The aim of this present study was to optimize the fermentation conditions (time and temperature) of amasi (a Southern African fermented dairy product) using response surface methodology (RSM), and to determine the physicochemical properties, as well as the microbial composition, using next generation sequencing. Fermentation time and temperature were optimized to produce different amasi samples and different parameters, including pH, total soluble solids (TSS), total titratable acids (TTA), and consistency. All the variables studied were found to show significant (p ≤ 0.05) changes with increasing fermentation time and temperature. Numerical optimization was used to obtain the optimal fermentation conditions for amasi; based on RSM, it was 32 °C for 140 h, while with k-means clustering, it was 25 °C for 120 h. Under both conditions for the optimal samples, the pH reduced from 6.64 to 3.99, TTA increased from 0.02 to 0.11 (% lactic acid), TSS decreased from 9.47 to 6.67 °Brix, and the consistency decreased from 23 to 15.23 cm/min. Most of the identified bacteria were linked to lactic acid bacteria, with the family Lactobacillaceae being the most predominant in amasi, while in raw milk, Prevotellaceae was the most abundant. The fermentation conditions (time and temperature) had a significant influence on the parameters investigated in this study. Results of this study could provide information for the commercialization of quality amasi.

This study investigated the effect of fermentation conditions (time and temperature) on the nutritional composition and health-promoting properties of raw milk, optimized naturally fermented amasi, and amasi fermented with starter culture. The composition of the products, including chemical analysis, amino acids, mineral composition, and fatty acids, as well as health-promoting properties (total phenolic content (TPC), total flavonoid content (TFC), and antioxidant activity assays (2,2-Azinobis (3-Ethyl-Benzothiazone-6-Sulphonic acid) (ABTS) and 2,2-diphenyl-1-picrylhydrazyl (DPPH)) were investigated. The results showed that the fermentation conditions (time and temperature) significantly affected the ash contents of both fermented samples. Fermentation was also observed to have increased the contents of most of the essential and non-essential amino acids. A high concentration of glutamic acid was observed in optimized naturally fermented amasi at 32 °C (OP32, 6.30 g/100 g), followed by starter-culture-fermented amasi at 25 °C (SC25, 5.58 g/100 g) and then starter-culture-fermented amasi at 32 °C (SC32, 5.35 g/100 g), with optimized naturally fermented amasi at 32 °C (OP25, 4.46 g/100 g) having the lowest concentration. A similar trend in the composition of minerals was also observed, with potassium being the most abundant (110.1–134.5 µg/g). The TPC, TFC, and the antioxidant activities were significantly improved by fermentation, while a significant reduction in free fatty acids was recorded. In conclusion, the fermentation conditions had a significant effect on the nutritional and health-promoting properties of amasi. The results provided in this study could provide nutritional information regarding the composition of amasi, which can be beneficial to produce a high-quality product.