Method for the determination of 5,5-diphenylbarbituric acid and its separation from 1,3-dimethoxymethyl-5,5-diphenylbarbituric acid in plasma by high-performance liquid chromatography.

A method is described for the measurement of 5,5-diphenylbarbituric acid in plasma using high-performance liquid chromatography with UV detection. Briefly, the compounds are separated on a C18 reversed-phase column using a mobile phase of 50 mM sodium acetate (pH 4.5) and methanol. The flow-rate is 1.0 ml/min and 25 microl are injected and detected at 215 nm. The method is specific and sensitive in the range of concentrations tested, with a limit of quantification of 0.25 microg/ml. The calibration curves are linear for concentrations between 0.25 and 10 microg/ml. Intra-day and inter-day coefficients of variation are less than 8.5 and 10.5%, respectively, over the linear range. Intra-day and inter-day bias are less than 7.0 and 8.0%, respectively. A pharmacokinetic study conducted in male Beagle dogs administered 10 mg/kg of 1,3-dimethoxymethyl-5,5-diphenylbarbituric acid or 8 mg/kg of 5,5-diphenylbarbituric acid intravenously demonstrates the utility of this method.

Early lactational response to supplemental protein by dairy cows fed grass-legume forage.

A total of 24 primiparous and 46 multiparous Holstein cows were in two trials to determine the influence of concentration of dietary protein on performance during early lactation. Treatments were 13.5 or 16.5% dietary crude protein for the first 12 wk of lactation. Complete mixed diets of grass-legume forage and concentrates were fed for ad libitum consumption. Treatment patterns repeated across trials. Yields of milk and 3.5% fat-corrected milk, intake of dry matter, and returns over feed costs were improved by feeding the diet with 16.5% crude protein. Percent milk fat, body weight change, and reproductive performance were unaffected by treatment. The results demonstrate benefit, during early lactation, from supplementing protein to grass-legume based diets containing 13.5% crude protein. Performance responses to increased concentrations of dietary crude protein during early lactation appear greater for multiparous than primiparous cows.

Influence of reducing dietary crude protein from 17 to 13.5 percent on early lactation.

A total of 117 Holstein cows were in two trials to determine how reduction in concentration of dietary protein influences early lactation. Treatments were feeding 17% crude protein for 3 mo or reducing dietary crude protein from 17 to 13.5% after 1st or 2nd mo postpartum. Results were similar in both trials. Reducing concentration of dietary crude protein to 13.5% at 1 mo postpartum tended to decrease intake of dry matter during the 2nd and 3rd mo of lactation. Milk production of multiparous, but not primiparous cows, was decreased. Dry matter intake and milk production did not decrease following a drop in concentration of dietary protein at 2 mo postpartum. Returns over feed costs were reduced by feeding the diet with 17% crude protein to primiparous cows longer than 1 mo or to multiparous cows longer than 2 mo postpartum. Milk composition, body weight change, and reproductive performance were not affected by treatment.

Growth rate and secretion of pituitary hormones in relation to age and chronic treatment with thyrotropin-releasing hormone in prepubertal dairy heifers.

Holstein heifers were treated with synthetic thyrotropin-releasing hormone (TRH) or saline twice daily from one week through 6 mo of age. Plasma concentrations of prolactin (PRL) and thyrotropin (TSH) were elevated (P less than .01) within 30 min after the first TRH injection (1 week of age). At 1 and 3 mo of treatment, PRL and TSH increased in response to TRH, although the TSH response was reduced (P less than .01) as compared to the first day of treatment. Although plasma growth hormone (GH) appeared to be elevated following the first TRH injection, this effect was not statistically significant (P less than .05), nor was it significantly influenced by treatment following subsequent TRH injections. None of the 3 hormones, PRL, TSH or GH, was elevated following the final TRH injection at 6 mo of age. In contrast, plasma concentrations of PRL and TSH were increased in a control heifer injected with TRH at 6 mo. These data indicate that hormonal responsiveness to TRH stimulation decreases with continued twice daily treatment at doses of TRH used in the present studies. Examination of weight gains indicated that chronic treatment with TRH was associated with increased growth rate through 6 mo of age (10.6% increased average daily gains P less than .10), which was exhibited in a steeper slope (P less than .05) of the growth curve in the TRH group. Feed intake was slightly greater in TRH heifers, although feed efficiency (kg feed/kg gain) was not different between the two groups. Plasma concentrations of PRL increased (P less than .01) with age (r = +0.938) in control heifers while plasma TSH and GH were not significantly related to age. This observation establishes a positive correlative relationship between PRL secretion and the approach of puberty in the dairly heirfer. It was also noted that elevation of PRL secretion by TRH treatment was associated with significant advancement of age at first observed estrus (9.4 vs. - 10.5 mo) suggesting that a functional relationship between PRL secretion and puberty may exist in dairy heifers.

Antibacterial host defense: in vitro interaction of bacteria, serum factors, and leukocytes from precolostral dairy calves and their dams.

The bactericidal capacity of serum and peripheral blood leukocytes from newborn dairy calves (before receiving colostrum) and their dams for a strain of Escherichia coli and Staphylococcus aureus has been investigated. Comparative studies were conducted, using either unheated (freshly collected) and heated serums alone or in combination with cells from adult and newborn animals. Neither serum from newborn calves was able to kill either microorganism. Both microorganisms were sensitive to bactericidal activity of antibody and complement in unheated serum from adult cattle. On heating, serum from adult cattle lost bactericidal properties of the antibody-complement system. A cellular deficiency in phagocyte function in cells of newborn dairy calves was not evident, since they were as able to kill E coli and S aureus as cells from adult cattle, when incubated in the presence of the same opsonic source. Bactericidial activity was not evident when cells from newborn calves were incubated with heated autologous calf serum and S aureus or E coli. Unheated, autologous calf serum combined with cells from newborn calves exerted bactericidal capacity against S aureus but not E coli. The bactericidal capacity against S aureus of cells from newborn calves incubated with unheated serum from newborn calves did not differ from heated serum from adult cattle. However, there was a difference with E coli. The reasons for this phenomenon have not been determined, but may relate to natural, specific immunity of the neonate. Immunoglobulin and antibody concentrations in serums from calves and dams were determined. Most of the dams had bacterial agglutination titers to both microorganisms; the calves had either undetectable or low levels of antibody.