Enrichment of apricot probiotic drink with sea buckthorn as a prebiotic and antioxidant source

ABSTRACT

Supplemented apricot probiotic drink (SAPD) was prepared with the addition of 2% lactobacillus rhamnosus and different percentages (2, 4, 8, 12%) of sea buckthorn powder, a total of five treatments were prepared To, T₁, T₂, T₃, and T₄ and stored at 4°C. The analysis of probiotic count was performed using total plate count and for the examination of antioxidant activity DPPH was done and total phenolic content was also conducted. Physiochemical, microbial (TPC), antioxidant (TPC, DPPH), and sensory evaluation of supplemented apricot probiotic drink were done at different storage periods (1, 14, and 28 days) and obtained data were subjected to the statistical design. There was a considerable change in total phenolic content (TPC) throughout storage in all treatments, but supplementation of sea buckthorn has increased the TPC and antioxidant activity of SAPD. The probiotic count of T2 showed the highest probiotics among all treatments with 6.70 log CFU/mL on the 1st day, 6.6 log CFU/mL on the 14th day, and 6.5 log CFU/mL on the 28th day. It was having 4% sea buckthorn followed by T1 having 6.56 log CFU/mL and T0 having 6.54 log CUF/mL of grand mean with little change during the storage period of 28 days. T3 and T4 showed decreased number of probiotic counts with an increase in the percentage of sea buckthorn. This research concluded that sea buckthorn can be supplemented in fruit drinks to provide probiotic and antioxidant benefits, but its increased ratio can reduce the overall acceptability of the drink.

KEYWORDS:

• Prebiotic
• probiotics
• supplementation
• sea buckthorn
• apricot

Introduction

People always found nutritionally enriched functional beverages that make novel innovations in food industries. The raised demand for healthy functional foods continuously shaped the food industry to prepare novel food products to meet desired qualities. This has resulted in a hike in demand for functional drinks specifically enriched with nutrients and functional properties. Functional drinks not only quench the thirst but also prevent diseases and improve physical and mental health. In the food market, functional drinks are nutritionally important and used for disease prevention.^[1]

Apricot (Prunus armeniaca) is a delectable fruit that first originated from temperate areas of the world. In Pakistan, it is mostly cultivated in northern areas and then transported for consumption in the whole country either in fresh or dried form. It is popular for its great nutritional and esthetic attributes, wonderful flavor, and eye-catching look. It is considered the “King of Dry Fruits” due to its high nutritional content, magnificent aroma, taste, and flavor. In 2018, Pakistan’s yearly apricot output was 178,957 tons, placing the country sixth in worldwide apricot productivity.^[2] C-1: References should be in number format. These God-gifted fruits have antioxidant potential due to the presence of carotenoids and organic acids.^[3] It can be used in multiple clinical trials for different diseases such as cardiovascular (CVD).^[4] It is a good source of vitamin A, therefore this can be used as a drug.^[5]

The word “Sea buckthorn” refers to a tough, thorny, deciduous shrub of the genus Hippophae, family Elaeagnaceae.^[6] When completely mature, sea buckthorn berries are 6–9 mm long, oval in form, and yellow, orange, or red. The fruit of the sea buckthorn tree is high in health-promoting compounds and nutrients. Tocopherols, ascorbic acid, carotenoids, and polyphenols are the most abundant, but B vitamin components, minerals, and lipids are found in sea buckthorn.^[7] The berries of sea buckthorn can be considered a vital source of all the essential nutrients required for humans. When compared with other fruits like oranges and peaches, the amount of several nutrients in sea buckthorn is greater. Sea buckthorn has therapeutic potential. When this fruit was taken with a diet, it reduced the low-density lipoprotein (LDL) and triglyceride levels in blood.^[8] It is beneficial to investigate novel methods of preparing different products by using sea buckthorn fruit to boost customer acceptability. Non-digestible oligosaccharides (NDO) present in sea buckthorn berries can increase the activity of health-promoting bacteria in the human digestive tract while decreasing the activity of health-harming bacteria, indicating prebiotic characteristics.^[9]

Prebiotics was first defined in 1995 as indigestible food item which is consumed by gut bacteria in human colon for their nourishment and growth thus contributing positively to the health of human. Prebiotics are classified into many groups of compounds that are very significant for the good health of gut microbiota.^[10]

