Nitritocobalamin and nitrosocobalamin may be confused with sulfitocobalamin using cation-exchange chromatography.

The previously reported ability of SP-Sephadex C25 column chromatography for partitioning biologically important cobalamins has been modified to include analytical separation of nitritocobalamin (NO2-Cbl) and nitrosocobalamin (NO-Cbl). Gel column dimensions (1.5 x 11.0 cm), a low eluent flow-rate (125 microliters/min), collection of small eluate fractions (160 microliters) plus maintenance of He saturated mobile and gel phases all combined to eliminate ordinarily confusing proximal elution of NO2-Cbl and NO-Cb1 with sulfitocobalamin (SO3-Cbl) and cyanocobalamin. Cobalamin elution profiles from the gel column were monitored by direct radiometric analysis of 57Co-labelled cobalamin standards or competitive intrinsic factor radioassays for cobalamin sample sizes up to 10.0 ng. Failure to implement the chromatographic conditions detailed here totally obscured analysis of NO2-Cbl coexisting with SO3-Cbl in brain tissues for chicks exposed to dietary sulfites and caused oversight of NO-Cb1 normally coexisting in prepared NO2-Cbl standards.

Optimization of protoplast formation, regeneration, and viability in Microsporum gypseum.

Factors affecting high yields, regeneration frequencies, and viability of protoplasts from clonal cultures of Microsporum gypseum were investigated. Maximum yields of protoplasts were obtained after 6 hrs digestion of 2-4 days old mycelium with Novozyme 234 using CaCl2 (0.4 M) as an osmotic stabilizer and glycine + HCl (pH 4.5) as the buffer system. Mercaptoethanol + dithiothreitol (0.01 M) proved to be the best pretreatment of mycelium prior to digestion with enzyme. A regeneration frequency of 94.4% was obtained using the top agar method with complete medium (pH 6.5) containing 0.5% agar and 0.4 M CaCl2 as an osmoticum. Colonies from regenerated protoplasts on medium containing CaCl2 were pigmented and completely powdery with high sporulation. Protoplast viability was studied in osmotic stabilizer supplemented with glucose or glutamine. After 24 hrs, glucose (2%) and glutamine (2%) enhanced protoplast viability by 22% and 23%, respectively. Protein synthesis, as measured by 3H-lysine uptake, matched the viability profile determined by fluorescence microscopy.

Permeability barriers to cytochrome-c in nerves of adult and immature rats.

Nerves in the tongues of adult and immature rats were examined with respect to their permeability to exogenous cytochrome-c (mol wt 12,000) injected into the tongue. The distribution of cytochrome-c was determined in cryostat sections on the basis of the peroxidase activity of this protein. Nerves of 14-day-old rats were permeable to injected cytochrome-c. The larger nerves of older animals showed only localized accumulations of cytochrome-c reaction product both between and within axons adjacent to endoneurial blood vessels. Reaction product was not found, however, in association with blood vessels penetrating nerves of the tongue that were not within the limits of tracer spread. In the smallest nerve brances, thin linear strands of reaction product filled the interstices between the nerve fibers.

Central nervous system: recovery of function.

Rats with one- or two-stage lesions in the hippocampus, anzygdala, or frontal cortex were tested on a variety of tasks. Even though locus and amount of tissue damage were very similar in the two groups, animals with one-stage lesions were impaired as judged by performance on all problems, while aninmals with two-stage lesionis were not.

Pemoline levels in brain: enhancement by dimethyl sulfoxide.

Pemoline-C(14) dissolved in dimethyl sulfoxide and injected intraperitoneally into rats was found in larger amounts in the brain than was a similar dose given in 0.3 percent tragacanth suspension. This appeared to be related to a partial breakdown of the blood-brain barrier in vivo by the dimethyl sulfoxide.