding capacities (WHC) of samples were measured by the centrifugation technique (Isanga and Zhang 2009) and values were calculated according to the Eq. 1:

$$WHC\left(\%\right)=\left(1-W1/W2\right)\times 100$$ 1

All analyses were performed in duplicate.

Microbial counts

10 g cheese samples were homogenized in a Stomacher with 90 mL Ringer solution (Bag Mixer 400 VW, Interscience, France) for 3 × 1.5 min. Serial dilutions were made in Ringer solution (Merck, Darmstadt, Germany) and plated in duplicate. Total aerobic mesophilic bacteria were determined on Tryptic Soya Agar including 1% skimmed-milk at 35 °C for 48 h. Lactococci were monitored on M17 agar containing 10% (w/v) sterile lactose solution which incubated under aerobic conditions at 37 °C for 48 h. In order to determine the Lactobacilli count MRS Agar were used under unaerobic incubation at 37 °C for 72 h. Filamentous fungi and yeasts were grown on Potato Dextrose Agar at 30 °C for 3–5 days. Coliform bacteria were enumerated on Violet Red Bile Agar following aerobic incubation at 37 °C for 24 h. Results were calculated in log₁₀ cfu/g.

Analysis of proteolysis (Peptide profile)

The proteolysis levels of samples were determined according to Kuchroo and Fox (1982). 20 g of Quark cheese was homogenized with 100 mL distilled water at 40 °C with Ultra-Turrax (Heidolph, Schwabach, Germany). The mixture was incubated at 40 °C for 1 h. The pH was adjusted to 4.6 with 1 M HCL followed by centrifugation at 5000 rpm for 15 min.

The peptides of the pH 4.6-soluble fraction of cheeses were measured using Reversed-Phase High Performance Liquid Chromatography (1100 series, Agilent Technology, CA, USA) equipped with a UV Dedector at 214 nm and a C18 (4.6 cm x 250 mm × 5 μm, 300Å pore size) column. The soluble fraction of sample was dissolved in a mixture of 0.2% trifluoroacetic acid and deionised water (1:1), subsequently 40μL sample from this mixture was injected to HPLC with a flow rate of 0.75 mL/min.

During the elution, solvent A (0.1% TFA (v/v) in deionised HPLC-grade water) and solvent B (0.1% TFA (v/v) in UV-grade acetonitrile) were used. The applied gradient programme was; 100% solvent A for 10 min, followed by a linear gradient to 50% B (v/v) over 80 min, increasing to 60% B (v/v) over 5 min and running at 60% B (v/v) for 5 min.

Volatile compound analyses

Volatile compound analyses of cheese samples were performed according to Lee et al. (2003) by employing the solid phase micro extraction (SPME) method, with minor modifications. 5 g cheese were weighted in a 20-mL vial and 10 μL of Internal Standard (2 methyl-3 heptanone and 2-methyl pentanoic acid) was added onto it. The vial was holded at 50 °C for 30 min for equilibrium of volatile compounds in the headspace by continuous stirring. Extraction of volatile compounds was carried out by injecting 75 μm of carboxen/polydimethylsiloxane SPME fiber (Supelco, Bellefonte, PA) into the vial.

The volatile compounds were desorbed by direct insertion of the fiber into the injection port of the Gas Chromatography coupled to a mass spectrometer (7890 GC system, Agilent Technologies, Santa Clara, CA). The GC was equipped with a DB-Wax column (30 m, 0.25 mm x 0.25 μm). Helium was used as carrier gas at a flow rate of 1 mL/min. The initial temperature program was employed at 45 °C for 10 min, followed by elevation of the temperature to 110 °C at a rate of 5 °C/min and 240 °C at a rate of 10 °C/min.

Volatile compounds were identified according to n-alkane standards (Sigma-Aldrich, St. Louis, MO) and by comparison of their mass spectra with those in the libraries of Wiley, NIST and Flavor. The concentration of compounds were determined by the calculation of the peak area of the internal standard to the area of the unknown compound. Samples were analyzed in duplicate.

Sensory analysis

Preference test was performed by 7 trained panelists from academic staff of Dairy Technology Department (Ankara University, Ankara, Turkey). Quark cheeses on day 1 and 14 were evaluated according to the scoring card described by Bodyfelt et al. (1988) with minor modifications. The samples were randomly coded and panelists were requested to evaluate color-appearance, body-texture and odour-flavor-taste of the samples in terms of sensory attributes.