Prebiotics and probiotics are very vital components of the human diet today. It has been seen these years that there is a huge development regarding the classification and verification of the advantages of probiotics and prebiotics related to human health.^[11,12] Prebiotics (as a source of energy in the process of fermentation) ingestion changes the composition of the gut by fermenting the substrates that increase the number of microbes. Prebiotics have a tremendous impact on the intestinal beneficial bacteria (Bifidobacteria and Lactobacilli) that improves physiological functions like short-chain fatty acids construction and spread the microorganism chain. Besides such profitable effects on the colon, prebiotics also plays a positive in the urogenital tract, skin, and oral cavity. The integral basis for counting other compounds such as polyphenols, whole grain, arabinoxylan, pectin non-carbohydrate, and starches in the list of prebiotics was promoted by authors.^[13]

Probiotic foods are frequently classified as dairy items, which are not suited for persons who are lactose intolerant. As a result, it makes sense to look for novel formulations and broaden the spectrum of probiotic products to include nondairy meals. Probiotics when added to any product produce antioxidants and vitamins after fermentation. It also helps to change the carbohydrate of high-calorie into low-calorie carbohydrates and can break down fats and cholesterol.^[14] Probiotic drinks are used as a potential functional food. In recent research, agricultural waste are used for the preparation of functional beverages. Voss et al.^[15] prepared the synbiotic beverage by using soybean waste “okara.” This multifunctional drink contains antioxidant and ACE inhibitory activities, which have promised a strategy for disease prevention.^[15] Concisely, this study aimed to develop a fermented functional drink from apricot and sea buckthorn and then evaluated the prebiotic and antioxidant potential of probiotic apricot drink.

Materials and methods

The present study was on the development of apricot drink with the addition of sea buckthorn powder as a prebiotic and antioxidant source and Lactobacillus rhamnosus as a probiotic and to analyze and evaluate physicochemical, microbiological, antioxidant, and sensory attributes.

Sun-dried sea buckthorn and apricot samples

Sun-dried sea buckthorn berries and apricot were collected from Skardu Gilgit Baltistan and cleaned manually by local people by separating thorns, leaves, and other impurities. Dried sea buckthorn was ground to form a powder using mortar and pistil and stored in a zip lock back for use in product development. Then, the washed apricots were dipped in distilled water overnight. The dipped apricot got moisture and became soft.

Activation of probiotic (lactobacillus rhamnosus)

The lactobacillus rhamnosus (dry powder) was purchased from the Ayub agriculture research center, Faisalabad. The dried powder inside one sachet (dose) of each probiotic formulation was aseptically weighed and activated before use, as recommended by the manufacturers. The activation method used was De Man Ragosa Sharpe (MRS) broth, using 1 g of probiotic for every 100 mL of broth, and incubated for 15 h at 37°C. After the incubation for activation, the culture together with the MRS broth was centrifuged at 4670 rmp for 15 min in a centrifuge, at 4°C and washed in NaCl solution (0.85% w/v) twice. Same method was used by Silva et al. (2021).

Chemical analysis of dried sea buckthorn and apricot

pH, acidity, moisture, ash, protein, fiber, fat, and carbohydrate analysis were performed according to AOAC.^[16]

Preparation of supplemented apricot probiotic drink

The socked apricot was blended by a blender and filtered with a muslin cloth to filter the coarse particles. About 200 mL of the apricot blend was filled in each sterilized bottle, which added 2% of probiotic lactobacillus rhamnosus from pure culture and 5 treatments were prepared (T₀, T₁, T₂, T₃ and T₄) where the ratio of sea buckthorn was added with different percentage (0%, 2%, 4%, 8% and 12%). All samples were stored at 4°C after 24 h of fermentation at room temperature. Each treatment was having three replications; a total of 15 samples were prepared and divided into 3 batches for storage study. The five samples were labeled as ‘’A’’ batch which were tested on the 1^st day for the probiotic count, TPC, TFC and antioxidant activity, and other tests. Batch B was tested on the 14^th day and Batch C was tested on the 28^th day.

