The associative effects (AE) between concentrate (C), peanut shell (P) and alfalfa (A) were investigated by means of an automated gas production (GP) system. The C, P and A were incubated alone or as 40 : 60 : 0, 40 : 45 : 15, 40 : 30 : 30, 40 : 15 : 45, 40 : 0 : 60 and 30 : 70 : 0, 30 : 55 : 15, 30 : 40 : 30, 30 : 25 : 45, 30 : 10 : 60, 30 : 0 : 70 mixtures where the C : roughage (R) ratios were 40 : 60 and 30 : 70. Samples (0.2000 ± 0.0010 g) of single feeds or mixtures were incubated for 96 h in individual bottles (100 ml) with 30 ml of buffered rumen fluid. GP parameters were analysed using a single exponential equation. After incubation, the residues were used to determine pH, dry matter digestibility (DMD), organic matter digestibility (OMD), volatile fatty acids (VFA) and ammonia nitrogen (NH₃-N) of the incubation fluid, and their single factor AE indices (SFAEI) and multiple-factors AE indices (MFAEI) were determined. The results showed that group of 30 peanut shell had higher SFAEI of GP_{48 h}, DMD, OMD and total volatile fatty acids (p < 0.05) and MFAEI (p < 0.05) than groups 60, 45 and 0 when C : R was 40 : 60. The group of 10 peanut shell showed higher SFAEI of GP_{48 h}, DMD and OMD (p < 0.05) than groups 70, 55 and 40 and MFAEI (p < 0.01) when C : R was 30 : 70. It is concluded that optimal SFAEI and MFAEI were obtained when the C : P : A ratios were 40 : 30 : 30 and 30 : 10 : 60.

This study aimed to determine the associative effects (AEs) of 28 feed combinations of concentrate/soybean pod/alfalfa at different concentrate-roughage ratios that were incubated for 72 h in single tubes (120 ml) which were added 30 ml rumen buffered fluid. The gas production (GP) at 0, 2, 4, 6, 9, 12, 24, 36, 48, 72 h was recorded. A single exponential equation was applied to calculate the GP parameters a (rapid GP), b (slow GP), a + b (GP potential) and c (rate constant of slow GP that can reflect the specific process of GP, rapid and slow GP and GP rate). The AEs were calculated by 72 h GP and weighted estimation value of each combination. After 72 h incubation, pH, volatile fatty acids (VFA) and ammonia nitrogen (NH₃–N), dry matter digestibility (DMD), organic matter digestibility (OMD) were determined the incubation fluid and residues. The single-factor AE index (SFAEI) and multiple-factor AE index (MFAEI) were computed. The results showed that the groups 50 : 50 : 0, 40 : 60 : 0, 60 : 20 : 20, 60 : 10 : 30, 50 : 30 : 20, 50 : 20 : 30, 40 : 50 : 10, 30 : 55 : 15, 30 : 40 : 30, 20 : 65 : 15, 20 : 50 : 30 had higher GP_72 h, a, b, DMD, OMD, NH₃–N, in addition, higher AE of GP, DMD, OMD, total VFA and NH₃–N than those of the other groups (P < 0.05 or P < 0.01), especially the group 30 : 55 : 15 was optimal. In conclusion, in vitro data reveal reliable fermentability and the highest SFAEI and MFAEI occurred when concentrate, soybean pod and alfalfa were combined at the ratios of 50 : 50 : 0, 40 : 60 : 0, 60 : 20 : 20, 60 : 10 : 30, 50 : 30 : 20, 50 : 20 : 30, 40 : 50 : 10, 30 : 55 : 15, 30 : 40 : 30, 20 : 65 : 15, 20 : 50 : 30.

This study aimed to evaluate the multiple-factor associative effects (AEs) of concentrate (C) to sunflower straw (SS) to alfalfa (A) ratios, concentrate to sunflower plate (SP) to alfalfa ratios, concentrate to sunflower seed shell (SSS) to alfalfa ratios at 40 : 60 : 0, 40 : 45 : 15, 40 : 30 : 30, 40 : 15 : 45, 40 : 0 : 60 and 30 : 70 : 0, 30 : 55 : 15, 30 : 40 : 30, 30 : 25 : 45, 30 : 10 : 60, 30 : 0 : 70, respectively, by using an in vitro gas production (GP) method. Thirty-three feed combinations and nine single feeds C, SS, A; C, SP, A; C, SSS, A were incubated respectively for 72 h in a GP tube. A total of 42 treatment combinations were tested, including 33 feed mix combinations [(5 + 6) × 3] and nine single raw materials, a total of 42 sample culture tubes. Each sample culture tube was repeated 20 times, totalling 840 (42 × 20) sample culture tubes. In this study, a total of 12 batches of in vitro culture tests were carried out. Seventy sample tubes and six blank tubes were cultured in each batch (eliminating systematic errors). A total of 912 sample culture tubes were used. The GP values of 0, 2, 4, 6, 9, 12, 24, 36, 48, 72 h were recorded. The GP parameters a, b, c, a + b were calculated by a single exponential equation. The AE values were calculated based on 72 h GP and weighted estimation value of 33 feed combinations. In the incubation fluid pH, ammonia nitrogen (NH₃–N) and volatile fatty acids (VFA) after 72 h incubation were determined and dry matter digestibility (DMD), organic matter digestibility (OMD) were measured in the residue. The single-factor associative effects index (SFAEI) and multiple-factor associative effects index were calculated. The results indicated that groups 40 : 45 : 15 (C:SS:A), 30 : 25 : 45 (C : SS : A), 40 : 60 : 0 (C : SP : A), 40 : 45 : 15 (C : SP : A), 30 : 40 : 30 (C : SP : A), 40 : 45 : 15 (C : SSS : A), 40 : 30 : 30 (C : SSS : A), 30 : 25 : 45 (C : SSS : A) and 30 : 10 : 60 (C : SSS : A) had higher a, b, GP_72h, NH₃–N, DMD, OMD, moreover, higher SFAEI (the AE of GP_72h, NH₃–N, total VFA, DMD and OMD) than other groups (P < 0.05). It was concluded that the optimal feed combinations occurred when concentrate/sunflower straw/alfalfa ratios were 40 : 45 : 15 and 30 : 25 : 45; concentrate/sunflower plate/alfalfa ratios were 40 : 60 : 0, 40 : 45 : 15 and 30 : 40 : 30; concentrate/sunflower seed shell/alfalfa ratios were 40 : 45 : 15, 40 : 30 : 30, 30 : 25 : 45 and 30 : 10 : 60.