Statistical analysis

Analysis of variance (ANOVA) was performed on data obtained from individual Quark cheeses made from buttermilk, kefir and cultured skim-milk on day 1 and 14, using Minitab statistical package (version 15.0, Minitab Inc., State College, PA). One-way ANOVA and Tukey’s multiple comparison tests were used to compare the data.

Results and discussion

Physicochemical parameters

Table 1 shows the average composition of quark cheeses made from various raw materials. Quark cheeses made from kefir and yayik buttermilk had lower pH values than Quark cheese made from skim-milk with mesophilic culture (P < 0.05). Treatment temperature of 100 °C caused a higher pH value in samples than 35 °C. The pH values of samples except for YBM35 did not show a significant change during the 14 days of storage (P > 0.05). However, when the pH values of cheeses from different raw materials were compared, cheeses made from SM were determined with higher pH values than the cheeses made from K and YBM as a result of the lactic starter culture effect.

Table 1.

Physicochemical properties of quark cheeses

	Period of storage
Samples	Day 1	Day 14
pH value
SM35	4.05 ± 0.020B	4.00 ± 0.030B
SM100	4.12 ± 0.020B	4.15 ± 0.005B
K35	3.71 ± 0.010B	3.75 ± 0.015B
K100	3.90 ± 0.035D	3.97 ± 0.005D
YBM35	3.91 ± 0.032D	3.93 ± 0.019A
YBM100	3.95 ± 0.025CD	3.96 ± 0.010CD
Titratable acidity (°SH)
SM35	86.6 ± 1.93BC	87.7 ± 2.28
SM100	83.1 ± 10.6C	83.0 ± 7.39
K35	100.6 ± 2.97AB	99.5 ± 4.10
K100	96.2 ± 2.20ABC	94.8 ± 4.19
YBM35	109.7 ± 6.84A	114.6 ± 5.33
YBM100	106.0 ± 5.05A	110.9 ± 1.08
Total solid (% w/w)
SM35	22.7 ± 1.28	22.5 ± 1.04
SM100	23.5 ± 0.91	23.0 ± 1.02
K35	21.4 ± 1.32	21.2 ± 0.57
K100	24.6 ± 1.33	24.3 ± 2.04
YBM35	21.6 ± 1.55	21.3 ± 1.42
YBM100	25.8 ± 5.75	24.6 ± 0.71
Fat (% w/w)
SM35	2.50 ± 1.50	2.25 ± 1.75
SM100	2.50 ± 1.50	2.25 ± 1.75
K35	3.75 ± 0.25	3.50 ± 0.00
K100	3.50 ± 0.50	3.50 ± 0.25
YBM35	3.75 ± 1.00	3.50 ± 1.25
YBM100	3.75 ± 1.25	3.50 ± 1.50
Total protein (% w/w)
SM35	13.4 ± 1.78	13.6 ± 2.01
SM100	14.4 ± 0.34	14.6 ± 1.22
K35	11.9 ± 0.78	11.8 ± 0.19
K100	14.4 ± 2.40	13.4 ± 1.34
YBM35	13.5 ± 0.42	13.4 ± 0.38
YBM100	17.9 ± 2.34	14.7 ± 1.26
Water holding capacity (%)
SM35	72.0 ± 3.42BC	68.5 ± 4.05
SM100	77.0 ± 4.25AB	73.5 ± 3.10
K35	74.0 ± 3.94BC	72.0 ± 2.40
K100	81.0 ± 1.75A	80.0 ± 1.75
YBM35	67.0 ± 2.13CD	65.0 ± 2.25
YBM100	70.0 ± 2.20C	67.7 ± 2.38

^A–DMeans with different uppercase letters significant differences between values of samples and storage days for each property (P < 0.05)

*Non-lettering columns or rows indicates the differences between scores are not found significant (P > 0.05)

Titratable acidities of Quark cheeses made from K and YBM were found higher compared to Quark cheese made from SM on day 1 and 14, while acidities were determined lower on day 14 compared to day 1, except for YBM (P < 0.05). This may be explained by the development of the total bacterial activity during the storage period. The decrease observed in the number of Lactococcus spp. and Lactobacillus spp. (Table 2) might be the reason of the reduction determined in the titratable acidity. This reduction was more pronounced in samples made from K in which some other microorganisms such as yeasts are also involved into the production process. Furthermore, Quark cheeses obtained by treatments at 100 °C showed a lower acidity compared to those treated at 35 °C in all samples. The reason of this result could be the lethal effect of high temperature on lactic bacteria.

