Antioxidant activity of labneh made from cashew milk and its combination with cow or camel milk using different starter cultures

Abstract

The aim of this study is to investigate the effects of three strains of probiotic Lactobacillus spp. i.e. L. plantarum (S1) L. casei (S2), and/or L. rhamnosus (S3) in co-cultures with Streptococcus thermophiles (St) and L. delbrueckii subsp. lactis (Ll) on the changes of total phenolic and flavonoid contents, and antioxidant activity of concentrated yogurt (labneh; L) made from individual cashew milk (EwM; 100:0 v/v) mixed with/without cow (Co) or camel (Ca) milk (75:25 v/v) during 0, 7, 14, and 21 days of storage. EwML100% S3 showed the highest total phenolic and flavonoid contents during 21 days of storage. Radical scavenging activity was improved in Ew-/Ca-ML (S2, S3, and S4) compared to control (containing St and Ll) on the 0th day. The maximum ferric reducing potential was observed in fresh Ew-/Ca-ML75% S1 (1.6 ± 0.05 mM Fe + 2 E/mL). All the starter cultures enhanced (p < 0.05) the chelating ability of EwML compared to control during the 7th and 14th day of storage. In conclusion, cashew milk labneh mixed with/without cow or camel milk containing probiotic Lactobacillus spp. can strengthen the health-promoting properties with antioxidant activity.

Introduction

Antioxidants are compounds that inhibit or delay the onset of oxidation and may be classified as natural or synthetic (Alenisan et al. 2017). There is an increasing demand for natural antioxidants due to the harmful and carcinogenic effects of synthetic antioxidants. Several studies have found that eating foods high in natural antioxidants can reduce the risk of malignancies, hypertension, diabetes, and cardiovascular diseases, particularly in underdeveloped countries where most people have limited finances and access to contemporary treatments (Mitterer-Daltoe et al. 2020; Macharia et al. 2022; Tain and Hsu 2022; Shori 2022).

Camel milk is a vital food supply for rural people in arid parts of Africa and the Middle East, where it may be consumed fresh or as fermented products (Shori 2017). There are around 28 million camels globally, and they produce about 3 million metric tonnes of milk each year (Polidori et al. 2021). The functional value of camel milk is greater than that of cow's milk. Dairy products derived from camel milk, including pasteurized milk, milk powder, fermented liquid milk, and cheese, are now widely available (Polidori et al. 2021). According to Alnohair (2021), camel milk has been used to treat a variety of metabolic and immunological disorders.

In recent years, the food industry has taken the lead in developing novel plant-based milk products. Alternatives to animal milk such as cashew milk are becoming increasingly popular among consumers (Olatidoye et al. 2020). Cashew milk is a good substitute for animal milk to produce certain dairy products. Cashew milk is used as a source of protein (5%), carbohydrates (5.95%), and fat (5.49%; Tamuno and Monday 2019). In addition, cashew milk contains calcium, potassium, and iron (4.75, 7.15, and 3.0 mg/100 g, respectively). Magnesium and phosphorus contents in cashew milk are about 2.0 mg/100 g each (Tamuno and Monday 2019).

Dairy products have been considered healthy and natural and when consumed regularly, it can actually prevent disease (Shori et al. 2022d). The buffering capacity of milk ensures microbial viability during fermentation and refrigerated storage. Therefore, fermented dairy products could be a potential vehicle for delivering probiotic bacteria to customers (Gil et al. 2019). Concentrated yogurt, also known as labneh, is an excellent source of proteins and minerals (Al-Rimawi et al. 2020). Labneh can be made with a variety of milk, including cow, sheep, and goat milk (Serhan et al. 2016). Labneh is a semi-solid creamy textured product with an acidic taste, containing about 23–25% (w/w) of total solid content (Al-Rimawi et al. 2020). Labneh was found to have superior nutritional and medicinal properties compared to yogurt. It contains 2.5 times higher protein content, 50% more minerals, and much more lactic acid bacteria (LAB) than regular yogurt (Binda and Ouwehand 2019‏). Labneh is suitable for lactose-intolerant individuals because it has a low lactose content of ~ 6% (Al-Rimawi et al. 2020). Therefore, the aim of this study is to investigate the effects of three strains of probiotic Lactobacillus spp. (i.e. L. rhamnosus, L. casei, and/or L. plantarum) in co-cultures with Streptococcus thermophilus and L. delbrueckii subsp. lactis on the changes of total phenolic and flavonoid contents, and antioxidant activity of concentrated yogurt (labneh) made from individual cashew milk (100:0) mixed with/without cow or camel milk (75:25) during 0, 7, 14, and 21 days of storage.

