Goat's milk has different composition compared to the more commonly consumed cow's milk. It has potentially better digestibility, higher buffering capacity and lower contents of αs1-casein (which may be responsible for causing allergic reactions) than cow's milk (Biadala and Konieczny, Reference Biadała and Konieczny2018). Goat's milk is also reported to contain more free amino acids and have higher antioxidant potential than cow's milk (Biadala and Konieczny, Reference Biadała and Konieczny2018). Simos et al. (Reference Simos, Metsios, Verginadis, D'Alessandro, Loiudice, Jirillo, Charalampidis, Koumanis, Boulaka, Martemucci and Karkabounas2011) also investigated the antioxidant activity of goat, donkey and cow's milk and concluded that goat's milk provided the best antioxidant capacity. Moreover, its casein precipitates more readily into more fine particles as compared to cow's milk (Ulusoy, Reference Ulusoy2015). Many potential health benefits, like for instance anti-diabetic, antihypertensive, antioxidant, anti-microbial and anti-inflammatory activities, have been attributed to goat's milk (Nayik et al., Reference Nayik, Jagdale, Gaikwad, Devkatte, Dar, Dezmirean, Bobis, Ranjha, Ansari, Hemeg and Alotaibi2021).
Goat's milk can be converted into many products, such as cheese, yogurt, milk powder, ice-cream, butter and fermented milk products (Ribeiro and Ribeiro, Reference Ribeiro and Ribeiro2010; Nayik et al., Reference Nayik, Jagdale, Gaikwad, Devkatte, Dar, Dezmirean, Bobis, Ranjha, Ansari, Hemeg and Alotaibi2021) and could potentially have use in the production of infant formula (Prosser, Reference Prosser2020; Vázquez-Garcíaet al. (Reference Vázquez-García, Cardador-Martínez, Orihuela-López, Ramos-Hernández and Martín-del-Campo2021) analyzed antioxidant peptides in Mexican goat cheese fractions (acid-soluble and non-protein nitrogen) during ripening. They suggested that the ripened Mexican goat cheese showed DPPH radical scavenging activity owing to the presence of peptides which were either present originally in the milk or released by the actions of starter culture during ripening. Another study also reported that goat cheese had high antioxidant activities due to peptides released by the action of rennet during cheese manufacturing (Hernández-Galán et al., Reference Hernández-Galán, Cardador-Martinez, Lopez-del-Castillo, Picque, Spinnler and del Campo2017). Further, López-Villafaña et al. (Reference López-Villafaña, Rojas-González, Elías-Román and Rodríguez-Hernández2023) also observed appreciable DPPH radical scavenging activity (%) of Mexican Panela goat's milk cheese.
Citrus fruit crops are being cultivated across the globe due to the economic and nutritional values of their fruits (Addi et al., Reference Addi, Elbouzidi, Abid, Tungmunnithum, Elamrani and Hano2022).The juices from different citrus fruits were observed to contain antioxidants and other functional compounds. For example, lemon (Citrus limon) juice has been characterized for its composition by Alfadul and Hassan (Reference Alfadul and Hassan2016) who found appreciable quantities of vitamin C (23.9 mg/100 g juice) and minerals (Ca, Mg, P, K, Na). Its juice was reported to contain tannins, flavonoids, terpenoids and phenols (Rauf et al., Reference Rauf, Uddin and Ali2014). Being a rich source of antioxidants, lemon juice has been found to have protective and antioxidant features concerning improved hematological and biochemical parameters of oxidatively stressed mice (Ali et al., Reference Ali, Obaid and Awaid2020). Grapefruit (Citrus × paradisi) juice was found to contain appreciable quantities of flavanones such as naringin and narirutin (Sicari et al., Reference Sicari, Pellicanò, Giuffrè, Zappia, Capocasale and Poiana2018). Sweet lime (Citrus limetta) is commonly known as ‘Mosambi’ in the culture of the Indian subcontinent region. This fruit has been reported to have multiple traditional uses such as in the treatment of scurvy, constipation, indigestion, ulcers, diabetes and urinary disorder (Khan et al., Reference Khan, Mahmood, Siddiqui H and Akhtar2016). The mandarin variety ‘Feutrell's Early’ (Citrus reticulata Blanco) is a delicious citrus fruit in terms of its sweet taste and has been reported to contain appreciable quantities of vitamin c, DPPH radical scavenging activities, total phenolics, flavonoids, flavonols and carotenoids (Nawaz et al., Reference Nawaz, Abbasi, Khan, Ali, Hasan and Hayat2020; Manzoor et al., Reference Manzoor, Hussain, Anjum, Naseer, Ahmad and Ziogas2023). Kinnow (Citrus reticulate L.) is a cross between the King Orange (Citrus nobilis) and Willow Leaf Orange (Citrus deliciosa) which was developed by Dr H.B. Frost in California in 1935 (Sharma et al., Reference Sharma, Awasthi and Kumar2016). It was brought to the Indian sub-continent and is now grown in Pakistan, India and Bangladesh. The juice of kinnow is rich in vitamin B, vitamin C, β-carotene, minerals, and other bioactive compounds (Mahawar et al., Reference Mahawar, Jalgaonkar, Bibwe, Bhushan, Meena and Sonkar2020). All the above mentioned citrus fruit varieties are grown in Pakistan, especially in Punjab and are available in abundant quantities during winter season.