Table 2.

Viable counts [log cfu/g] of microbial populations in Quark cheeses

	Period of storage
Samples	Day 1	Day 14
Lactococcus spp.
SM35	7.79 ± 0.96	6.19 ± 0.19
SM100	7.32 ± 0.53	6.39 ± 0.39
K35	7.05 ± 0.31	5.43 ± 0.06
K100	6.54 ± 0.24	6.42 ± 0.04
YBM35	7.26 ± 0.88	6.22 ± 0.97
YBM100	6.76 ± 0.14	6.70 ± 0.16
Lactobacillus spp.
SM35	6.36 ± 1.05AB	4.85 ± 0.15C
SM100	6.08 ± 0.14AB	5.77 ± 0.18ABC
K35	6.61 ± 0.19AB	6.80 ± 0.14A
K100	6.62 ± 0.08AB	6.85 ± 0.72A
YBM35	6.61 ± 0.42AB	6.80 ± 0.93A
YBM100	5.47 ± 0.04BC	5.83 ± 0.06ABC
Total aerobic bacteria
SM35	6.80 ± 0.61	8.19 ± 0.39
SM100	6.97 ± 0.57	7.05 ± 0.02
K35	7.26 ± 0.22	6.72 ± 0.07
K100	7.11 ± 1.16	7.24 ± 0.03
YBM35	7.02 ± 1.03	7.19 ± 0.95
YBM100	6.60 ± 0.25	6.90 ± 0.15
Yeast and mold
SM35	2.56 ± 0.04CD	3.30 ± 0.70BCD
SM100	1.54 ± 0.24D	2.07 ± 0.29D
K35	6.40 ± 0.72AB	7.45 ± 0.02A
K100	5.59 ± 0.42ABC	6.59 ± 0.14A
YBM35	3.09 ± 0.88A	3.85 ± 1.06A
YBM100	2.95 ± 0.06CD	7.12 ± 0.10A
Coliform bacteria
SM35	nd	nd
SM100	nd	nd
K35	nd	nd
K100	nd	nd
YBM35	nd	nd
YBM100	nd	nd

^A–DMeans with different uppercase letters significant differences between values of samples and storage days for each property (P < 0.05)

*Non-lettering columns or rows indicates the differences between scores are not found significant (P > 0.05)

The total solid (TS) and fat contents of cheeses were variable among samples. These differences were most probably related to the different raw materials used for preparation of Quark cheeses. Cheeses produced from YBM and K at 100 °C showed the highest TS content. However, TS, protein and fat values did not show any significant change (P > 0.05) during the whole storage. Whereas, samples treated at 100 °C had higher TS and protein contents when compared to 35 °C. Moreover, when the values of samples treated at 35 °C and 100 °C were compared, it was found that a higher TS content determined in the products processed at 100 °C had lower acidity values. Similar observations were reported by Kristo et al. (2003) who found a lower TS content indicates a higher titratable acidity values. These results were determined in parallel with the variation determined in water holding capacities (WHC). TS and fat contents of raw materials and accordingly the samples produced from them were determined as an important factor on the WHC values of samples. Hinrichs et al. (2004) also reported that higher TS content promotes the interactions between serum phase and solids which resulted in a higher WHC. Moreover, effect of treatment was found significant on WHC. The samples treated at 100 °C demonstrated a higher WHC due to the higher protein contents that they have, which indicates a low possibility of the breakdown of protein network and a firmer structure which retain the serum phase (Lucey 2002). Similar findings were recently reported by Miloradovic et al. (2018) that protein matrices had a higher water holding capacities as a result of protein aggregation when milk was treated at 90 °C for 5 min before cheese making. A decrease was observed in WHC values of all samples on day 14 indicating the disintegration of the structure during storage.

