Production, physiological response, and calcium and magnesium balance of lactating Holstein cows fed different sources of supplemental magnesium with or without ruminal buffer.

The objective of this study was to evaluate the effects of dietary replacement of magnesium oxide (MgO) with calcium-magnesium hydroxide [CaMg(OH)2] and its interaction with ruminal buffer (sodium sesquicarbonate) supplementation on production, Ca and Mg balance, and overall physiological response of mid-lactation Holstein dairy cows. Sixty cows averaging 40.5 ± 7.0 kg of milk/d were used. Treatments were assigned following a 2 × 2 factorial arrangement: (1) MgO, (2) MgO + buffer, (3) CaMg(OH)2, or (4) CaMg(OH)2 + buffer. Diets were formulated to have 16.5% of crude protein, 1.82 Mcal/kg of net energy for lactation, 0.67% Ca, 0.39% P, and 0.25% Mg, all on a dry matter (DM) basis. Treatments were individually top dressed. Milk production, composition, and DM intake were evaluated. A subsample of 20 cows were randomly selected for the evaluation of Ca and Mg balance, blood gases, and electrolytes. Ruminal fluid was also collected for evaluation of pH and Ca and Mg solubility. Effects of Mg source, buffer, and the interaction Mg source × buffer were analyzed through orthogonal contrasts. An interaction of Mg source × buffer was found for DM intake and feed efficiency, in which cows fed CaMg(OH)2 had a similar feed efficiency regardless of ruminal buffer inclusion; however, when cows were fed MgO, the inclusion of buffer reduced feed efficiency. No effects on body weight and milk yield were observed. Buffer addition tended to increase the concentrations of fat, protein, and solids-not-fat, without affecting the yields of these milk components. Magnesium source and buffer did not affect ruminal fluid, blood, urine, or fecal pH; however, buffer supplementation increased urinary pH. Treatment with CaMg(OH)2 increased blood concentration of HCO3-, total CO2, and base excess compared with cows fed MgO. No differences were observed in the ruminal solubility of Ca and Mg or on milk or urinary Ca and Mg excretion. Greater plasma Mg concentration was observed for animals fed MgO compared with cows fed CaMg(OH)2; however, both sources were above the threshold recommended in the literature for dairy cows. Also, a reduction in fecal Mg excretion was observed in animals fed CaMg(OH)2. In summary, we provide evidence that CaMg(OH)2 could replace MgO without affecting performance, overall physiological response, or Ca and Mg balance of mid-lactating dairy Holstein cows.

Effects of exogenous amylolytic or fibrolytic enzymes inclusion on in vitro fermentation of lactating dairy cow diets in a dual-flow continuous-culture system.

The objective of this study was to determine the effects of including exogenous amylolytic or fibrolytic enzymes in a diet for high-producing dairy cows on in vitro ruminal fermentation. Eight dual-flow continuous-culture fermentors were used in a replicated 4 × 4 Latin square. The treatments were control (CON), a xylanase and glucanase mixture (T1), an α-amylase mixture (T2), or a xylanase, glucanase, and α-amylase mixture (T3). Treatments were included at a rate of 0.008% of diet dry matter (DM) for T1 and T2 and at 0.02% for T3. All treatments replaced the equivalent amount of soybean meal in the diet compared with CON. All diets were balanced to have the same nutrient composition [30.2% neutral detergent fiber (NDF), 16.1% crude protein (CP), and 30% starch; DM basis], and fermentors were fed 106 g/d divided into 2 feedings. At each feeding, T2 was pipetted into the respective fermentor and an equivalent amount of deionized water was added to each fermentor to eliminate potential variation. Experimental periods were 10 d (7 d for adaptation and 3 d for sample collection). Composite samples of daily effluent were collected and analyzed for volatile fatty acids (VFA), NH3-N, and lactate concentrations, degradability of DM, organic matter, NDF, CP, and starch, and flow and metabolism of N. Samples of fermentor contents were collected from each fermentor at 0, 1, 2, 4, 6, and 8 h after feeding to determine kinetics of pH, NH3-N, lactate, and VFA concentrations over time. All data were analyzed using PROC GLIMMIX of SAS (SAS Institute Inc.), and the repeated variable of time was included for kinetics measurements. Treatment did not affect mean pH, degradability, N flow and metabolism, or the concentrations of VFA, NH3-N, or lactate in the effluent samples. Treatment did not affect pH, acetate:propionate ratio, or the concentrations of lactate, NH3-N, total VFA, acetate, propionate, butyrate, isobutyrate, valerate, or caproate. However, the concentration of total VFA tended to change at each time point depending upon the treatment, and T2 tended to have a greater proportion of 2-methylbutyrate and isovalerate than CON, T1, or T3. As 2-methylbutyrate and isovalerate are branched-chain VFA that are synthesized from branched-chain amino acids, T2 may have an increased fermentation of branched-chain amino acids or decreased uptake by fibrolytic microorganisms. Although we did not observe changes in N metabolism due to the enzymes, there could be changes in microbial populations that utilize branched-chain VFA. Overall, the tested enzymes did not improve in vitro ruminal fermentation in the diet of high-producing dairy cows.