Among the dairy products, paneer is considered a primitive cheese variety which is produced through heat and acid coagulation of milk (Qureshi et al., Reference Qureshi, Amjad, Nadeem, Murtaza and Munir2019). People in Pakistan do not have a tradition to consume paneer either alone or with bread. In our previous study, traditional spices were incorporated into buffalo milk paneer matrix so as to increase its consumption (Qureshi et al., Reference Qureshi, Nadeem, Iftikhar, Salim-ur-Rehman, Ibrahim, Majeed and Sultan2023). Concerning goat's milk, some people avoid it due to its distinct flavor owing to the presence of more free amino acids and higher amounts of short-chain fatty acids (Ulusoy, Reference Ulusoy2015). In this context, it would be of interest to be able to convert goat's milk into value added product(s) so as to boost its consumption. Synthetic citric acid is usually used for the coagulation of milk in order to produce paneer at sweet shops or bakeries. Citrus fruit juices contain abundant quantities of natural ascorbic acid which may also act as a coagulant in the preparation of paneer. Moreover, citrus fruit juices are potentially health promoting due to the presence of abundant quantities of bioactive compounds. Therefore, in order to avoid deleterious effects of synthetic acids (for coagulation) as well as to enhance the nutrition and taste of goat's milk paneer, it would be advantageous to coagulate goat's milk with natural citrus fruit juices. To our knowledge, no study has been previously published regarding the quality and antioxidant potential of goat's milk paneer coagulated with juices from citrus varieties. Thus, the objective of the present study was to explore the physico-chemical characteristics, microbiological quality, sensorial acceptability and antioxidant potential of goat's milk paneer prepared through coagulation by the juices from different citrus varieties.
Materials and methods
Procurement of raw materials for development of paneer
Goat's milk used for paneer manufacturing was collected from a goat farm in a village of Bahawalpur city near Cholistan University of Veterinary and Animal Sciences, Bahawalpur. It was filtered using muslin cloth before being standardized: fresh goat's milk was separated to obtain skim milk and cream, which were then used to standardize the milk to 2.5% fat (7.5% solids-not-fat) before paneer manufacture. Different citrus fruits were purchased from the local fruit market.
Chemicals and apparatus
DPPH (2, 2-diphenyl-1-picrylhydrazyl), ascorbic acid, quercetin, Folin-Ciocalteu reagent and various other chemicals/standards were purchased from Merck (KgaA, Darmstadt, Germany). All other chemicals were also of analytical grade. All the apparatus and glassware was properly cleaned and manufacturing was done under hygienic condition in the laboratory. For the packaging of paneer, plastic sheet and aluminum foil were also bought from the local market.
Treatment plan and manufacturing
The citrus fruit juices and milk were used for the manufacturing of paneer. The six different paneers were made with citric acid (control), lemon juice, grapefruit juice, musambi juice, mandarin juice (Feutrell's Early) and kinnow juice. The pH and total soluble solids (°Brix) of the juices were determined previously, and in our preliminary trials the quantities of the different citrus juices required for coagulation were noted. These formed the basis of the paneer preparation, as detailed in online Supplementary Table S1. The pH of the whole paneer mixture was adjusted to 5.70–5.80 because proper coagulation occurred at that pH, and at that point addition of coagulants was stopped so that the coagulum attained should not be hard.