Viability of microbial populations

As shown in Table 3, count of Lactococcus ssp. and Lactobacillus ssp. populations on day 1 were determined highest in SM and K, respectively. The number of Lactococcus ssp. decreased during the storage time of all samples. Similarly, Lactobacillus ssp. count demonstrated a decrease in samples made from SM during the storage, whereas an increase was observed in cheeses made from K and YBM. The populations of total bacteria were similar in all cheeses in the first day of storage and each showed an increase during the storage period except for K35. However, the increase rate in cheeses treated at 100 °C was lower than the cheeses treated at 35 °C. Similar to this trend, the population of each microbial group were determined lower in the samples treated at 100 °C than the samples treated at 35 °C. This result may be explained by the protection effect of higher temperatures against bacteria. Furthermore, the low total bacteria number in K35 on day 14 could be most probably due to the competitive effect of the yeast and mold detected at a high number. Accordingly, the highest counts of yeast and mold were observed on the 14th day of storage for all samples. Coliform bacteria were not determined in any of the Quark samples.

Table 3.

Relative abundance of volatile compounds [µg/100 g] isolated from Quark cheeses (mean ± SD)

	SM35		SM100		K35		K100		YBM35		YBM100
	Day 1	day 14	day 1	day 14	day 1	day 14	day 1	day 14	day 1	day 14	day 1	day 14
Carboxylic acids
Acetic acid	466 ± 1.60C	914 ± 2.57C	nd	nd	6336 ± 13.4A	2925 ± 81.3B	1982 ± 109.1BC	936 ± 41.6C	nd	nd	nd	nd
Hexanoic acid	962 ± 46.4B	1098 ± 38.6AB	1055 ± 1.2AB	1178 ± 2.42A	305 ± 7.60D	395 ± 7.30D	575 ± 37.3C	1159 ± 23.9A	nd	nd	nd	nd
Octanoic acid	2223 ± 47.2AB	2576 ± 12.9A	380 ± 31G	630 ± 43.8FG	2083 ± 8.42BC	944 ± 155.9EF	1274 ± 8.64DE	1634 ± 8.43CD	nd	nd	nd	nd
3-methyl-butanoic acid	nd	nd	nd	nd	2912 ± 9.16A	3316 ± 4.90B	nd	nd	nd	nd	nd	nd
2-methyl-pentanoic acid	988 ± 1.95C	1802 ± 2.12B	1005 ± 2.1C	1920 ± 3.1A	nd	nd	251.7 ± 1.13E	436 ± 0.9D	nd	nd	nd	nd
Alcohols
3-methyl-1-butanol	nd	nd	nd	1025 ± 2.0C	4875 ± 21.4A	2519 ± 23.5B	620 ± 13.6C	2541 ± 25.6B	nd	nd	nd	nd
Phenethyl alcohol	nd	nd	nd	303 ± 0.6C	2730 ± 42.2B	10974 ± 80.4A	882 ± 34.7C	2553 ± 6.67B	nd	nd	nd	1262 ± 1.15BC
Ethyl alcohol	nd	nd	nd	nd	191 ± 5.30D	382 ± 13.2C	993 ± 2.1B	1716 ± 4.60A	nd	nd	nd	nd
2,3-Butanediol	nd	nd	nd	nd	600 ± 2.80A	318 ± 11.9C	nd	501 ± 24.2B	nd	nd	nd	nd
Esters
Ethyl acetate	nd	350 ± 5.60D	nd	nd	465 ± 3.20D	1457 ± 2.19C	2710 ± 22.3B	15848 ± 32.2A	nd	nd	nd	nd
Ethyl octanoate	nd	nd	nd	nd	1054 ± 55.5B	2402 ± 17.9A	130 ± 1.18D	598 ± 3.30C	nd	nd	nd	nd
Phenethyl acetate	nd	999 ± 3.73C	nd	nd	1819 ± 143.7C	10374 ± 408.4A	1182 ± 29.2C	3727 ± 71.2B	nd	nd	nd	nd
Ketones
2-Nonanone	1167 ± 2.1A	290 ± 2.8D	545 ± 4.10C	238 ± 1.94DE	791 ± 14.6B	580 ± 5.95C	184 ± 1.98E	102 ± 1.60F	nd	nd	nd	nd
2-butanone, 3-hydroxy	nd	nd	714 ± 2.26C	221 ± 1.61D	1190 ± 13.6B	579 ± 3.72CD	261 ± 8.90D	nd	2761 ± 8.90A	1213 ± 7.75B	2055 ± 9.8A	993 ± 3.6B
2,3-butanedione	nd	nd	nd	nd	nd	nd	nd	nd	4328 ± 6.97A	3215 ± 5.67B	2888 ± 4.65B	1675 ± 2.29B
Aldehydes
Acetaldehyde	nd	nd	nd	nd	nd	nd	nd	nd	231 ± 6.50D	534 ± 7.30C	966 ± 2.2A	1143 ± 1.22B
Benzaldehyde	16477 ± 81.2A	198 ± 46.2B		nd	nd	nd	nd	nd	905 ± 8.14B	nd	682 ± 2.3B	813 ± 2.70B
Hydrocarbones
Styrene	nd	nd	nd	189 ± 2.64C	3128 ± 15.6A	3601 ± 31.0A	3652 ± 8.30A	1464 ± 10.15B	nd	nd	nd	nd
Toluene	nd	nd	nd	nd	nd	1977 ± 10.4B	nd	3374 ± 14.3A	nd	nd	nd	nd