Materials and methods

Materials

Pure strains of S. thermophilus St1342 and L. delbrueckii ssp. lactis ATCC 7830, L. plantreum ATCC 14917, L. casei (393) and L. rhamnosus ATCC 53103 were obtained from the Microbiological Resources Center (Cairo mircen) in Ain Shams University. All strains of bacteria were grown in selected media under anaerobic conditions at 37 ± 2 °C for 48 h. Cashew nuts, cow milk (Almarei®), and camel milk (Al-turath®) were purchased from a local food store. Anhydrous sodium carbonate, gallic acid (GA), (+)-catechin hydrate (CH), ethanol, 2,2-diphenyl-1-picrylhydrazyl (DPPH), folin-Ciocalteu reagent, potassium ferricyanide, ferrous chloride, ferric chloride, ferrozine, phosphate buffer, trichloroacetic acid (TCA), sodium nitrite, sodium hydroxide, and aluminum chloride were purchased from Sigma-Aldrich, St. Louis, MO, USA.

Methods

Preparation of starter culture

Streptococcus thermophilus, L. casei, L. plantreum, L. rhamnosus, and L. delbrueckii ssp. lactis were all stored at − 80 °C. Sterile 10 mL aliquots of MRS broth inoculated with 1% (v/v) of each strain individually were incubated at 37 °C except for L. delbrueckii ssp. lactis which was incubated at 42 °C (Shori et al. 2022b). The activated organisms after three successive transfers were used to prepare starter cultures for labneh making. The cultures were prepared by inoculating 1% (v/v) in 10 mL aliquots of sterile reconstituted skim milk (72 °C for 15 s) supplemented with 2% glucose and 1% yeast extract.

Preparation of cashew extract

Cashew nuts (10 g) were soaked in 100 mL of distilled water for 12 h (Shori et al. 2022b). The mixture was blended and filtered using a clean Muslin cloth, followed by centrifugation (4080 g, 4 °C) for 15 min. The supernatant was harvested and pasteurized at 90 °C for 5 min. The clear solution was refrigerated (4 °C) and used within 3 days as cashew milk in the making of labneh.

Preparation of labneh

Labneh was prepared according to Serhan et al. (2016) with some modifications. Labneh (L) was prepared from individual cashew milk (EwM) mixed with/without cow milk (CoM) or camel milk (CaM) in two different concentrations (100:0 and 75:25; v/v). In addition, 2% of starter culture (S) including S1 = L. plantarum (Lp), S2 = L. casei (Lc), S3 = L. rhamnosus (Lr), and S4 = (Lr, Lc, and Lp) in co-cultures with S. thermophilus (St) and L. delbrueckii subsp. lactis (Ll) was added to each formula. Then, all the mixtures were mixed thoroughly and incubated at 41 °C for 3 h. The prepared yogurt was centrifuged (4080 g; 4 °C) for 10 min and the whey was discarded. The solid product was collected namely (labneh) and stored at 4 ± 1 °C for 0, 7, 14, and 21 days. Labneh (control) containing co-cultures (St and Ll) was prepared in the same manner using EwM mixed with/without CoM or CaM in a ratio of 100:0 and 75:25 (v/v).

Preparation of labneh water extract

Ten grams of each labneh sample was homogenized with 2.5 mL dH₂O (Shori et al. 2018). The samples were acidified to pH 4.0 by adding HCl (0.1 M) and incubated in a water bath at 45 °C for 10 min. The samples were centrifuged (5000 g, 4 °C) for 10 min and the supernatant was then adjusted to pH 7.0 using NaOH (0.1 M). This was followed by second centrifugation (5000 g, 4 °C) for 10 min and the clear supernatant was used for further analysis.