The standardized milk was transferred to a stainless steel container and heated for 5 min at 85°C, then cooled to 75°C with continual stirring before coagulation with different coagulants. The whey was then drained out by pouring it into the muslin fabric, and the resulting curd was placed in a hoop lined with the muslin cloth. Then, the paneer of each treatment was separately pressed for 30 min, yielding a small block of paneer. The paneer block was cut into five equal pieces. For each treatment, the same method was implemented. The cut paneer pieces were then carefully wrapped, first in a plastic sheet to prevent moisture loss and contamination, and then in aluminum foil. The wrapped pieces were stored at refrigeration temperature (4°C) for 3, 6, 9 or 12 d. One piece from each treatment was immediately frozen as fresh (day 0).
Physico-chemical analysis
Physico-chemical analysis of paneer was carried out during storage period. Gerber method was used to determine the fat in paneer. Moisture contents of paneer were determined according to IDF standard 4/ISO 5534 (IDF, 2004) through oven drying. After calibrating with buffers of pH 4.0 and pH 7.0, the pH of paneer was measured using a pH meter. With a few drops of water, the electrode was placed in grated paneer (20 g) (Ardö and Polychroniadou, Reference Ardö and Polychroniadou1999).Total nitrogen was calculated using the IDF standard 20B (IDF, 1993). Protein contents (%) were determined by multiplying TN (%) by 6.38. The titratable acidity was measured according to the method described in AOAC (Reference Latimer2016).
Microbiological analysis
Paneer was microbiologically analyzed for total plate counts (TPC, log CFU/g of paneer), yeasts and molds (YMC, log CFU/g of paneer) during storage period as described by Broadbent et al. (Reference Broadbent, Brighton and McMahon2013).
Sensory evaluation
Paneer was evaluated for sensorial quality using a 9 point hedonic scale (1 = dislike extremely, 2 = dislike very much, 3 = dislike moderately, 4 = dislike slightly 5 = neither like nor dislike, 6 = like slightly, 7 = like moderately, 8 = like very much, 9 = like extremely) for appearance and color, taste, texture and overall acceptability (Dongare and Syed, Reference Dongare and Syed2018). The sensory characteristics of prepared paneer samples were assessed by 20 trained assessors including graduates and faculty from the Department of Food Sciences at our university who know about the scaling procedures of paneer samples.
Other measurements
Water soluble extracts (WSEs) of paneer were prepared according to the procedure described by Gupta et al. (Reference Gupta, Mann, Kumar and Sangwan2013) with a few modifications, and used for further analysis. The total phenolic content (TP) of paneer or juices was determined using the method described by Reis et al. (Reference Reis, Stojković, Soković, Glamočlija, Ćirić, Barros and Ferreira2012). The TP contents were calculated as μg gallic acid equivalent (GAE)/g of paneer or μg GAE/ml of juice. Total flavonoid (TF) contents were measured via the spectrophotometric method according to Jia et al. (Reference Jia, Mengcheng and Jianming1999). The results were articulated as μg quercetin equivalent (QE)/g of paneer or μg QE/ml of juice. All determinations were carried out in triplicate. The ferric reducing antioxidant power (FRAP) was computed as described by Reis et al. (Reference Reis, Stojković, Soković, Glamočlija, Ćirić, Barros and Ferreira2012) with slight modifications. The results were calculated as μg ascorbic acid equivalent (AAE)/g or μg AAE/ml.
Radical scavenging activities were determined in two assays. The capability of samples to scavenge 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) was determined following the method of Zeghad et al. (Reference Zeghad, Ahmed, Belkhiri, Heyden and Demeyer2019). The ABTS radical scavenging activity was calculated as mg ascorbic acid equivalent (AAE)/100 g or mg AAE/100 ml. The potential for WSE of paneer or juices to scavenge 2,2-diphenyl-1-picrylhydrazyl radical (DPPH) was measured according to the method of Yi et al. (Reference Yi, Yu, Liang and Zeng2008) with some modifications. The DPPH radical scavenging activity was calculated as mg ascorbic acid equivalent (AAE)/100 g or mg AAE/100 ml. Finally, the total antioxidant capacity (TAC) was determined as described by Prieto et al. (Reference Prieto, Pineda and Aguilar1999). The TAC was calculated as mg ascorbic acid equivalent (AAE)/100 g or mg AAE/100 mL.
