The investigation aimed at assessing a comparative study on the production and characterization of ACE inhibitory, anti-diabetic, and anti-inflammatory activities, along with the production of ACE inhibitory and anti-diabetic peptides through the fermentation of buffalo and camel milk by Limosilactobacillus fermentum (KGL4) and Saccharomyces cerevisiae (WBS2A). The angiotensin-converting enzyme (ACE) inhibitory and anti-diabetic properties were evaluated at particular time intervals (12, 24, 36, and 48 h) at 37 °C, and we discovered maximum activity at 37 °C after 48 h of incubation. The maximum ACE inhibitory, lipase inhibitory activities, alpha-glucosidase inhibitory, and alpha-amylase inhibitory activities were found in the fermented camel milk (77.96 ± 2.61, 73.85 ± 1.19, 85.37 ± 2.15, and 70.86 ± 1.02), as compared to the fermented buffalo milk (FBM) (75.25 ± 1.72, 61.79 ± 2.14, 80.09 ± 0.51, and 67.29 ± 1.75). Proteolytic activity was measured with different inoculation rates (1.5%, 2.0%, and 2.5%) and incubation times (12, 24, 36, and 48 h) to optimize the growth conditions. Maximum proteolysis was found at a 2.5% inoculation rate and at a 48 h incubation period in both fermented buffalo (9.14 ± 0.06) and camel milk (9.10 ± 0.17). SDS-PAGE and 2D gel electrophoresis were conducted for protein purification. The camel and buffalo milk that had not been fermented revealed protein bands ranging from 10 to 100 kDa and 10 to 75 kDa, respectively, whereas all the fermented samples showed bands ranging from 10 to 75 kDa. There were no visible protein bands in the permeates on SDS-PAGE. When fermented buffalo and camel milk were electrophoresed in 2D gel, 15 and 20 protein spots were detected, respectively. The protein spots in the 2D gel electrophoresis ranged in size from 20 to 75 kDa. To distinguish between different peptide fractions, water-soluble extract (WSE) fractions of ultrafiltration (3 and 10 kDa retentate and permeate) of fermented camel and buffalo milk were employed in RP-HPLC (reversed-phase high-performance liquid chromatography). The impact of fermented buffalo and camel milk on inflammation induced by LPS (lipopolysaccharide) was also investigated in the RAW 264.7 cell line. Novel peptide sequences with ACE inhibitory and anti-diabetic properties were also analyzed on the anti-hypertensive database (AHTDB) and bioactive peptide (BIOPEP) database. We found the sequences SCQAQPTTMTR, EMPFPK, TTMPLW, HPHPHLSFMAIPPK, FFNDKIAK, ALPMHIR, IPAVFK, LDQWLCEK, and AVPYPQR from the fermented buffalo milk and the sequences TDVMPQWW, EKTFLLYSCPHR, SSHPYLEQLY, IDSGLYLGSNYITAIR, and FDEFLSQSCAPGSDPR from the fermented camel milk.

Isabgol (Plantago ovata Forsk.) is an important medicinal plant having high pharmacological activity in its seed husk, which is substantially used in the food, beverages and packaging industries. Nevertheless, isabgol lags behind in research, particularly for genomic resources, like molecular markers, genetic maps, etc. Presently, molecular markers can be easily developed through next generation sequencing technologies, more efficiently, cost effectively and in less time than ever before. This study was framed keeping in view the need to develop molecular markers for this economically important crop by employing a microsatellite enrichment protocol using a next generation sequencing platform (ion torrent PGM™) to obtain simple sequence repeats (SSRs) for Plantago ovata for the very first time. A total of 3447 contigs were assembled, which contained 249 SSRs. Thirty seven loci were randomly selected for primer development; of which, 30 loci were successfully amplified. The developed microsatellite markers showed the amplification of the expected size and cross-amplification in another six species of Plantago. The SSR markers were unable to show polymorphism within P. ovata, suggesting that low variability exists within genotypes of P. ovata. This study suggests that PGM™ sequencing is a rapid and cost-effective tool for developing SSR markers for non-model species, and the markers so-observed could be useful in the molecular breeding of P. ovata.

In this study, camel milk (CM) and Gir cow milk (GCM) were fermented through cofermentation via yeast–lactic cultures, i.e., Lacticaseibacillus rhamnosus (M9, MTCC 25516) and Saccharomyces cerevisiae (WBS2A, MG101828), and their antioxidant and antidiabetic effectiveness were studied. To optimize the growth conditions, the level of proteolysis was evaluated by exploring various inoculation levels (1.5, 2.0 and 2.5%) as well as incubation durations (0, 12, 24, 36 and 48 h). Peptides were extracted and purified through 2D gel electrophoresis as well as SDS–PAGE. Water-soluble extracts (WSEs) of ultrafiltered (UF) peptide fractions were evaluated via reversed-phase high-performance liquid chromatography (RP-HPLC) to identify the peptide segments. By applying the Peakview tool, peptide sequences obtained from liquid chromatography–mass spectrometry (LC/MS) were reviewed by comparison with those in the BIOPEP database. Furthermore, the elevated levels of TNF-α, IL-6, IL-1β and nitric oxide (NO) in RAW 267.4 cells treated with lipopolysaccharide (LPS) are considerably lower than those in cultured CM and GCM. Protein macromolecules in CMs and GCMs have been captured via confocal laser scanning microscopy (CLSM) and Fourier transform infrared (FTIR) spectroscopy both before and after fermentation.