Total phenolic content (TPC)

2N Folin-Ciocalteu reagent (0.5 mL; 50% v/v) was added to 5 mL dH₂O, 1 mL of 95% ethanol, and 1 mL of labneh water extract or gallic acid standard solutions (10–60 μg/mL; Shori 2013). The mixture was kept at room temperature for 5 min before 1 mL of sodium carbonate solution (5% w/v) was added to the sample. The mixture was left in the dark at 25 °C for 1 h and the absorbance was measured at 725 nm using a spectrophotometer (Shimadzu UV Mini 1240, Duisburg, Germany). The total phenolic content was estimated against the gallic acid standard curve and expressed as µg gallic acid equivalent per gram fresh labneh (µg GAE/g fresh labneh).

Total flavonoid content (TFC)

The total flavonoid content was determined as described by Bouterfas et al. (2016). Labneh sample (250 μl) was mixed with 1.25 mL dH2O and 75 μl sodium nitrite for 5 min. Then, 150 μl of 10% aluminum chloride, 0.5 mL of 1 M sodium hydroxide, and 275 μl of dH2O were added to the mixture. The absorbance was read at 510 nm and a standard curve was prepared from a serial dilution (0–500 μg/mL) of catechin. The total flavonoid content was estimated against the catechin standard curve and expressed as µg catechin equivalent per gram fresh labneh (µg CE/g fresh labneh).

Determination of antioxidant activity

DPPH radical scavenging activity assay

DPPH reagent (3 mL; 60 mM) dissolved in 95% ethanol was added to 250 μl of labneh water extract (Muniandy et al. 2016). The mixture was incubated in the dark at room temperature for 1 h. Absorbance (Abs) at 517 nm was measured against control (250 μl of ethanol instead of the extract) using a spectrophotometer (Genesys 10 UV–vis, Thermo Fisher Scientific, Inc., USA). The DPPH radical scavenging activity was calculated as below:

$$\text{Radical scavenging activity}\left(\%\right)=\left({\text{Abs}}_{\text{control}@\text{517nm}}-{\text{Abs}}_{\text{sample}@\text{517nm}}/{\text{Abs}}_{\text{control}@\text{517 nm}}\right)\times 100$$

Ferric reducing antioxidant potential (FRAP) assay

FRAP reagent (3.6 mL) containing 300 mM acetate buffer, 8 mM 2,4,6-tri(2-pyridyl)-s-triazine (TPTZ) reagent, and 20 mM FeCl₃ solutions at a ratio of 10:1:1 was mixed with 400 μl of labneh water extract or iron(II) sulfate heptahydrate (FeSO₄·7H₂O) standard solutions (0.3–1.0 μg/mL; Muniandy et al. 2016). The mixture was incubated in a water bath (37 °C) for 10 min before absorbance was read at 593 using a spectrophotometer (Genesys 10 UV–vis, Thermo Fisher Scientific, Inc., USA). The results were estimated against FeSO₄·7H₂O standard curve and expressed as mM Fe²⁺ equivalent/mL (mM Fe²⁺ E/mL).

Ferrous ion chelating (FIC) ability assay

The 2 mM iron(II) sulfate hydrate (FeSO₄·xH₂O) solution and 5 mM ferrozine solution were diluted 20 times prior to use (Shori 2020). Then, 1 mL of diluted FeSO₄·xH₂O solution was mixed with 1 mL of labneh water extract and 1 mL of diluted ferrozine solution. This was followed by incubation for 10 min at 25 °C. Absorbance (Abs) at 562 nm was measured against control (1 mL of dH₂O instead of the extract) using a spectrophotometer (Genesys 10 UV–vis, Thermo Fisher Scientific, Inc., USA). The FIC ability of samples was calculated as below:

$$\text{FIC ability}\left(\%\right)=\left({\text{Abs}}_{\text{control}@\text{562nm}}-{\text{Abs}}_{\text{sample}@\text{562nm}}/{\text{Abs}}_{\text{control}@\text{562 nm}}\right)\times 100$$

Statistical analysis

All data were presented as means ± standard error (SE) and performed in duplicates for three batches (n = 3). IBM SPSS statistics software version 20.0 was used and one-way analysis of variance (ANOVA) as well as the significance of the mean differences were determined using Duncan's test (p < 0.05).