Physiochemical analysis of supplemented apricot probiotic drink

pH, acidity, and total soluble solids were measured according to the protocols of AOAC^[16] using a pH meter, phenolphthalein indicator, and refractometer, respectively. The colors of all samples were measured using a colorimeter; this practice was performed according to the modified protocol of Nour.^[17]

Antioxidant analysis of apricot probiotic drink

Total polyphenolic content (TPC)

The Folin-Ciocalteu technique, as reported by Naz et al.,^[18] was used to quantify the total phenolic contents with varying amounts of gallic acid in which the calibration curve was created. The gallic acid solution in methanol at concentrations of 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, and 0.10 mg/mL was combined with 5 mL of tenfold diluted Folin-Ciocalteu reagent and 4 mL of sodium carbonate (10%). After recording the absorbance at 760 nm for an hour, the calibration curve was drawn using the absorbance as a function of concentration. The same reagent as stated above was combined with 1 mL of a sample (0.001 g/mL), and after an hour the absorbance of the resultant blue color complex was measured at 760 nm. Each determination was made three times. Quantification was performed using gallic acid as the standard.^[19] The following formula was used to determine the total amount of phenolic compounds present in the plant extracts as gallic acid equivalents per gram (GAE/g).

T= C$\times$V/M

DPPH scavenging activity

According to Usman et al.,^[20] the antioxidant potential of the product was assessed using the DPPH radical scavenging test with slight modification. Following this approach, 3 mL of the sample was combined with 1 mL of newly made, 0.004% DPPH in methanol solution. The combination was then left in the dark for 30 min. Then at 517 nm, absorbance was detected. A reaction combination with a lower absorbance has a high capacity to scavenge free radicals. There were three copies of each test run. These calculations were used to determine the % inhibition of DPPH radical samples.

$DPPHInhibition\left(\mathrm{\%}\right)\frac{Blankabsorbance\left({\mathrm{A}}_0\right)-Sampleabsorbance\left({\mathrm{A}}_1\right)\times 100}{Blankabsorbance\left({\mathrm{A}}_0\right)}$

Microbial analysis of apricot probiotic drink

Using Man Rogosa Sharpe agar (MRS), medium Lactobacillus rhamnosus was enumerated. Incubation of plates was done at 37ºC for 48 h.^[21] The number of bacteria is estimated indirectly based on the number of colonies generated by the cells of microorganisms after thermostatization at 37°C for 48 h. MRS agar was used to count the total plate count as described by Caleja et al..^[22] The 20 g of plate MRS agar was taken and then dissolved in the 300 mL of distilled water. It was then placed in an autoclave. After autoclaving, it was kept for some time for lowering its temperature and after cooling, sixfold dilutions of all samples were prepared. A 9 mL of saline in six test tubes each was taken. In 1^st test tube, a 1 mL Apricot drink sample was added making the volume up to 10 mL. In the next step, 1 mL out of this test tube was added into the second test tube having 9 mL of saline. Further, 1 mL out of the 2^nd test tube was taken and poured into the 3^rd test tube and so on. A 0.1 mL out of these serially diluted samples was transferred to petri plates. These petri dishes were incubated at 37°C for 24–48 h. These plates were observed under the colony counter to count the probiotic colonies and results were recorded.

Sensory analysis

Ten panelists, including graduate students from the Department of Food Science and Technology at the University of Agriculture Faisalabad, completed sensory evaluations of the supplemented apricot probiotic drink on the first, fourteenth, and twenty-eighth days of its storage term of 4 weeks. A nine-point hedonic scale was used for the test, with one denoting dislike extremely and nine denoting like extremely. Water was offered in between samples to wash the palate, and the sequence in which the items were presented was randomly chosen.^[23]

Statistical analysis

Data collected were analyzed and interpreted by a suitable statistical design to evaluate the level of significance.^[24]