^A–GMeans with different uppercase letters significant differences between values of samples and storage days for each property (P < 0.05)

*Non-lettering columns or rows indicates the differences between scores are not found significant (P > 0.05)

Peptide profile

RP-HPLC peptide fractions of Quark cheeses at the 1st and 14th days of storage are shown in Fig. 2. Analyses were performed in a totally 85 min. While the peaks eluted in the early stage (first 30 min) of chromatogram correspond to hydrophilic peptides, peaks eluted at the later retention times (55–85 min) demonstrated the hydrophobic peptides. Furthermore, the intermediate time from 30 to 55 min corresponds to low molecular weight hydrophobic peptides. Fragmentation of proteins or large peptides into smaller ones and amino acids is seen with a decrease in the height/area of hydrophobic peaks and an increase of that in hydrophilic peaks (McSweeney 2004).

Fig. 2.

Peptide profiles of quark cheeses

In terms of peptide profile, qualitative and quantitative differences were observed both among sample types and over the course of storage. Moreover, the temperature degree applied to the raw materials during productions revealed a pronounced difference in the peptide profiles of all samples (Fig. 2). 35 °C were determined to be more convenient than 100 °C for the sample of SM in which the mesophilic lactic cultures were used with regard to both the numbers and the areas of peaks. However, for the samples of K and YBM an increase was observed in the number of peptides in the hydrophilic and intermediate regions, and a decrease was seen in the hydrophobic region. The observed changes in the hydrophilic and hydrophobic regions is the indicator of the hydrolysis of proteins or larger peptides into smaller ones and amino acids.

Peptide profiles of K includes more various peaks both in the sample produced at 35 °C and 100 °C. The reason of this situation could be related to the diversity of microorganisms available in kefir. Microorganism types are directly affect the peptide profiles of samples in terms of their varied peptidase activities. Furthermore, the reason of the decrease in the number and the amount of peptides in the samples subjected to 100 °C might also be the inhibition of peptidase activities at high temperatures. Similarly, McSweeney et al. (1993) found qualitative and quantitative differences between the peptide profiles of raw and heat-treated milk cheeses.

The areas of peaks eluting at the same time demonstrated an increase in all samples after 14 days of storage. Moreover, new peaks were occurred during the storage period. This formation is clearly seen in the sample of YBM100. Peptide profiles of samples treated at 100 °C also have higher protein contents which could also be a factor of the increased peptide areas and numbers.

According to the results, the variation of peptide profiles of samples depending on raw material, time or temperature indicates differences in the peptidase activity of different cultures present in different samples.

K35 contains much more peptide number than SM100, and an increase was observed in the areas of peaks after the storage time of 14 days. Similar results were obtained for K and YBM. In all samples treated at 35 °C the extent of proteolysis were obviously seen in the early hydrophilic region of chromatograms with the increased number of peptides (peaks) and lack or decreased area of peptides in the hydrophobic zone, most probably because of the vitality and the activity of the microorganisms at this temperature norm. Similar to our results, Amani et al. (2016) confirmed that lower proteolytic activity caused a higher WHC in yoghurt samples while during storage due to the increase in proteolytic activity and disintegration of the structure samples showed lower WHC.