Structure of a ribulose 5-phosphate 3-epimerase from Plasmodium falciparum.

The crystal structure of Pfal009167AAA, a putative ribulose 5-phosphate 3-epimerase (PfalRPE) from Plasmodium falciparum, has been determined to 2 A resolution. RPE represents an exciting potential drug target for developing antimalarials because it is involved in the shikimate and the pentose phosphate pathways. The structure is a classic TIM-barrel fold. A coordinated Zn ion and a bound sulfate ion in the active site of the enzyme allow for a greater understanding of the mechanism of action of this enzyme. This structure is solved in the framework of the Structural Genomics of Pathogenic Protozoa (SGPP) consortium.

Phase I trial of micronized formulation carboxyamidotriazole in patients with refractory solid tumors: pharmacokinetics, clinical outcome, and comparison of formulations.

PURPOSE: Cytostatic agents targeted against angiogenesis and tumor cell invasive potential form a new class of investigational drugs. Orally administered carboxyamidotriazole (CAI) (NSC609974) is both antiangiogenic and antimetastatic. An encapsulated micronized powder formulation has been developed to optimize CAI administration. A phase I dose escalation trial with pharmacokinetic analysis has been performed. PATIENTS AND METHODS: Twenty-one patients with refractory solid tumors and good end organ function and performance status were enrolled onto the study. Patients received a test dose followed 1 week later by daily administration of CAI in the encapsulated micronized formulation at doses of 100 to 350 mg/m2. Patients remained on CAI until disease progression or dose-limiting toxicity. Plasma samples were taken to characterize the pharmacokinetic parameters of this formulation of CAI. RESULTS: All patients were assessable for toxicity and 18 were assessable for pharmacokinetics and response analysis. Grade 1 and 2 gastrointestinal side effects were observed in up to 50% of patients. Dose-limiting toxicity was observed in both patients treated at 350 mg/m2/d, consisting of reversible grade 2 to 3 cerebellar ataxia (n = 1) and confusion (n = 1). One minor response (MR) was observed in a patient with renal cell carcinoma and another nine patients had disease stabilization (MR + SD = 47%). Pharmacokinetic analysis demonstrated reduced bioavailability (58% reduction) compared with the PEG-400 liquid formulation previously reported. CONCLUSION: The better toxicity profile of encapsulated micronized CAI with similar frequency of disease stabilization and ease of administration compared with the liquid or gelatin capsule, suggests that the micronized formulation is a preferable formulation for subsequent studies. A dose of 300 mg/m2/d is proposed for phase II investigations.

Pharmacokinetics of orally administered carboxyamido-triazole, an inhibitor of calcium-mediated signal transduction.

Carboxyamido-triazole (CAI), inhibits proliferation, invasion, and metastatic potential of a number of cancer cell lines at concentrations greater than 0.4 microgram/ml. The objective of this study was to characterize the pharmacokinetic profile from the first Phase I clinical trial of CAI for the single test dose and multiple daily dosing schedule. Two different p.o. formulations (liquid and gelcap) of CAI were administered. Thirty-nine patients with cancer were enrolled. The dose escalation schema was 100, 125, and 150 mg/m2/day and 200 and 330 mg/m2 every other day of the liquid formulation, plus 100 and 125 mg/m2/day and 200 mg/m2 every other day of the gelcap. The CAI pharmacokinetics are best described by a two-compartment open linear model. The gelcap was more rapidly absorbed than the liquid [time to maximum plasma concentration (Tmax) = 2.06 +/- 1.02 versus 5.31 +/- 3.59 h, P2 = 0.0012] which resulted in higher peak plasma concentrations. There was no evidence of saturable elimination as the dose was increased. The mean steady-state peak concentration was 5.1 +/- 1.0 microgram/ml for the 150 mg/m2/day multiple daily dosing regimen. The terminal half-life of CAI was relatively prolonged, 111 h, and the total body p.o. clearance was low (1.87 liters/h). The peak concentration for all dose levels explored was greater than the targeted concentration suggested by in vitro data for activity. Thus, these data suggest that an effective cytostatic exposure of CAI may be obtained with daily or every other day dosing without severe toxicity.

Collaborative atmosphere: rectal pouching in the burn center.

Collaborative practice between nursing specialties opens up opportunities for expanded and improved quality care. Rectal pouching in the burn center is one of these options; it was used in this case with a patient who received cultured epithelial autograft. Pouching practices from the literature are reviewed. A description of rectal pouching to provide stool containment and protection of the wound bed is detailed. Problems with the pouching procedure and potential solutions are also discussed. A summary statement of this highly successful method of stool containment, which is used to guard against wound infection, concludes the article.

Inhibition of testicular steroid metabolism by administration of 1-aminobenzotriazole to rats.

