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1. Chloroplasts isolated from spinach leaves by using the low-ionic-strength buffers of Nakatani & Barber [(1977) Biochim. Biophys. Acta.461, 510-512] had higher rates of HCO(3) (-)-dependent oxygen evolution (up to 369mumol/h per mg of chlorophyll) and higher rates of [1-(14)C]acetate incorporation into long-chain fatty acids (up to 1500nmol/h per mg of chlorophyll) than chloroplasts isolated by using alternative procedures. 2. Acetate appeared to be the preferred substrate for fatty acid synthesis by isolated chloroplasts, although high rates of synthesis were also measured from H(14)CO(3) (-) in assays permitting high rats of photosynthesis. Incorporation of H(14)CO(3) (-) into fatty acids was decreased by relatively low concentrations of unlabelled acetate. Acetyl-CoA synthetase activity was present 3-4 times in excess of that required to account for rates of [1-(14)C]acetate incorporation into fatty acids, but pyruvate dehydrogenase was either absent or present in very low activity in spinach chloroplasts. 3. Rates of long-chain-fatty acid synthesis from [1-(14)C]acetate in the highly active chloroplast preparations, compared with those used previously, were less dependent on added cofactors, but showed a greater response to light. The effects of added CoA plus ATP, Triton X-100 and sn-glycerol 3-phosphate on the products of [1-(14)C]acetate incorporation were similar to those reported for less active chloroplast preparations. 4. Endogenous [(14)C]acetyl-CoA plus [(14)C]malonyl-CoA was maintained at a constant low level even when fatty acid synthesis was limited by low HCO(3) (-) concentrations. Endogenous [(14)C]acyl-(acyl-carrier protein) concentrations increased with increasing HCO(3) (-) concentration and higher rates of fatty acid synthesis, but were slightly lower in the presence of Triton X-100. It is proposed that rates of long-chain-fatty acid synthesis in isolated chloroplasts at saturating [1-(14)C]acetate concentrations and optimal HCO(3) (-) concentrations may be primarily controlled by rates of removal of the products of the fatty acid synthetase.

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A potent cellulase solution was prepared from culture filtrates of an artificiallyproduced mutant of Trichoderma species. The filtrates were diluted to provide a standardised, simulated rumen liquor which was then used to study the correlation between cellulase digestibility and in-vivo digestibility of a range of plant materials. Cell walls of whole, dried plant material were either not attacked by the cellulase or were attacked only very slowly, but cell walls isolated by neutral-detergent extraction were readily hydrolysed. Cellulase digestibility, defined as the percentage of whole, dry plant material solubilised by neutral-detergent extraction followed by exhaustive hydrolysis with standardised cellulase, was highlycorrelatedwith in-vivo dry matter digestibility (DMD) (r = 0.98) and predicted that parameter with reasonable accuracy (r.s.d,, residual standard deviation = 2.83). The form of the regression equation was in-vivo DMD=0.98 x cellulase solubility-10.12, suggesting that the same factors limited cellulase and in-vivo digestibility. The method was simple and reliable and results were known within 48 h.