Statistical analysis
Statistical analysis was performed using Minitab statistical software version 16 (Minitab Inc., State College, PA, USA) by employing two-way ANOVA and Tukey's test for pair wise comparison at a level of P < 0.05.
Results
pH and total soluble solids (°Brix) of coagulants
Online Supplementary Fig. S1 shows pH and °Brix of coagulants used in the preparation of goat's milk paneer. Among the coagulants, lemon juice showed the minimum pH (2.42) whereas the maximum pH was shown by Musambi juice (4.23).The citric acid solution (2%) showed higher pH values (2.92) than lemon juice but lower values compared to juices from other citrus varieties (3.43–4.23). The minimum quantity of lemon juice was used in the development of goat's milk paneer followed by citric acid solution. The maximum quantity of musambi juice was used in the preparation of goat's milk paneer. Among the natural coagulants used in the present study, lemon juice showed the minimum °Brix (8.21) whereas the maximum °Brix was shown by kinnow juice (10.94). The citric acid solution (2%) showed the minimum °Brix (1.52).
Physico-chemical characteristics of goat's milk paneer
Data regarding the physico-chemical characteristics of the goat's milk paneers are shown in Table 1. All the treatments showed significant decreasing trend of pH during storage. The pH was slightly decreased after successive storage period. There was slight variation in acidity between each treatment. All fresh paneer in the present study showed pH values in the range 5.79–5.87. All the treatments showed significant increasing trend of acidity (%) during storage. The acidity was slightly increased after successive storage period. There was slight variation in acidity between each treatment. All fresh paneer in the present study showed acidity values more or less 0.30%. Even though, there was significant effect of storage on the moisture contents of paneer but that decrement was slight after successive storage period. The decreasing trend of moisture during storage thereby led to gradual increase in dry matter of paneer with the passage of time. All fresh paneer in the present study showed more than 53% moisture contents. All the treatments showed significant increasing trend of protein (%) contents during storage. The protein contents were slightly increased after successive storage period. There was slight variation in protein contents between each treatment. All fresh paneer in the present study showed fat contents in the range 17–19%. The fat contents were slightly increased after subsequent storage period. There was slight variation in the fat contents between each treatment. All fresh paneer in the present study showed fat content in the range 18–20%. All the treatments showed very slight increasing (non-significant) trend of ash (%) contents during storage. There was slight variation in ash contents between each treatment. All fresh paneer in the present study showed ash contents values more or less 2.0%.
Proximal characteristics of goat's milk paneers prepared through coagulation by different citrus juices, during storage
Means with different letters in the same column show significant (P < 0.05) differences between treatments and storage period.
Microbiological quality of goat's milk paneer
Microbial count data are given in Fig. 1. The TPC values of freshly prepared control paneer were higher (2.40 log10 CFU/g) than all other paneer prepared through coagulation of citrus juices (except musambi juice paneer). Generally, it was observed that the TPC values of all treatments varied significantly during storage. At the end of the storage period, most of the treatments showed TPC values in the range 5.61–5.81 log10 CFU/g. A similar trend was also seen regarding yeast and mold counts (YMC: Fig. 1). YMC of freshly prepared control paneer were 1.45 (log10 CFU/g) whereas all other fresh paneer coagulated by citrus juices showed slightly lower values. The values also increased significantly during storage in all the treatments and reached up to the range of 4.46–4.86 log10 CFU/g. All the goat's milk paneer prepared using different coagulants showed higher TPC values than the counts of Y & M. It was found that visible fungus started to appear on the surfaces of paneer stored for more than 15 d.
Total plate counts (TPC, means ± sd) (black bars) and yeast and molds (YMC, means ± sd) (gray bars) of goat's milk paneer prepared through coagulation by different citrus juices, during storage at 4°C.
Sensorial quality of goat's milk paneer
Sensory scores are given in Fig. 2 for appearance and color, flavor, texture and overall acceptability. Generally, all fresh paneer coagulated using different coagulants had higher sensory scores for all the sensory attributes. The paneer prepared using different citrus juices had higher sensory scores than control paneer at each storage period. Among all the treatments, the paneer prepared using mandarin juice gave the highest score for all sensory attributes. The paneer stored for 12 d was liked the least by the judges due to its slightly deteriorated quality.
Sensory acceptance scores (means ± sd) regarding appearance and color (black bars), flavor (dark gray bars), texture (light gray bars) and overall acceptability (white bars) of goat's milk paneer prepared through coagulation by different citrus juices, during storage at 4°C.