Effects of 1-aminobenzotriazole (ABT) on testicular steroid metabolism were evaluated in rats. Administration of ABT to adult male rats caused dose-dependent decreases in testicular microsomal and mitochondrial cytochrome P450 concentrations. Significant losses of P450 occurred within 8 h of ABT treatment. Accompanying the declines in testicular P450 content were decreases in microsomal 17 alpha-hydroxylase and mitochondrial cholesterol sidechain cleavage activities. Incubation of testicular microsomes or mitochondria in vitro with ABT plus an NADPH-generating system had no effect on P450 concentrations or on rates of steroid metabolism. By contrast, incubation of hepatic microsomes with ABT under the same conditions decreased P450 levels and xenobiotic-metabolizing activity. The results indicate that ABT in vivo causes inactivation of steroidogenic P450 isozymes in the testis, but the mechanism of inactivation differs from that on xenobiotic-metabolizing isozymes.

Cloning and nucleotide sequence of the type E staphylococcal enterotoxin gene.

The gene for staphylococcal enterotoxin type E (entE) was cloned from Staphylococcus aureus into plasmid vector pBR322 and introduced into Escherichia coli. A staphylococcal enterotoxin type E-producing E. coli strain was isolated. The complete nucleotide sequence of the cloned structural entE gene and the N-terminal amino acid sequence of mature staphylococcal enterotoxin type E were determined. The entE gene contained 771 base pairs that encoded a protein with a molecular weight of 29,358 which was apparently processed to a mature extracellular form with a molecular weight of 26,425. DNA sequence comparisons indicated that staphylococcal enterotoxins type E and A are closely related. There was 84% nucleotide sequence homology between entE and the gene for staphylococcal enterotoxin type A; these genes encoded protein products that had 214 (83%) homologous amino acid residues (mature forms had 188 [82%] homologous amino acid residues).

Identification of a bacteriophage containing a silent staphylococcal variant enterotoxin gene (sezA+).

A variant enterotoxin gene, referred to as sezA+, has been identified. Staphylococcus aureus FRI1106, a staphylococcal enterotoxin type D producer (Sed+), contained HindIII fragments of 3.8 and 9.4 kilobase pairs (kbp) that hybridized in Southern blot analysis to a probe containing only staphylococcal enterotoxin type A structural gene sequences. Presumably, probe A-624 hybridized to the 9.4-kbp HindIII fragment because of the sequence homology between sea+ and sed+. This 9.4-kbp HindIII fragment, which was part of a staphylococcal plasmid, was isolated and ligated into an Escherichia coli plasmid vector; Sed+ E. coli recombinant clones were isolated. The 3.8-kbp HindIII fragment was shown to be part of a viable lysogenic bacteriophage, and it contained sezA+. This sezA(+)-containing fragment was cloned into E. coli, and its DNA sequence was determined. Examination of the nucleotide sequence revealed a 771-bp region that contained an open reading frame with 85 and 77% nucleotide and derived amino acid sequence identifies with sea+ and staphylococcal enterotoxin type A, respectively. This open reading frame has 83 to 50% nucleotide sequence identities with the other types of staphylococcal enterotoxin genes. sezA+ was shown to be transcribed into stable mRNA. However, the sezA+ mRNA was not translated into an enterotoxinlike protein because it lacks an appropriate translation initiation codon.

Identification and characterization of human metabolites of CAI [5-amino-1-1(4'-chlorobenzoyl-3,5-dichlorobenzyl)-1,2,3-triazole- 4-carboxamide).

The calcium influx inhibitor and cytostatic agent, 5-amino-1-1(4'-chlorobenzoyl-3,5-dichlorobenzyl)-1,2,3-triazole-4-carboxamide (CAI), is in phase I clinical trial for patients with refractory cancer. Additional chromatography peaks were observed during HPLC analysis of patient samples. Identification and characterization of physiological metabolites were undertaken using HPLC techniques developed for their purification from blood, pleural fluid, and urine samples. A hydrophobic metabolite, M1, was purified and functionally characterized. Structural analysis of the purified compound indicated that it is a 3,5-dichloro-4(p-chlorobenzoyl)-benzoic acid. Quantitative analysis of M1 concentration during CAI administration indicated that the rise in M1 concentration lagged behind that of CAI and persisted after CAI was no longer detectable. No clear relationship between CAI or M1 and either toxicity or efficacy was observed. Chromatography of patient blood and urine samples under conditions favoring hydrophilic metabolite detection suggested the presence of a glucuronide compound; this was also indicated by sample treatment with beta-glucuronidase. Attempts at purification did not yield a compound stable for structural analysis. The benzophenone metabolite, M1, was nonfunctional in assays of calcium influx inhibition or proliferation. No pharmacodynamic associations were observed for these metabolites, nor was there pharmacological activity of the M1 as an individual agent. These data suggest that CAI is processed into triazole and benzophenone moieties by phase I metabolism, and these metabolites or the parent compound may be conjugated for excretion by glucuronidation.