Results

Total phenolic and flavonoid contents of labneh

Table 1 shows the TPC and TFC in labneh made from individual cashew milk (100%) mixed with/without cow or camel milk (75:25) using three types of probiotics and their combination compared to control during 21 days of storage. Control labneh made from cashew milk showed significantly (p < 0.05) higher phenolic content (34.70 ± 1.90 μg GAE/g fresh labneh), followed by cashew and camel milk labneh (25.06 ± 4.70 µg GAE/g fresh labneh) at day 0 (Table 1). Comparatively, the presence of starter cultures S1, S2, S3, and S4 in all labneh samples displayed an increase (p < 0.05) in TPC compared to their respective controls during 21 days of storage. The higher phenolic content (p < 0.05) in all labneh samples was shown in the presence of S3 during the storage period (Table 1). All labneh samples showed a maximum increase (p < 0.05) in TPC on the 14th day, which was about twofolds, threefolds, and fourfolds higher than day 0 for all EwML, Ew-/Ca-ML, and Ew-/Co-ML; respectively. A non-significant reduction was noticed in all labneh samples on the 21st day (Table 1). However, control labneh made from individual cashew milk mixed with/without cow or camel milk showed a significant (p < 0.05) reduction in TPC at the end of storage.

Table 1.

Total phenolic (µg GAE/g fresh labneh) and flavonoid (µg CE/g fresh labneh) contents of labneh (L) made from individual cashew milk (100%) and its mixture with cow or camel milk (75:25) in the presence of starter cultures Lactobacillus plantarum (S1), L. casei (S2), and L. rhamnosus (S3) and their combination (S4) compared to control during 21 days of storage at 4 °C

Samples	Total phenolic content (µg GAE/ g fresh labneh)				Total flavonoid content (µg CE/g fresh labneh)
Day	0	7	14	21	0	7	14	21
EwML (control)*	34.7 ± 1.90a	43.7 ± 9.00a	62.5 ± 1.19a	43.7 ± 0.10a	30.5 ± 0.30a	38.2 ± 0.80a	41.5 ± 10.6a	24.8 ± 1.80a
EwML S1	42.2 ± 0.14b	54.15 ± 0.10b	92.7 ± 0.50b	90.7 ± 0.10b	33.5 ± 0.21a	50.0 ± 0.90b	75.9 ± 2.30b	40.9 ± 0.31b
EwML S2	41.4 ± 0.20b	50.5 ± 3.50b	88.8 ± 5.10b	92.6 ± 4.88b	31.6 ± 0.07a	40.7 ± 0.90a	42.4 ± 4.20a	37.9 ± 0.49b
EwML S3	49.2 ± 2.80c	71.0 ± 0.39c	107.7 ± 7.60c	100.6 ± 2.81b	40.4 ± 0.14b	69.6 ± 0.87c	106.4 ± 1.13c	85.5 ± 1.13c
EwML S4	42.3 ± 1.80b	68.7 ± 0.39c	97.0 ± 0.50b	94.7 ± 0.77b	33.9 ± 0.4a	69.9 ± 1.06c	87.6 ± 0.07d	77.7 ± 4.45c
Ew-/Co-ML (control)*	20.5 ± 2.60a	31.6 ± 0.10a	58.5 ± 1.10a	32.4 ± 4.30a	19.8 ± 0.14a	24.4 ± 0.14a	28.7 ± 3.40a	21.1 ± 0.09a
Ew-/Co-ML S1	22.15 ± 12.7a	52.3 ± 1.50b	86.7 ± 1.60b	80.23 ± 0.82b	25.1 ± 0.80b	32.8 ± 0.49b	55.5 ± 2.60b	29.9 ± 0.3b
Ew-/Co-ML S2	21.4 ± 4.03a	50.8 ± 0.60b	84.5 ± 1.60b	88.5 ± 1.10b	24.9 ± 0.18b	30.3 ± 0.14b	38.1 ± 0.87c	24.7 ± 2.40c
Ew-/Co-ML S3	24.5 ± 0.70b	54.5 ± 0.70b	102.7 ± 0.50c	92.5 ± 1.19b	33.5 ± 0.21c	55.4 ± 0.25c	99.5 ± 0.14d	80.0 ± 1.13d
Ew-/Co-ML S4	23.2 ± 1.55b	53.32 ± 0.11b	91.2 ± 46.8b	84.5 ± 0.60b	27.1 ± 0.60b	58.2 ± 1.10c	67.9 ± 1.06e	44.4 ± 0.14e
Ew-/Ca-ML (control)*	25.06 ± 4.70a	35.7 ± 2.00a	61.6 ± 1.10a	38.7 ± 6.50a	21.3 ± 1.16a	27.5 ± 0.21a	31.7 ± 0.20a	23.9 ± 0.76a
Ew-/Ca-ML S1	26.9 ± 1.90a	52.3 ± 1.09b	90.8 ± 6.40b	86.2 ± 3.40b	26.2 ± 1.60b	34.0 ± 0.63b	75.2 ± 5.20b	30.4 ± 0.14b
Ew-/Ca-ML S2	25.95 ± 2.33a	45.2 ± 1.70c	85.6 ± 4.93b	91.13 ± 0.09b	26.8 ± 5.16b	33.4 ± 0.95b	40.4 ± 0.98c	28.9 ± 2.40b
Ew-/Ca-ML S3	30.0 ± 6.08b	68.6 ± 1.09d	105.6 ± 6.12c	94.5 ± 1.40b	34.4 ± 0.14c	67.9 ± 0.77c	102.6 ± 1.50d	81.5 ± 1.06c
Ew-/Ca-ML S4	29.7 ± 4.90b	65.7 ± 3.30d	94.8 ± 1.80c	87.9 ± 9.10b	29.7 ± 2.60b	67.7 ± 6.40c	85.1 ± 0.42e	65.6 ± 1.06d

