Assay of the concentration and 13C-isotopic enrichment of malonyl-coenzyme A by gas chromatography-mass spectrometry.

We developed gas chromatography-mass spectrometry assays for the concentration and mass isotopomer distribution of malonyl-CoA in tissues. The assay involves perchloric acid extraction of the tissue, spiking the extract with [U-13C3]malonyl-CoA or dimethylmalonyl-CoA internal standard, isolation of short-chain acyl-CoA fraction on an oligonucleotide purification cartridge, alkaline hydrolysis to malonate, trimethylsilyl derivatization, and analysis of the mass isotopomer distribution of malonate. The assay was applied to labeling of malonyl-CoA from various [13C]substrates in perfused rat livers and hearts. In livers perfused with [1,2-13C2]acetate, malonyl-CoA is doubly labeled from [1,2-13C2]acetate and singly labeled from 13CO2. In livers perfused with either NaH13CO3 or [3-13C]lactate + [3-13C]pyruvate, the half-lives of singly labeled malonyl-CoA were less than 20 s and 6.95 min, respectively. In rat heart, the half-life of malonyl-CoA, traced with NaH13CO3, was about 1.25 min. Thus, our assay allows us to measure the turnover of tissue malonyl-CoA, the contribution of various [13C]substrates to its production in lipogenic and nonlipogenic organs, and the cycling between acetyl-CoA and malonyl-CoA in nonlipogenic organs.

The effects of acid precipitation on amphibians.

: A large number of studies conclusively demonstrate that low pH conditions have important ecological effects on amphibian communities. These effects vary developmentally and depend on complex interactions with other physical, chemical, and biological parameters. Acid sensitivity varies widely within and among species of amphibians. In spite of the importance of acidity to their reproductive success and the presence of many acidic amphibian habitats, there is, at present, little direct evidence that acid precipitation has produced widespread declines in amphibian populations. However, the potential role of sublethal effects of acidity on amphibian declines has not been studied thoroughly.

Allozyme heterozygosity and growth in the tiger salamander, Ambystoma tigrinum.

Statistically significant, positive correlations were found between individual heterozygosity at seven protein polymorphisms and size in five natural populations of tiger salamanders. When these same populations were sampled later in the same year, the positive associations between heterozygosity and size were no longer evident. A pair mating was conducted in the laboratory, and offspring were distributed randomly to four replicate populations maintained in the laboratory. In two to these four replicates, a significant, positive association between protein heterozygosity and size was observed. The laboratory results suggest that the associations seen in natural populations are due to associations between protein heterozygosity and growth rate early in the larval period.

The distribution of individual heterozygosity in natural populations.

Estimation of the distribution of the level of individual heterozygosity within natural populations is explored with both Monte-Carlo simulation studies and data from natural populations. Simulations indicate that heterozygosities estimated from as few as a dozen randomly chosen loci may, to some degree, reflect (r = 0.35) heterozygosity determined by 100 independent loci. The shape of the expected distribution of heterozygosity is heavily dependent upon levels of heterozygosity at the loci. Complete genetic data for 12 loci from 997 Fundulus heteroclitus are used to describe the distributions of heterozygosity for different localities, for age classes and for sexes. The distributions deviate from normality. Distributions from different localities are not different, but the distributions are heterogeneous among age classes at one of two localities and are heterogeneous between the sexes.