The least peak numbers and areas were belong to samples of SM in which the commercial mesophilic lactic acid bacteria were used. Moreover, much more loss of activity against the high temperature were observed in these samples. As a result we can conclude alternatively produced products such as K and YBM are more rich in peptide profile.

Studies demonstrated when α-, β- and κ-caseins are subjected to temperatures of 100 °C or higher showed an increase in the newly formed peptides which were generated from the fragmentation of caseins (Meltretter et al. 2008). Similar results were found by Morales and Jimenez-Perez (1998) about the two newly formed peptides after heating of milk. Contrary to these findings, Grappin and Beuvier (1997) reported the milk pasteurization leads to a significant decrease in the amount of peaks in hydrophilic region such as small peptides and amino acids which obtained from HPLC profiles.

Volatile flavor compounds

Table 3 shows the volatile flavor compounds that mainly differentiated the Quark cheeses made from SM, K, and YBM treated at different temperatures. These volatiles contain carboxylic acids, alcohols, ketones, esters, aldehydes and hydrocarbones. Mostly, all the groups demonstrated a increase during storage, except for ketones. Alcohols were identified as the most abundant class for K35 and K100. The levels of alcohols tended to increase until the 14th day (P < 0.05). However, acids revealed an increasing abundance for both the products of SM and K for the same period. Esters were also found in higher amounts.

Both carboxylic acids and alcohols detected higher in K35 than K100. This shows the inhibition effect of higher temperature on beneficial microorganisms and the prevention of metabolite production. However, in terms of the product types, Quark cheeses made from kefir were found as having the richest volatile profile, most probably depending on the raw material which includes a mixed and varied microbiota. This result accords with another study (Beshkova et al. 2003) which found the compounds of acetaldehyde, ethyl acetate, 2- butanone, diacetyl, and ethanol in the volatile profile of kefir.

Among carboxylic acids, particularly, short chain fatty acids such as acetic acid, hexanoic acid and octanoic acid were determined in high concentrations in the samples of SM and K, which shows the hydrolysis of triglycerides by microbial enzymes and formation of the fatty acids. Lipases from fungal strains such as yeasts are playing an important role during these reactions. This can be explained by the higher abundance of short chain fatty acids in K35 and K100, in which the raw material of kefir includes yeast species. Due to the low detection of short-chain fatty acids, they are known as significant contributors of aroma formation in cheeses (Tavaria et al. 2004). Among linear chain fatty acids, hexanoic acid were also found to be predominant components of aroma similar to other cheese types such as Torta del Casar (Delgado et al. 2010), traditional Greek Xinotyri cheese (Bontinis et al. 2012) and Teleme cheese (Massouras et al. 2006). Having a pungent odour and taste, acetic acid is usually formed by the activity of lactic acid bacteria, and provides a specific aroma to brine-cured cheeses including Feta and Domiati (Hayaloglu and Karabulut 2011). 3-methyl-butanoic (isovaleric) acid and 2-methyl-pentanoic acid were the branched-chain fatty acids, that were identified in samples. 3-methyl-butanoic acid, which gives a rancid cheese-like flavour to some kind of cheeses (Yvon and Rijnen 2001) was also determined in K35.

Alcohols were only measured in the samples K. The use of kefir which contain yeast species is likely to be the reason of this situation. Alcohol concentrations detected in the products of K could originate from the activity of lactic acid bacteria (lactococci, lactobacilli), enterococci and yeast which are found in the microbial populations of kefir (Wang et al. 2012). Particularly, yeasts are playing an important role in kefir fermentation and produce ethanol and carbon dioxide. Kefir grains contain some yeasts that are fermenting lactose such as Kluyveromyces lactis, Kluyveromyces marxianus, Torula kefir. The higher ratio of alcohols in K35 and K100 pointed out that the presence of yeasts arising from kefir and their activity in these products. Ethyl alcohol was particularly determined as the main alcohol in Brie, Gorgonzola, Camambert and Blue cheeses which are ripened by some fungal species (Bontinis et al. 2012). Secondary alcohols such as 2-phenylethanol (phenethyl alcohol), 2,3-butanediol were also determined in products.