Antioxidant potential of juice from citrus varieties
Online Supplementary Fig. S2 presents the results concerning antioxidant potential of the different goat's milk paneers. The juice from all the citrus varieties had high TP contents with the highest shown by kinnow juice (1556.67 μg GAE/ml) followed by grapefruit juice (1356.55 μg GAE/ml). Regarding TF values, the highest value (566.86 μg QE/ml) was for kinnow juice followed by grapefruit juice (466.36 μg QE/ml). The juices from other citrus varieties also showed appreciable quantities of TF. Similarly, the highest value of FRAP was for kinnow juice (1310.09 μg AAE/ml) followed by mandarin (1309.05 μg AAE/ml). The juices from other citrus varieties also showed considerable FRAP values. The highest value of ABTS radical scavenging activity was for kinnow juice (224.33 mg AAE/100 ml) followed by lemon juice (199.80 mg AAE/100 ml). Good values of ABTS radical scavenging activity were also obtained by the juices from other citrus varieties. Regarding DPPH radical scavenging activity, the highest value was for mandarin juice (145.30 mg ascorbic acid equivalent (AAE)/100 ml). Good values of DPPH radical scavenging activity were also obtained by the juices from other citrus varieties. Concerning total antioxidant capacity, the highest value was for Feutrell juice (145.20 mg AAE/100 ml) followed by lemon juice (142.10 mg AAE/100 ml). Good values of TAC were also obtained by the juices from other citrus varieties.
Antioxidant potential of goat's milk paneer
Figure 3 shows the total phenolic and flavonoid contents as well as FRAP values of WSE from goat's milk paneer. The TP value of freshly prepared control paneer showed the lowest value (269.67 μg GAE/g) whereas the highest value (455.34 μg GAE/g) was for fresh paneer prepared from lemon juice. The paneer prepared using other citrus fruit juices also showed good TP contents. The TP values decreased gradually during storage in all the treatments. Regarding TF contents, the highest value was obtained by fresh paneer using kinnow juice (169.33 μg QE/g) followed by respective paneer made by grapefruit juice (144.70 μg QE/g). The lowest contents of TF were shown by the control (citric acid) paneer. The TF values also decreased gradually during storage in all the treatments. The highest value (140.88 μg AAE/g) of FRAP was for fresh paneer coagulated through lemon juice (T 1) whereas the lowest values were for control paneer. In this way, paneer prepared using kinnow juice and lemon juice showed promising results concerning TP and TF contents as well as FRAP values.
Total phenolics (TP, means ± sd, μg GAE/g) (gray bars), total flavonoids (TF, 
, means ± sd, μg QE/g) and ferric reducing antioxidant power (FRAP, 
, means ± sd, μg AAE/g) (white bars) of goat's milk paneer prepared through coagulation by different citrus juices, during storage at 4°C.
Figure 4 shows the ABTS and DPPH radical scavenging activities as well as TAC values of the paneers. In all three measures, the highest value was for fresh paneer prepared using kinnow juice followed by grapefruit juice, whereas the lowest values were shown by the control paneer. All three characteristics decreased gradually during storage in all the treatments. Overall, paneer prepared using kinnow juice and grapefruit juice showed promising results concerning DPPH and ABTS radical scavenging activities as well as TAC values.
DPPH radical scavenging activity (means ± sd, mg AAE/100 g) (black bars), ABTS radical scavenging activity (means ± sd, mg AAE/100 g) (gray bars), and total antioxidant capacity (TAC) (
, means ± sd, mg AAE/100 g) of goat's milk paneer prepared through coagulation by different citrus juices, during storage at 4°C.
Antioxidant potential of whey from goat's milk paneer
Figure 5 shows the results for TP and TF contents, FRAP and TAC values, DPPH and ABTS radical scavenging activity of whey collected during manufacturing of goat's milk paneer. The TP and also TF values of the control (citric acid) whey were the lowest whereas the highest values were obtained by paneer prepared using kinnow juice. The paneer prepared using juices of other citrus varieties also showed considerable TP contents. Whey from paneer made with grapefruit juice had the second highest TF value. Similarly, the highest value (17.01 μg AAE/g) of FRAP was obtained by whey from paneer coagulated with kinnow juice whereas the lowest value was shown by whey from control paneer. Whey from paneer prepared using kinnow juice and grapefruit juice showed promising results concerning TP and TF contents as well as FRAP values.