Data are presented as mean ± SEM. M

a,b,c,d means with different superscript letters in a column are significantly different (p < 0.05)

Cow- = Co, camel- = Ca, cashew- = Ew, milk = M, labneh = L, starter culture = S, Lactobacillus plantarum = 1, L. casei = 2, L. rhamnosus = 3, and 1 + 2 + 3 = 4

*Control containing Streptococcus thermophilus and L. delbrueckii subsp. Lactis

The presence of starter cultures S1, S2, S3, and S4 increased (p < 0.05) TFC in fresh labneh made from 100% cashew milk (34–40 µg CE/g fresh labneh) compared to control (30.5 ± 0.30 µg CE/g fresh labneh; Table 1). However, only starter cultures S3 and S4 enhanced (p < 0.05) TFC in Ew-/Co-ML and Ew-/Ca-ML compared to their respective controls on the 0th day (Table 1). In addition, the increase in the TFC of labneh samples was influenced by the starter cultures used during the storage period (Table 1). In cashew milk labneh mixed with camel milk, S3 had the highest TFC (p < 0.05) followed by S4 as compared to other samples during the 7th and 14th day of storage. In addition, all labneh samples showed the highest TFC (p < 0.05) on the 14th day with maximum values shown in the presence of S3. However, all labneh samples showed no significant decline (p > 0.05) in TFC on the 21st day compared to their respective controls, which showed a sharp decrease (p < 0.05).

Antioxidant properties

DPPH radical scavenging activity of labneh

Figure 1 shows the DPPH radical scavenging activity in labneh made from individual cashew milk (100%) mixed with/without cow or camel milk (75:25) using three types of probiotics and their combination compared to control during 21 days of storage. EwML S1, S2, and S3 showed the highest radical scavenging activity (95–93%; p < 0.05) on the 0th day as compared to control (83.7 ± 0.40%; Fig. 1a). The presence of starter cultures (S1–S4) in cashew milk labneh did not significantly improve (p > 0.05) radical scavenging activity compared to control during 21 days of storage at 4 °C. Ew-/Co-ML (control) showed radical scavenging activity of 82.9 ± 0.11% on the 0th day (Fig. 1b). However, the presence of starter cultures increased (p < 0.05) the activity in Ew-/Co-ML between 90 and 93%. A slight reduction (p < 0.05) of radical scavenging activity was observed in Ew-/Co-ML S2 and S3 (82.1 ± 0.07 and 85.6 ± 0.05; respectively) compared to control (91.0 ± 0.02%) on the 7th day. Ew-/Ca-ML (S2, S3, and S4) showed significantly higher radical scavenging activity (94%; p < 0.05) than control (86.1 ± 0.90%) on the 0th day (Fig. 1c). Ew-/Ca-ML S1 showed a significant reduction (p < 0.05) in radical scavenging activity compared to control on the 7th day but increased (p < 0.05) on the 14th day. Radical scavenging activity of Ew-/Ca-ML S1 and S2 was reduced by 11% (p < 0.05) compared to control at the end of storage (Fig. 1c).