2,3 butanediol is formed from 2,3-butanedione (diacetyl) by the bacterial enzymes. 2-butanone, 3-hydroxy (acetoin) is the midproduct of this reaction. Acetoin was determined in all samples. It is thought to be produced from lactose, pyruvate or citrate by LAB (Di Cagno et al. 2003) at the production stage. 2,3 butanedione, which provides an intensely buttery flavour to products, was only detected in the samples of YBM. However, the other ketones such as 2-butanone, 3-hydroxy and 2-nonanone were determined in all products. The levels of acetoin and diacetyl tended to decrease, most probably because of the formation of 2,3 butanediol from these compounds. Furthermore, the decrease in the amount of acetoin during the storage could be the reason of the reduction observed in lactic acid bacteria. Methyl ketones are also generated by the enzymatic activity of fungal strains (Molimard and Spinnler 1996) and have low perception threshold with a specific odour. They were particularly detected in cheeses such as Camambert and Blue cheeses (Di Cagno et al. 2003).

Mainly ethyl esters were determined in samples. Esters are generated by the esterification of short-chain fatty acids with alcohols (Delgado et al. 2010). Similar to ketones, esters also have a siginificant effect on the aroma of dairy products (Molimard and Spinnler 1996). The increase observed in the concentrations of esters are due to the continuous esterification reaction of short chain fatty acids with alcohols.

Similar to 2,3 butanedione, acetaldehyde from aldehydes group was also identified merely in the products of YBM. The best perceived ‘odour-flavour and taste’ property of Quark cheese made from YBM is most probably provided by the components that were not detected in other samples such as acetaldehyde and 2,3-butanedione, which are usually typical for yoghurt. Ott et al. (2000) also reported that acetaldehyde was the main carbonyl compound that makes a contribution to yoghurt aroma.

Sensory evaluation

Figure 3 shows the results of sensory evaluation of Quark samples. Production of cheeses from different raw materials resulted in variations on judgement preferences for sensorial attributes of the samples. Panelists preferred K35 and YBM100 in terms of color-appearance and body-texture, respectively on day 1. Whereas, Quark cheese made from YBM100 and K100 were the best perceived products with respect to odor-flavor-taste on the same day. This was probably due to the inhibition of unwanted metabolite producer microorganisms which changed the flavour by producing different metabolites during storage. Panelists also indicated a creamy and a mild acidity taste belong to YBM100 that made it the most preferable sample. K35 had an attraction for its color-appearance on the first day, but when storage progressed, the other samples were preferred by panelists more than K35. Furthermore, panelists notified to perceive a slight bitterness with the taste of K35. Odour-flavor-taste scores of K35 were determined lower than all the other samples (P < 0.05). This could be due to the fragmentation of proteins to smaller peptides and aminoacids which is stimulated by the varied microorganisms present in K35. This result also observed with the more peak numbers in the early retention time of the HPLC chromatogram of K35 (Fig. 2). In addition to this, regardless of the raw material, all samples produced at 35 °C had the lowest scores for both color-appearance and body-texture on day 14 because of the whey separation formation. Odour-flavor-taste scores of all 6 samples decreased with the prolonged of storage time, whereas the scores for body-texture increased up till the end of storage for the samples produced at 100 °C.

Fig. 3.

Sensory properties of quark cheeses

Conclusion

The results demonstrate that alternatively produced products of K and YBM had advantages over SM in terms of investigated properties. Both types of raw materials enhanced not only the flavor and taste but also the texture of samples. Each property was found to be related both to the applied temperature and type of raw material. For all product types, samples produced at 100 °C were preferred to samples produced at 35 °C in terms of microbiological, physicochemical and sensory properties. The usage of either K or YBM resulted in a higher TS content and WHC values. The temperature of 100 °C had a favourable detractive effect on the total microbial counts during storage when compared to cheeses treated at 35 °C. Due to the presence of diverse microorganisms in kefir, both samples of K showed a developed proteolysis which is similar to the trend determined in the proteolysis level of samples obtained at 35 °C for all three product types. Products made from K100 and YBM100 were the favorite products in terms of odour-flavour and taste, probably due to the detected flavour compounds such as 2,3-butanedione, acetaldehyde and ethanol which were not detected in SM. K and YBM also showed a more rich peptide profile which could be the reason of the slight bitterness, perceived particularly during the taste of K. In general, samples produced at 100 °C were preferred to samples produced at 35 °C with regard to texture, flavor and taste. As a conclusion, Quark cheese could be produced from kefir and yayik buttermilk instead of cultured milk in order to develop the sensorial, technological and functional product characteristics.