Total phenolics (TP, 
, means ± sd, μg GAE/ml)), total flavonoids (TF, 
, means ± sd, μg QE/ml), ferric reducing antioxidant power (FRAP, means ± sd, μg AAE/ml) (black bars), ABTS radical scavenging activity (means ± sd, mg AAE/100 ml) (dark gray bars), DPPH radical scavenging activity (means ± sd, mg AAE/100 ml) (light gray bars) and total antioxidant capacity (TAC) (means ± sd, mg AAE/100 ml) (white bars) of whey from goat's milk paneer prepared through coagulation by different citrus juices.
Regarding ABTS and DPPH radical scavenging activities, once again whey from kinnow paneer had the highest value (3.43 and 1.63 mg AAE/100 g, respectively) and that from control paneer had the lowest. The whey collected from other paneer coagulated by different citrus juices also showed promising results for ABTS values. Lemon juice paneer also had a high DPPH activity (1.62 mg AAE/100 g). Not surprisingly the highest value of TAC was for whey from paneer prepared using kinnow juice (22.03 mg AAE/100 g) and the lowest was for control paneer whey. Whey from paneer prepared using kinnow juice showed more promising results concerning DPPH and ABTS radical scavenging activity as well as TAC values.
Discussion
The pH of the juices to be used as coagulants determined the quantity required in the preparation of paneer. The lower the pH, the lesser was the quantity of coagulants used for making paneer with the specified quantity of milk. A lower pH value of any coagulant might be due to presence of more citric acid. The musambi juice probably contained least natural citric acid because the maximum quantity of musambi juice was used in the preparation of paneer (700 ml vs., for example, 65 ml of lemon juice: online Supplementary Table S1). Our pH values were similar to published reports (grapefruit juice: Ahmed et al. (Reference Ahmed, Rattanpal, Gul, Dar and Sharma2019); lemon juice: Al-Juhaimi and Ghafoor (Reference Al-Juhaimi and Ghafoor2013); mandarin juice: Manzoor et al. (Reference Manzoor, Hussain, Anjum, Naseer, Ahmad and Ziogas2023) and Nawaz et al. (Reference Nawaz, Abbasi, Khan, Ali, Hasan and Hayat2020)). During our paneer manufacturing, the pH of the whole paneer mixture was adjusted to 5.70–5.80 (coagulation point) irrespective of which juice was being used. This is why different quantities of the citrus juices were used, and meant acidity values in the different fresh paneer matrices were all more or less same. Badola et al. (Reference Badola, Danish, Kumar, Fahad, Kanade, Upadhayay, Kohli and Rautela2018) observed a pH of around 5.65 at the time of milk coagulation during manufacturing of paneer which is a little bit lower than we used.
The °Brix value equates to the total soluble solids of the juice. The freshly squeezed natural juices used here would have contained sugars along with other compounds, including citric acid and other organic acids, whilst the °Brix of citric acid solution was due only to citric acid itself. As for pH, our °Brix values were similar to published data (lemon juice: Aguilar-Hernández et al. (Reference Aguilar-Hernández, Núñez-Gómez, Forner-Giner, Hernández, Pastor-Pérez and Legua2020), Al-Juhaimi and Ghafoor (Reference Al-Juhaimi and Ghafoor2013) and Pham et al. (Reference Pham, Nguyen, Dinh, Kieu, Bach, Phong and Truc2020); grapefruit juice: Ahmed et al. (Reference Ahmed, Rattanpal, Gul, Dar and Sharma2019); kinnow juice: Mahawar et al. (Reference Mahawar, Jalgaonkar, Bibwe, Bhushan, Meena and Sonkar2020); mandarin juice: Manzoor et al. (Reference Manzoor, Hussain, Anjum, Naseer, Ahmad and Ziogas2023) and Nawaz et al. (Reference Nawaz, Abbasi, Khan, Ali, Hasan and Hayat2020)).
The decreasing trend of pH of paneer with storage time is in accordance with the results of Shanaziya et al. (Reference Shanaziya, Mangalika and Nayananjalie2018) and Singh et al. (Reference Singh, Singh and Shakya2014). This might be due to the activities of indigenous microorganisms in paneer, causing more accumulation of acids during the storage period. There was some loss of moisture contents during storage of paneer, thereby increasing the dry matter content. Again, our data are consistent with published reports (Khatkar et al., Reference Khatkar, Ray and Kaur2017). As dry matter of paneer matrix increased during storage, then it positively influenced the protein, fat and ash contents.
