Fractionation of nitrogen isotopes by mixed ruminal bacteria

Mixed ruminal bacteria were cultured with glucose, cellulose or no carbohydrate, and ammonium bicarbonate or casein hydrolysate. Changes in amounts of bacterial ammonia and non-ammonia N were measured. Ratios of N isotopes expressed as delta 15N (delta 15N) were measured by isotope ratio mass spectrometry. When bacteria were cultured with glucose and ammonium bicarbonate, bacterial delta 15N decreased from .9 to -5.8/1000 and residual ammonia delta 15N increased from -1.4 to 12.7/1000. Fractionation of N isotope occurred during ammonia incorporation because the difference between delta 15N of ammonia and delta 15N of bacteria (delta 15N) was 18.8/1000 (P < .01). However, when casein hydrolysate was the N source, delta 15N between non-ammonia and bacteria averaged only 1.3/1000 (P > .1), indicating no fractionation of N isotopes occurred during utilization of amino acids. The amount of bacterial N was highest at 24 h of incubation when cellulose was the carbohydrate source. At that time, delta 15N between ammonia and bacteria was 8.9/1000 when ammonia was the N source, but delta 15N between non-ammonia and bacteria was 1.7/1000 when casein hydrolysate was the N source. Bacterial N decreased after 24 h when cellulose was the source of carbohydrate. Results indicate that fractionation of N isotopes occurred during ammonia incorporation, but not during incorporation of N from amino acids, deamination, and release of ammonia. Fractionation of N isotopes during incorporation of ammonia N may be used as a marker to study N metabolism by ruminal bacteria.