To study the coagulation performance of rennet from Bacillus velezensis DS-1, we evaluated the effect of its different dosages (9、12、15、18、21 and 24 U/mL) on the turbidity, viscosity, rheology, optical density at 500 nm (OD_{500 nm}) and microstructure of curd. To this end, commercial rennet was used as control. The results showed that when the rennet dosage ranged from 9 to 21 U/mL, the viscosity of coagulated milk initially increased and then leveled off, and the turbidity increased with increasing curdling time. When the rennet dosage was 24 U/mL, the turbidity increased at first and then decreased with increasing curdling time. The rheological characteristics showed that peak storage modulus of milk coagulated with rennet from strain DS-1 at dosages below 18 U/mL was smaller than that with commercial rennet and it gradually increased with increasing dosage of DS-1 rennet. As the dosage rose from 9 to 21 U/mL, the water-holding capacity increased significantly (P < 0.05). The OD_{500 nm} value declined first and then increased slightly with increasing rennet dosage. The microstructure analysis showed that casein particles were more densely with increasing rennet dosage, and milk curd formed by rennet from Bacillus velezensis DS-1 exhibited a more porous structure. The results of this study demonstrate that Bacillus velezensis DS-1 rennet has good coagulation performance, showing minimal differences in coagulation parameters compared with commercial rennet.

This study was conducted to reveal the effect of milk-clotting enzyme (MCE) produced by Bacillus licheniformis on the texture of Cheddar cheese during ripening. Cheddar cheese and Cheddar cheese analogue were prepared with the MCE and designated as C_DF and C_D3, respectively, and commercial chymosin was used as a control (C_CF). The changes in the texture, rheology and microstructure of cheese during ripening were analyzed. The results were as follows: The hardness of the three cheeses increased with maturation time, while the elasticity and chewiness presented a downward trend. The hardness, springiness and chewiness of C_DF were the lowest. The scanning electron microscope (SEM) results showed that the compactness of the protein structure decreased with increasing maturity, and the structure of C_DF was looser than that of C_CF. The rheological results showed that the storage modulus and loss modulus decreased with increasing ripening time up to six months and increasing sweep temperature. The tan δ value decreased during the ripening process, indicating that the fluidity of cheese decreased with increasing ripening time. The fluidity and fusibility of C_DF were better than those of the other groups. The above results indicated that the mechanical properties of cheese made with Bacillus licheniformis MCE were better than those made with commercial rennet, while the latter had harder texture and denser protein network structure.

The present study was designed to evaluate the effect of the chymosin from Bacillus licheniformis on the proteolysis of Cheddar cheese during ripening. The chymosin was used to produce cheddar cheese (C_DF) and Cheddar cheese analogue (C_D3), and a commercial milk-clotting enzyme preparation was also used to prepare Cheddar cheese (C_CF). The variation of proteolysis indexes was analyzed during cheese ripening. The results showed that the contents of casein, soluble nitrogen at pH 4.6 (pH 4.6-SN), 12% trichloroacetic acid-soluble nitrogen (12% TCA-SN), 5% phosphotungstic acid-soluble nitrogen (PTA-SN) and total free amino acid in C_DF, C_D3 and C_CF increased significantly with maturation time, and were significantly higher in C_DF than in C_CF during maturation (P < 0.05). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis showed that the degree of hydrolysis of α-casein (α-CN) was higher in C_DF. The results of pH 4.6-SN analysis showed that the total peptide content increased first and then decreased with ripening time, but the ratio of hydrophobic peptide to hydrophilic peptide showed a continuous downward trend. At the sixth month of ripening, the ratios of hydrophobic peptide to hydrophilic peptide in C_DF, C_D3 and C_CF were 2.668, 2.822 and 3.788, respectively. Principal component analysis (PCA) showed that for all three cheeses, the proteolysis degree was positively correlated withmaturity, but negatively correlated with the ratio of hydrophobic peptides to hydrophilic peptides. The above results showed that cheese produced with Bacillus licheniformis chymosin had higher degree of proteolysis, but smaller proportion of hydrophobic peptides. Our findings could provide a theoretical basis for the application of Bacillus licheniformis chymosin in cheese production.

Tenebrio molitor rennet (TMR) was prepared and the effects of preheating milk at different temperatures on its coagulation behavior and hydrolytic properties were evaluated, and the structure of milk curds was characterized by Fourier transform infrared (FTIR) spectroscopy and laser scanning confocal microscopy (LSCM). The results showed that the water-holding capacity of curds increased significantly with the increase in preheating temperature (P < 0.05), while the particle size, apparent viscosity and storage modulus G´ increased initially and then decreased (P < 0.05); the particle size and apparent viscosity reached their maximum of 15.07 and 0.51 at 45 ℃, respectively. The production of casein glycomacropeptide (CGMP) and the degree of hydrolysis (DH) were not significantly changed with increasing temperature from 45 to 55 ℃ (P > 0.05). The secondary structure and microstructure indicated that the flexibility, stability and compactness of curds were the best at 45 ℃. At this temperature, the water-holding capacity and whey OD value of TMR-coagulated curd tended to be stable when compared with that coagulated by commercial rennet (CR), and the particle size significantly increased. Meanwhile, TMR-coagulated curd exhibited a continuous, irregular and compact network structure, indicating good coagulation properties. Therefore, the optimum coagulation temperature for TMR was 45 ℃. This study provides a theoretical basis and reference for the development and application of new insect-derived rennet.