Fig. 1.

Changes in radical scavenging activity of labneh (L) made from cashew milk (EwM; 100%; A) and its mixture with cow (Co; B) or camel (Ca; C) milk (M; 75:25) in the presence of starter cultures Lactobacillus plantarum (S1), L. casei (S2), and L. rhamnosus (S3) and their combination (S4) compared to control during 21 days of storage at 4 °C. Data are presented as mean ± SEM. The level of significance was preset at p < 0.05 compared to control at the same storage period

Ferric reducing antioxidant potential (FRAP) of labneh

Figure 2 shows ferric reducing antioxidant potential of labneh made from individual cashew milk (100%) mixed with/without cow or camel milk (75:25) using three types of probiotics and their combination compared to control during 21 days of storage. EwML (control) showed ferric reducing potential of 0.602 ± 3.60 mM Fe⁺² E/mL on the 0th day (Fig. 2a). Maximum ferric reducing potential was observed in EwML S4 (0.920 ± 0.00 mM Fe⁺² E/mL), followed by EwMLS3 (0.810 ± 0.00 mM Fe⁺² E/mL) on the 0th day. EwML S3 and S4 showed a significant decline (p < 0.05) in ferric reducing potential on the 7th day, whereas EwMLS2 maintained ferric reducing potential over 2 weeks (Fig. 2a). The presence of starter cultures (S1–S4) in Ew-/Co-ML increased (p < 0.05) ferric reducing potential compared to control on the 0th day with Ew-/Co-MLS4 showing the highest ferric reducing potential (1.200 ± 0.05 mM Fe⁺² E/mL; Fig. 2b). Similarly, starter cultures (S1–S4) in Ew-/Ca-ML showed greater ferric reducing potential than controls on the 0th day with S1 showing maximum ferric reducing potential (1.600 ± 0.05 mM Fe⁺² E/mL; Fig. 2c). Nevertheless, a significant (p < 0.05) loss of ferric reducing potential was observed in both Ew-/Co-ML and Ew-/Ca-ML using different starter cultures during 21 days of storage (Fig. 2b and c).

Fig. 2.

Changes in ferric reducing antioxidant potential (FRAP) of labneh (L) made from cashew milk (EwM; 100%; A) and its mixture with cow (Co; B) or camel (Ca; C) milk (M; 75:25) in the presence of starter cultures Lactobacillus plantarum (S1), L. casei (S2), and L. rhamnosus (S3) and their combination (S4) compared to control during 21 days of storage at 4 °C. Data are presented as mean ± SEM. The level of significance was preset at p < 0.05 compared to control at the same storage period

Ferrous ion chelation (FIC) of labneh

Figure 3 shows the ferrous ion-chelating (FIC) of labneh made from individual cashew milk (100%) mixed with/without cow or camel milk (75:25) using three types of probiotics and their combination compared to control during 21 days of storage. As compared to other samples, EwML (control) had the highest chelating ability (76.00 ± 0.30%) on the 0th day, which was followed by a reduction (p < 0.05) to 63% on the 14th day and an increase to 99.70 ± 0.05% on the 21st day (Fig. 3a). All EwML samples showed higher (p < 0.05) chelating ability than control during the 7th and 14th day of storage. EwMLS4 showed a significant reduction (23.90 ± 0.10%; p < 0.05) in chelating ability on the 21st day of storage. The addition of starter cultures did not affect (p < 0.05) the chelating ability of Ew-/Co-ML compared to control during 21 days of storage (Fig. 3b). However, the chelating ability of 7-day-stored Ew-/Co-MLS3 and 21-day-stored Ew-/Co-MLS1 (74.40 ± 1.50 and 92.50 ± 0.40%; respectively) was almost similar to that of control. Ew-/Ca-ML (control) showed significantly (p < 0.05) higher chelating ability than other samples on the 0th day (Fig. 3c). The chelating ability of Ew-/Ca-ML S1 and S3 increased (p < 0.05) to 95.00 ± 1.10% and 96.40 ± 0.10%; respectively on the 7th day and maintained these levels for 21 days of storage. In addition, Ew-/Ca-MLS2 showed higher chelating ability (99.33 ± 0.20%; p < 0.05) than the control (85.40 ± 0.10%) on the 21st day (Fig. 3c).