The presence of TPC and YMC of the freshly prepared paneers might be due to presence of microbiota in goat's milk. Moreover, microbes may reinfect paneer during post manufacture conditions of handling, packaging and storage. Moreover, no starter or probiotic culture is used in the manufacturing of paneer, and it is recognized that such cultures can inhibit the proliferation of harmful microbes. For this reason, the shelf life of paneer is very low as compared to cheese. The increasing trend of TPC and YMC during storage of paneer as observed in the present study is consistent with the findings of some previous studies (Singh et al., Reference Singh, Singh and Shakya2014; Mishra et al., Reference Mishra, Rao, Bhardwaj, Sutariya, Kavitkar and Subhash2016), whilst the lower TPC and YMC counts in fresh paneer using citrus juices than in control paneer might be due to antimicrobial factors other than acidity. In this respect, lemon juice gave the best outcome.
The sensory attributes usually illustrate the quality characteristics of prepared paneer. López-Villafaña et al. (Reference López-Villafaña, Rojas-González, Elías-Román and Rodríguez-Hernández2023) observed sensory score values above 6.4 for freshly produced goat's milk cheese whilst our initial values were generally above this. Our observation of decreasing sensory attributes during storage agrees with earlier reports (Singh et al., Reference Singh, Singh and Shakya2014 and Das et al., Reference Das, Chauhan, Agrawal, Tomar and Sirajuddin2018). The microbial counts of paneer increased to an unacceptable limit after 15 d of storage. Therefore, paneer stored for 15 d were not included in the present study for sensory evaluation.
The juices from different citrus fruits contain antioxidants and other functional compounds. For example, lemon (Citrus limon) juice contains appreciable quantities of vitamin C (Alfadul and Hassan, Reference Alfadul and Hassan2016), tannins, flavonoids, terpenoids and phenols (Rauf et al., Reference Rauf, Uddin and Ali2014). Grapefruit (Citrus × paradisi) juice has appreciable quantities of flavanones such as naringin and narirutin (Sicari et al., Reference Sicari, Pellicanò, Giuffrè, Zappia, Capocasale and Poiana2018). Mandarins such as Feutrell's Early (Citrus reticulata Blanco) contains appreciable quantities of vitamin c, DPPH radical scavenging activities, total phenolics, flavonoids, flavonols and carotenoids (Nawaz et al., Reference Nawaz, Abbasi, Khan, Ali, Hasan and Hayat2020; Manzoor et al., Reference Manzoor, Hussain, Anjum, Naseer, Ahmad and Ziogas2023). Kinnow (Citrus reticulate L.) juice is rich in vitamin B, vitamin C, β-carotene, minerals and other bioactive compounds (Mahawar et al., Reference Mahawar, Jalgaonkar, Bibwe, Bhushan, Meena and Sonkar2020). The promising results of citrus fruit juices concerning TP and TF contents, FRAP values, ABTS and DPPH radical scavenging activity in the present study might be due to the presence of one or more of these compounds. The TP contents of lemon and kinnow juice obtained in the present study were slightly higher than the contents found by Al-Juhaimi and Ghafoor (Reference Al-Juhaimi and Ghafoor2013) which might be due to geographical variations, and it should be remembered that the antioxidant activity of each citrus juice itself was many fold higher than the WSE of the goat's milk paneer prepared through coagulation of their respective juice, nevertheless the results of DPPH/ABTS and TAC of the latter were very promising. The high antioxidant potential of the juices might be attributed to the presence of high contents of phenolics as well as flavonoids. So, paneers coagulated by citrus juices may retain phenolics and flavonoids in their matrix. However, other factors might be in play as well. Casein protein fractions were reported to be responsible for the antioxidant potential of goat's milk (Mal et al., Reference Mal, Singh, Mane, Sharma, Sharma, Bhar and Dhar2018), as were free amino acids (Biadala and Konieczny, Reference Biadała and Konieczny2018). Hernández-Galán et al. (Reference Hernández-Galán, Cardador-Martinez, Lopez-del-Castillo, Picque, Spinnler and del Campo2017) also reported antioxidant potential of goat's milk owing to the presence of peptides originally present in the milk. In another study, Vázquez-Garcí et al. (Reference Vázquez-García, Cardador-Martínez, Orihuela-López, Ramos-Hernández and Martín-del-Campo2021) also related DPPH radical scavenging activity of Mexican goat cheese to the peptides originally present in goat's milk. Hence, it may be assumed that the antioxidant potential of goat's milk paneer prepared in the present study might be due to presence of different proteins, their peptides and free amino acids, as well as the contribution from the citrus juices.