Results and discussion

Chemical analysis of dried sea buckthorn and apricot

Table 1, which displays the mean values of the physicochemical parameters of sea buckthorn, illustrates the findings of the physicochemical examination of sea buckthorn berries (ash, moisture, fiber, acidity, pH, and TSS). The findings showed that sea buckthorn berries that have been sun-dried have 5.5% moisture, 4.6% ash, 4.5% fiber, 27.2% lipids, 54.3% carbs, 2.85 pH, and 1.88% acidity. Tkacz et al.^[25] investigated the physicochemical and phytochemical characteristics of sea buckthorn juice during the storage period and fresh to determine the stability of components. The outcome of the proximate analysis of sun-dried apricot is shown in Table 1. According to it, the apricot’s moisture content was 12.75%. The sample that experienced open sun-drying yielded 3.8% ash. The results showed the greatest mean values of 80.67% for carbohydrates, followed by 3.1% crude fiber and 1.83% crude lipids. In samples that were dehydrated in the sun, the crude protein % was 0.95. The outcomes of the proximate analysis of sun-dried apricots closely matched the findings of the investigation of Khan et al..^[26] Drying causes the removal of moisture from apricot due to which the percentage of carbohydrates increased. It is common knowledge that foods with minimal fat content typically include a lot of moisture. Fruit juicing as a supplement is best due to fruits’ greatest moisture content. High moisture content also tends to encourage chemical deterioration and microbiological infection as stated by Khan et al. (2010). The pH of dried apricot was 2.65 and the acidity was 3.78, the results correlate with the findings of Wani et al.^[27] who studied the physicochemical properties of apricot.

Table 1. Chemical analysis of sea buckthorn and apricot powder, results are expressed in Mean $\pm$standard deviation.

Parameters	Sea buckthorn	Apricot
Moisture	$5.5\pm$ 0.57%	12.75 $\pm$1.72%
Ash	4.6 $\pm$1.25%	3.8$\pm$0.76%
Fibre	4.5 $\pm 0.55$ %	3.1$\pm$0.57%
Fat	27.2 $\pm 1.9$ %	1.83$\pm$0.5%
pH	2.85 $\pm$0.74%	2.65 $\pm$0.74%
Acidity	1.88$\pm$0.52%	3.78$\pm$0.42%
Carbohydrate	54.33 $\pm 2.88$ %	80.07$\pm$0.56%

Physicochemical analysis of apricot probiotic drink

Proximate analysis

The results related to pH and acidity which were calculated through analysis of variance for apricot probiotic drink developed by the addition of various concentrations of sea buckthorn and 2% Lactobacillus rhamnosus are given away in Table 2. The data presented in Table 2 show that drink pH resides between 4.20 and 3.24 within the treatments and throughout storage days. Among various concentrations of sea buckthorn, pH fluctuates significantly (p < .05) and efficiently highest decline in pH was noted in T₄ within treatments and along storage days. According to the result of the study, a significant decreasing trend was exhibited in the pH of the drink. The mean values for the pH showed a decreasing trend in whole treatments and after 28 days with a maximum drop in pH was observed in T₄ (3.24) which is further chased by T₃ (3.28), whereas the lowest fall in pH was detected in the control treatment (3.40). Probiotics used simple sugars during fermentation and produce organic acids. These organic acids reduced the pH of the product during storage.

Table 2. Mean values of pH, acidity and TSS of a supplemented apricot probiotic drink. Values with different letters were significantly different (p ˂ 0.05).

	pH			Acidity			TSS
Treatment	1 Day	14 Days	28 Days	1 Day	14 Days	28 Days	1 Day	14 days	28 days
T0	4.2$\pm 0.53$^a	3.81$\pm 0.60$^e	3.40$\pm 0.48$^i	0.50$\pm$0.51^g	0.58$\pm$0.65^ef	0.66$\pm$0.59^c	7.52$\pm$0.50^hi	8.13$\pm$0.63^defg	8.42$\pm$0.61^bcd
T1	4.1$\pm 0.89$^ab	3.78$\pm 0.91$^ef	3.38$\pm 0.61$^i	0.51$\pm$0.63^g	0.59$\pm$0.77^de	0.67$\pm$0.58^c	7.43$\pm 0.76$i	7.92$\pm$0.84^efgh	8.52$\pm$0.65^abcd
T2	4.17$\pm$0.62^bc	3.77$\pm 0.51$^fg	3.33$\pm 0.66$^j	0.55$\pm$0.49^f	0.62$\pm$0.77^d	0.73$\pm$1.3^b	7.68$\pm$0.83^ghi	8.21$\pm 1.50$^def	8.73$\pm$0.59^abc
T3	4.15$\pm 0.71$^c	3.75$\pm 0.71$^g	3.28$\pm 0.94$^k	0.58$\pm$0.59^ef	0.67$\pm$0.50^c	0.77$\pm$0.68^a	8.12$\pm$0.58^defg	8.33$\pm$0.79^cde	8.85$\pm$0.62^ab
T4	4.12$\pm 0.82$^d	3.68$\pm 0.51$^h	3.24$\pm 0.51$^l	0.62$\pm$0.70^d	0.68$\pm$1.01^c	0.80$\pm$0.88^a	7.87$\pm$0.61^fghi	8.41$\pm$0.61^bcd	8.92$\pm$0.43^a