Fig. 3.

Changes in ferrous ion-chelating (FIC) of labneh (L) made from cashew milk (EwM; 100%; A) and its mixture with cow (Co; B) or camel (Ca; C) milk (M; 75:25) in the presence of starter cultures Lactobacillus plantarum (S1), L. casei (S2), and L. rhamnosus (S3) and their combination (S4) compared to control during 21 days of storage at 4 °C. Data are presented as mean ± SEM. The level of significance was preset at p < 0.05 compared to control at the same storage period

Discussions

A number of scientific investigations have been conducted on the development and production of novel probiotic fermented foods with functional properties (Shori et al 2022c; Al-Sulbi and Shori 2022; Shori and Al zahrani 2022). Fermented foods are influenced by degradation, structural changes, and the development of new molecules that occur during fermentation which are related to several parameters such as microbial strains, substrate content, and fermentation conditions (Mikulajová et al. 2021). Lactic acid bacteria are frequently used during fermentation to ensure the safety and stability of the final products and to promote health benefits to the host (Liptakova et al. 2017).

Plant-based milk (PBM) products have been developed to offer an alternative to animal-based dairy products and have become increasingly popular (Montemurro et al. 2021). Despite a lengthy history of non-dairy products, the development of new PBM products has seen a surge in interest because of their health benefits (Shori et al. 2022a). In our analysis, labneh made from 100% cashew milk had higher phenolic and flavonoid contents than labneh made from cashew milk mixed with cow or camel milk (Table 1). The inherent phenolic and flavonoid compounds in cashew milk could be responsible for high phenolic and flavonoid contents in the labneh. The composition of phenolic acids in cashew extract included caffeic acid, p-coumaric acid, ferulic acid, gallic acid, syringic acid, naringenin, catechin, and quercetin (Różańska et al. 2021). In addition, flavonoids in cashew extract such as 3-O-galactoside, 3-O-glucoside, 3-O-rhamnoside, 3-O-xylopyranoside, 3-O-arabinopyranoside and 3-O-arabinofuranoside of quercetin and myricetin, as well as kaempferol 3-O-glucoside were also identified (Salehi et al. 2019). Further study is required to determine the TPC and TFC as well as identify the phenolic and flavonoid compounds profiles in cashew milk before and after fermentation. The reduction of phenolic and flavonoid contents in labneh made from cashew milk mixed with cow or camel milk could possibly occur as a result of the interaction of milk proteins with reactive phenolic compounds from the cashew milk. However, the amalgamation of cashew and camel milk during labneh preparation with different strains of lactobacillus probiotics showed promising results for total phenols, suggesting the greater availability of nutrients and milk proteins.