Most earlier studies reported antioxidant potential of goat's milk cheese but no comprehensive study has previously been done on the antioxidant activity of goat's milk paneer. Goat's milk cheese is usually prepared using specific starter cultures (such as MO-1, LHB02 from Chr. Hansen, Denmark: Tabla et al., Reference Tabla, Gomez, Simancas, Rebollo, Molina and Roa2018) and rennet (50% chymosin and 50% pepsin: Baquero et al., Reference Baquero, Alverez Rios, Rodriguez, Romero and Martin2011) and it is these that are usually responsible for the release of bioactive peptides during production as well as during ripening. Therefore, the antioxidant potential of cheese would be different from paneer because paneer does not contain such kind of starter cultures and rennet. The total polyphenol content of goat's milk cheese obtained from the pasteurized milk of grazing goats was 300 μg GAE/g (Hilario et al., Reference Hilario, Puga, Ocana and Romo2010), similar to our data for paneer coagulated by citrus juices. They observed high concentrations of some phenolic compounds such as caffeic acid, chlorogenic acid and ferulic acid but only minute quantities of the flavonoid compounds, catequine and quercetin. Such bioactive compounds are probably present in paneer, and it may be that the decrease of antioxidant potential during storage might be due to degradation of such compounds in the paneer matrix. Most of the phenolics and flavonoids would be expected to leach into the whey of the paneer, but the higher antioxidant potential of the paneer itself (as opposed to the whey) may be expected due to the presence of casein and its fractions (peptides, amino acids) in the paneer matrix. Whey is the by-product of cheese and paneer and can be easily digested and absorbed. It contains vitamin-binding proteins and bioactive proteins, such as beta-lactoglobulin, alpha-lactalbumin, immunoglobulin, serum albumin, and lactoferrin. These components may exhibit a range of immune enhancing and antioxidant activities (Tsutsumi and Tsutsumi, Reference Tsutsumi and Tsutsumi2013). Dryáková et al. (Reference Dryáková, Pihlanto, Marnila, Čurda and Korhonen2010) and Mann et al. (Reference Mann, Kumari, Kumar, Sharma, Prajapati, Mahboob and Athira2015) also reported that whey protein hydrolysates were responsible for DPPH radical scavenging activity. The whey from all the paneer (except the control paneer) had a good taste presumably due to the presence of more quantities of fruit sugars in addition to lactose. Presence of vitamin C, sugars and bioactive compounds confer improved nutritional value to such whey, which could potentially be converted into valuable infant formulations or drinks. In this respect, preparation of functional milk beverages from whey has also been reported by Mann et al. (Reference Mann, Kumari, Kumar, Sharma, Prajapati, Mahboob and Athira2015).
In conclusion, all the paneers prepared using different citrus juices for coagulation were acceptable concerning sensory attributes. The results of all the antioxidant potential analyses were promising. Citrus fruit juice paneers had greater antioxidant potential than the control paneer, presumably due to the presence of phenolics, flavonoids and other bioactive compounds. Moreover, paneer coagulated by kinnow juice showed the maximum values regarding TF contents, TAC values and DPPH/ABTS radical scavenging activities. It may be recommended that citrus juice paneer as well as its whey should be produced by the dairy industry.
Supplementary material
The supplementary material for this article can be found at https://doi.org/10.1017/S0022029924000190
Acknowledgments
The authors acknowledge the financial support from higher education commission (HEC), Pakistan (under the project No. 10383) for carrying out research activities. This publication was made possible by grant number NC.X-267-5-12-170-1 from the National Institute of Food and Agriculture (NIFA) and the Department of Family and Consumer Sciences and the Agriculture Research Station at North Carolina Agriculture and Technical State University (Greensboro, NC, USA 27411). This work was also supported, in part, by 1890 Capacity Building Program grant no. (2020-38821-31113/project accession no. 021765).