The outcome of the current study is strongly linked to the results of Khan et al.^[28] who prepared mango sea buckthorn blended juice and studied the stability of juice during a storage period of 3 months, studied the physicochemical parameters of juice and noticed a decrease in pH of juice during storage. The decrease is due to the increase in organic acids as sea buckthorn has a pH equal to 2 and has a high amount of organic acids.

Table 2 presents the results regarding acidity. With changes in the concentration of sea buckthorn and storage days, acidity varies significantly (p < .05) as shown by statistical results. Combinations among dual variables were non-significant. Based on the data displayed in Table 2, it was indicated that the new drink acidity starts from 0.50 to 0.80 throughout the storage. According to the mean values of the table, a significant (p < .05) increased trend was observed in the acidity of the drink except for T₀ and T₁ which showed no significant difference. In T_4, later than 28 days of cold storage has the highest acidity followed by T₃ and T₂. The pH and acidity have an inverse relation to each other along with the development of acid, acidity increased so pH reduce. The data obtained in the current study are precisely related to the work of Gunenc et al.^[29]; they used sea buckthorn as a fresh source of prebiotics that enhanced the probiotic survival in yogurt by the addition of bioactive natural products from sea buckthorn and also studied physicochemical parameters of yogurt and noticed an increase in acidity of yogurt during storage. They concluded that microbial activity causes the production of acids that increase the acidity of probiotics yogurt during storage. Acids have a strong impact on pH values, as they together are in opposite linkage to one another, one is increasing, and another is decreasing. Another study conducted by Khan et al.^[28] showed a similar trend of change in the acidity of juice made from sea buckthorn and mango.

There was a significant effect of treatments on the TSS of the drink and the effect of storage days’ also showed a significance (p < .05) level. However, the interaction between treatment and days showed a non-significant result. Table 2 describes the mean value of treatments over the period. According to the overall mean of treatments in the table, the highest TSS value was recorded in T₃ which did not differ significantly from T₄, again little difference was seen in T₄ and T₂. T₂ showed similarity with T₀ and T₁ got the least TSS. An increase in trend can be observed in the treatments during storage and within treatments, there is fluctuation where there was no proper trend. T₀ has high TSS in every treatment, T₁ showed reduced TSS compared to T₀, T₂ and T₃ showed a rise in TSS and T₄ TSS again reduced. The conversion of polysaccharides to sugars may be the cause of the rise in TSS seen after storage. Additionally, this could have increased the amount of total sugars in storage.^[30] The current study shares similarities with the research of Khan et al.,^[31] which found the breakdown of polysaccharides into sugars during storage, increased the TSS content in drinks. This research examined the organoleptic and sensory properties of drinks made with sea buckthorn and aloe gel.

Color analysis

The color of food products is as central as a taste for the quality guarantee of the food. According to marketers, color is one of the major factors that affect the buying behavior of customers and consumers. Most consumers decide on the flavor of food products based on their color. The ANOVA (Table 3) describes the variance of analysis of color for L*, a* and b* values. The effect of treatments on color of the drink is significant (p < .05) for L*, a* and b × . However, the effect of days on the drink with sea buckthorn supplementation is significant (p < .05) for L* and a*, but b* values showed non-significant results for the days and interaction between treatments and days. The interaction between days and treatments on the color characteristics of the drink for L* and a* is significant as well. The mean value describes that the supplementation of sea buckthorn has decreased the acceptability for consumers by the increase in the storage days. The mean values clearly show the major impact of treatment on color since it reduces drink lightness during treatment and as time goes on during storage. The change in color light (orange to dark) can also be a reason for low acceptability. The range of values was between 32.7 and 25.0. Values drop during storage because of a change in the L* value of color brought on by a rise in acidity and a fall in pH.