Shori et al. (2022b) reported that the growth and metabolism of LAB have been enhanced in the presence of cashew milk. To ensure high-quality products, it is important to select the right starter cultures. The present study found that the addition of three strains of Lactobacillus spp and their combination increased the phenolic and flavonoid contents in labneh samples compared to control throughout storage periods. In addition, labneh samples containing L. rhamnosus exhibited the highest TFC and TPC among other samples during 21 days of storage. This indicated that the addition of L. rhamnosus in individual cashew milk and its mixture with cow or camel milk during fermentation contributed to an increase in phenolic and flavonoid contents in labneh which can lead to potential health benefits such as antioxidants. A similar observation was also reported for cashew milk yogurt fermented with L. rhamnosus (Shori et al. 2022b). A study conducted on soy-based milk fermented with S. boulardii and probiotic Lactobacillus spp. showed an increase in flavonoid content (especially isoflavone) by 95% (Rekha and Vijayalakshmi 2010). In addition, Lactobacillus-fermentation of soymilk increased phenolic acids, isoflavone, and antioxidant activity (Chavan et al. 2018). This phenomenon has been described as a general impact of LAB-induced acidification, which increases phenol solubilization and extractability. In addition, some LAB produces certain enzymes like feruroyl esterases that promote the release of antioxidant phenols, thereby improving phenolic content (Verni et al. 2019). Currently, there are several species of LAB that have been proved to produce feruloyl esterase including L. johnsonii (Lai et al. 2009), L. plantarum (Esteban-Torres et al. 2013), Pediococcus acidilactici (Kaur et al. 2014), L. amylovorus (Xu et al. 2017), Bifidobacterium animalis subsp. lactis (Fritsch et al. 2017) and L. helveticus (Song and Baik 2017). In addition, it is reported that L. rhamnosus can produce much lower but active feruroyl esterases during the fermentation of wheat bran, rye bran, corn pectin, or brewer's spent grain to release phenolic acids (Szwajgier and Jakubczyk 2011). Further study is needed to investigate the ability of the three strains of probiotic Lactobacillus spp. to produce feruroyl esterases during labneh fermentation from individual cashew milk and its combination with cow or camel milk. Higher protein content and dietary fiber in cashew milk fermented with different LAB strains may be associated with a higher phenolic content (Tlais et al. 2022). The decrease in TFC of labneh samples at the end of storage may be explained by the action of the starter culture to further break down the polymeric phenolics in the cashew milk. A similar observation was reported in a previous study (Shori et al. 2022b).

It was reported that fruits, vegetables, and nuts could exhibit antioxidant properties due to their phenolic and flavonoids compounds (Shori 2015). Rice-Evan et al. (1997) reported that phenolic and flavonoid compounds have redox properties, which allow them to act as reducing agents, hydrogen donators, and singlet oxygen quenchers. Thus, the redox potential of these compounds plays an important role in determining the antioxidant capacity of the plant. A variety of tests are used to determine antioxidant activity (Malomo et al. 2020). DPPH and FRAP are the most widely used methods to assess the antioxidant activity of food products. Antioxidants are chemical compounds that neutralize and scavenge free radicals, constantly generated in the body (Shori et al. 2022b). It was shown that the antioxidant content of goat's milk yogurt was 93% higher than camel milk yogurt when Pediococcus pentosaceus was used as a starter culture. This finding implies that probiotic bacteria boost yogurt's antioxidant properties (Balakrishnan and Agrawal 2014). In our current study, the labneh made from cashew and camel milk combination showed the highest chelating ability when fermented with different starter cultures. FIC assay was used to evaluate metal ion binding by antioxidant components. Ew-/Ca-ML was found to possess the ability to prevent or inhibit reactions such as a Fenton's type reaction, which generates reactive hydroxyl radicals. In a previous study, tocopherols and tocopheryl acetate (vitamin E) were found to have antioxidant properties in cashew extract (Yang et al. 2009). Accordingly, the cashew milk labneh prepared in our study showed high antioxidant activity.

Conclusion

TPC and TFC were increased significantly in the presence of L. rhamnouses in all labneh samples during the two weeks of storage. The presence of starter cultures (S2, S3, and S4) in fresh Ew-/Ca-ML significantly increased radical scavenging activity. L. planterum enhanced ferric reducing potential in Ew-/Ca-ML. Cashew milk labneh mixed with/without cow or camel milk using three strains of Lactobacillus spp. and their combination has significantly boosted the total phenols and flavonoid content, as well as the antioxidant activity. The inoculation of L. plantarum, L. casei, and/or L. rhamnosus in labneh samples during storage could improve total phenolic and flavonoid content as well as act as a potential source of antioxidants. Therefore, labneh produced from cashew milk mixed with/without cow or camel milk using S1, S2, S3, or S4 can strengthen the health-promoting properties with anti-oxidant activity.

Authors' contributions

ABS Conception and design, drafting the article, critical review of important intellectual content, final approval of the version to be published. OSA-s Acquisition of data, analysis and interpretation of data, final approval of the version to be published.

Funding

This project was funded by Deanship of Scientific Research (DSR), King Abdulaziz University, Jeddah, under grant No. (IFPIP-17-247-1443). The authors, therefore, gratefully acknowledgment the DSR technical and financial support.

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The authors have no conflicts of interest to declare.

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