Effect of storage duration on frozen inoculum to be used for the in vitro gas production technique in rabbit

The present study aimed to investigate the effect of storage duration of frozen inoculum on fermentation parameters obtained with the in vitro gas production technique. Two non-predigested diets differing in chemical composition and especially crude fibre content (low fibre diet: 13.8%; high-fibre diet: 22.6%) were ground to pass a 1 mm screen and subjected to fermentation with the same inoculum frozen for different periods: after 1 month (inoculum 1), after 2 months (inoculum 2) and after 3 months (inoculum 3). The inoculum used was obtained from the caecal content of 75-day-old NZW rabbits. After defrosting, the caecal content was diluted with the medium 1: 1 (V/V) and squeezed through six layers of gauze to obtain the inoculum. The substrate affected several fermentation parameters. In particular, the high-fibre diet had lower potential and cumulative gas production (A = ml/g 220 vs 256; P < 0.01 and OMCV ml/g 185 vs 221; P < 0.01), lower organic matter degradability (OMl 67.2% vs 58.0%; P < 0.01) and production of volatile fatty acids (mmol/g 56.2 vs 49.8; P < 0.01), and took more time to obtain gas production equal to A/2 (B = h 11.8 vs 10.1; P < 0.01) compared with the low-fibre diet. However, the three inocula had very similar gas production kinetics, overlapping values of degraded organic matter (62.4%, 62.7% and 62.7% respectively for inocula 1, 2 and 3) and similar production of VFA (54.0, 52.2 and 52.8 mmol/g, respectively for inocula 1, 2 and 3). This research showed it is possible to use frozen inoculum for at least 3 months and in this time interval obtain the same parameters of in vitro fermentation.