Table 3. Mean values of color analysis (L*, a*, b*) of a supplemented apricot probiotic drink. Values with different letters were non-significantly different (p > .05).

Treatment	L	a*	b*
T0	32.26$\pm 1.04$^A	10.14$\pm 0.92$^A	32.85$\pm 1.15$^A
T1	30.10$\pm 0.67$^B	12.02$\pm 0.69$^B	30.92$\pm 0.47$^B
T2	28.14$\pm 1.18$^C	14.07$\pm 0.56$^C	29.05$\pm 0.75$^C
T3	26.20$\pm 0.76$^D	16.27$\pm 1.51$^B	27.22$\pm 0.58$^D
T4	24.07$\pm 0.79$^E	18.10$\pm 0.97$^A	25.00$\pm 1.01$^E

Microbial examination of apricot probiotic drink

The outcome of probiotic count regarding analysis of variance for a supplemented apricot probiotic drink is given in Table 4. The statistical findings showed that treatment and storage days had a significant (p < .05) impact on the probiotic count of drinks. A non-significant interaction was seen between the two factors of treatment and days.

Table 4. Mean values of total plate count (log CFU/mL) of a supplemented apricot probiotic drink. Values with different letters were significantly different (p ˂ 0.05).

Treatment	1 Day	14 Days	28 Days
T0	6.5$\pm$0.61^ab	6.4$\pm$0.58^ab	6.3$\pm$0.21^b
T1	6.7$\pm$0.43^a	6.6$\pm$0.21^ab	6.4$\pm$0.70^ab
T2	6.7$\pm$0.51^a	6.6$\pm$0.34^ab	6.5$\pm$0.62^ab
T3	5.3$\pm$0.43^c	5.2$\pm$0.32^c	5.2$\pm$0.55^c
T4	5.2$\pm$0.33^c	5.2$\pm$0.42^c	5.1$\pm$0.45^c

The final results are presented in Table 4 and demonstrated that the viability of bacteria for the probiotic apricot drink slightly fluctuated between different treatments and throughout the storage period. The number of probiotics resides between (5.1–6.7). According to the mean values of the table, T₃ and T₄ showed a decrease in the number of probiotics with treatments but no significant (p > .05) change was noticed concerning storage days, except T₂ and T₁ which contain 4 and 2% sea buckthorn. T₂ and T₁ showed the highest probiotic count throughout the storage period as compared to other treatments followed by T₀. Both of them were not significantly different (p > .05) with the increase in storage days. The grand mean values of storage indicated that there was a little change in probiotic number as the 1^st day and 14^th day showed the same results and on the 18^th day, the number reduced but still the reduction was very less. It was 6.08 log CFU/mL on the 1^st day, 6.00 log CFU/mL on the 14^th day and 5.90 log CFU/mL on the 28^th day. Within the 1^st day, T₀ has 6.5 logs CFU/mL then an increase in number was seen in T₁ and T₂, after which the number decreased for T₃ and T₄. The same behavior was shown by each treatment in the table with some fluctuations. It can be concluded from the results that treatments with less percentage of sea buckthorn showed better results and increased the number of probiotics and it has prebiotic qualities. But it can reduce probiotics if used in a high percentage, and it could be due to its acidic nature.

Tkacz et al.^[25] monitored the growth of Lactobacillus plantarum in a different mixture of sea buckthorn and apple juice and described the course of malolactic fermentation for 72 h. After 72 h, the number of colony-forming units decreased by approximately 4.7 logs for the 10% sea buckthorn juice, by 5.3 logs for the 20% juice, by 5.5 logs for the 30% juice, and by 4.6 logs for the 40% juice. It has been shown that Lactobacillus plantarum cells going through the decline phase have active metabolic pathways associated with malolactic fermentation. The presence of high fibers, high acidity, and vitamin C content in apricot can be correlated to the survival rates of bacteria. Organic acid in sea buckthorn results in the acidic nature of the drink, which reduced the number of bacteria with treatment and with time.^[30]

Antioxidant analysis of apricot probiotic drink

Total polyphenols contents (TPC)

Table 5 describes the variance analysis of polyphenols in this drink and the effects of treatment and storage are at a high significance level (p < .05). Interactions between treatments and storage period also indicate high significance. Table 5 demonstrates the mean resides between 483.69 and 55.14 values of polyphenols in apricot probiotic drinks over the storage period of 1, 14 and 28 days. These values indicate greater differences during 1, 14 and 28 days of storage. The phenolic content within the treatments was increasing with the increase in the percentage of sea buckthorn but a high decreasing trend was observed with the increase in storage days. The mean value of total phenolic content on the 1^st day was 463.32 which was reduced to 385.71 on the 14^th day and followed by 52.29 on the 28^th day. However, the overall mean value for treatment showed an increasing trend. The highest TPC was observed in T₄ which was enriched with 12% sea buckthorn but the least was observed in T₀ having 0% sea buckthorn.

Table 5. Mean values of total phenolic contents (mg GAE/L) and DPPH (%) of a supplemented apricot probiotic drink. Values with different letters were significantly different (p ˂ 0.05).

	TPC (mg GAE/L)			DPPH (%)
Treatment	1 Day	14 Days	28day	1 Day	14 Days	28 Days
T0	436.60$\pm$2.32^d	264.41$\pm$6.47^f	16.14$\pm$0.96^h	16.00$\pm$0.51^k	14.01$\pm 0.88$^l	12.34$\pm$0.56^m
T1	447.26$\pm$1.69^c	408.70$\pm$0.25^e	28.53$\pm$0.45^i	43.65$\pm 1.58$^h	40.01$\pm$0.72^i	37.00$\pm$0.81^j
T2	470.47$\pm$0.63^b	410.17$\pm$1.00^e	44.83$\pm$0.58^k	50.33$\pm$0.58^f	47.01$\pm$0.81^g	44.01$\pm$0.65^h
T3	478.59$\pm$1.11^ab	433.26$\pm$10.40^d	55.05$\pm$0.18^j	61.81$\pm$0.89^c	57.00$\pm 1.01$^d	52.33$\pm$0.57^e
T4	483.69$\pm$3.99^a	412.05$\pm$0.31^e	116.90$\pm$0.58^g	70.01$\pm$1.00^a	66.66$\pm$0.57^b	63.01$\pm$0.99^c

The results of the current study show a similar trend as those of Selvamuthu and Khanum,^[32] who worked on the development of spiced sea buckthorn mixed fruit squash. The total phenols content of the spiced squash was estimated to be 125.67 mg/100 gm initially, and this was found to decrease significantly (p < .05) to 62.83 and 28.90 mg/100 gm, after 6 months of storage at ambient temperature and 37°C, respectively. This data indicates that a considerable loss in total phenols occurred during the storage period. The loss in total phenols accounted for 50% and 77% under ambient temperature and at 37°C, respectively, after 6 months of storage.

Negi and Dey,^[33] performed a comparative analysis of the total phenolic content of sea buckthorn wine and other selected fruit wines. For this, individual fruit juices were used to set up wine fermentation using the same lab-isolated strain. The most remarkable finding of the study was that the total phenolic content of sea buckthorn wine (689 mg GAE/L) was comparable to grape wine (647 mg GAE/L). Similar studies have shown similar trends.^[33]

DPPH

The results in Table 5 demonstrate an analysis of the variance of DPPH content in different concentrations of sea buckthorn-supplemented apricot probiotic drink. According to (Table 5), the effect of different concentrations of treatments shows high significance. The effect of days and interaction between days and treatments of sea buckthorn apricot probiotic drink is also highly significant.

Table 5 demonstrates the mean scores of DPPH of treatments during different storage days from 12.34 to 70.01. The minimum value was exhibited on the 28^th day of the control group and the maximum value was observed on the 1^st day of T₄ which has 12% of sea buckthorn in the apricot probiotic drink. The higher antioxidant activity in treatments was due to the addition of sea buckthorn at different concentrations of 2%, 4%, 8%, and 12%. There was a declining trend in the antioxidant activity of the sea buckthorn-supplemented apricot probiotic drink during the storage periods. The decrease in polyphenols may be the reason for decreased antioxidant activity. Klimczak et al.^[34] studied the effect of storage time and temperature on the phenolic content and antioxidant activity of orange juice for 6 months. He found out that with the increase in months, the antioxidant activity decreased from 53.2% to 41.2% and then to 27.3% on the 2^nd, 4^th, and 6^th months. The decrease in antioxidant activity is linked with the phenolic content which is reducing constantly with the storage period. Accordingly, juices that show high antioxidant activity contain a high concentration of Vitamin C and phenolic compounds.

Sensory analysis

From a marketing perspective, food acceptability is crucial. Consumer preference is influenced by the acceptability and appropriateness of food goods. The key elements of sensory evaluation to assure the high quality of food products are taste, aroma, appearance, and overall acceptability. Taste is usually defined as the perception of the flavor of eatable products either food or a drink when exposed to the mouth. All the treatments showed acceptable scores with the addition of different concentrations of sea buckthorn. According to Figure 1 analysis of variance, the effects of treatments on taste, aroma, and appearance were significant (P < .05). Figure 1 describes the values of treatments T₀, T₂, T_3, and T₄ showing a significantly decreasing trend except for T₁ which has increased with the addition of sea buckthorn as compared to T₀. The treatments having less concentration of sea buckthorn had a high preference score as compared to an apricot drink with high sea buckthorn. T₁ and T₂ had non-significant (P > .05) differences as compared to other treatments because they did not change the acidity of the drink. Similarly, due to the increase in organic acids and number of bacteria, the mean score of overall acceptability of T₁ and T₂ got high scores 6.4 and 6.2 from judges, which were slightly lower than the control treatment (6.8). The results of the present study have resemblances with the study of Shawi et al..^[35] During the storage periods, there was a change in the chemical composition of supplemented sea buckthorn yogurt, where increased acidity and a decrease in pH make yogurt sour or acidic and which ultimately led to an undesirable taste for consumers.

Figure 1. Mean preferences score for sensory attributes for a supplemented apricot probiotic drink.

Conclusion

The current research aimed to assess the prebiotic capability of sea buckthorn on the growth and survival of Lactobacillus rhamnosus and also enhance the antioxidant activity of apricot probiotic drinks. The acidity of supplemented apricot probiotic drink was recorded which supports the growth and persistence of Lactobacillus rhamnosus. An increase in Lactobacillus rhamnosus count was observed with an increase in sea buckthorn concentration up to 2 to 4% in treatment T₁ and T₂ followed by T₀ having 0% sea buckthorn, Further increase in the concentration of sea buckthorn powder exhibited decreasing array in microbial percent of apricot probiotic drink due to antimicrobial and acidic property of sea buckthorn. The decrease in the probiotic count was very less with storage days. According to various results of the study sea buckthorn added drink presented an increase in polyphenols with the increase in the concentration of sea buckthorn but a considerable reduction in polyphenols was noticed with an increase in storage days. Decreased pH throughout treatments and with storage day can cause increased titratable acidity of the drink. The TSS of apricot probiotic drink showed an increasing trend with some fluctuation with the addition of sea buckthorn and storage days. The highest TSS was observed in T₄ after 28 days it could be because of the breakdown of polysaccharides into sugars during storage. Supplementation of sea buckthorn has a significant effect on the color of the drink the lightness L* value was decreasing in each treatment, the redness a* of the drink has been increased and the yellowness b* of the drink showed decreasing trend due to which the light orange color of drink changes to dark orange in treatment T₄. The results of sensory evaluation for overall acceptability indicated that T₀ ranked 1^st, T₁ 2^nd and T₂ 3^rd, T₃ 4^th and T₄ was given the lowest score which was supplemented with 12% sea buckthorn that might be due to the high acidic and sour taste, all treatments showed change compared to controlled one. A similar trend was observed for aroma and appearance but the taste of T₁ was given a high score among other treatments followed by T₀, T₂ T₃ and T₄. Sea buckthorn has prebiotic and antioxidant properties but its low concentration can exhibit good sensory quality products that can be acceptable for consumers.

Data availability

Although adequate data has been given in the form of tables and figures, however, all authors declare that if more data are required then the data will be provided on a request basis.

Acknowledgments

The authors would like to thank the Researchers Supporting Project number (RSP2023R138), King Saud University, Riyadh, Saudi Arabia.

Disclosure statement

No potential conflict of interest was reported by the authors.

Additional information

Funding

The authors declare that no funds, grants, or other support were received during the preparation of this manuscript.