Improved swimming pool achieves higher reproducibility and sensitivity to effect of food components as ergogenic AIDS.

A previously developed current swimming pool for mice has been used to evaluate many food components that enhance endurance exercise performance. In this article, to improve reproducibility, reliability and sensitivity of this assay system, we improved the spout part to generate a uniform current and divided the pool into six lanes to avoid physical interference between swimming mice. The stability of the current flow was assessed by measuring the surface current speed and water volume from the spout part. Maximum swimming times of ddY and BALB/c mice were measured to assess the reproducibility of the maximum swimming time. The improvement in sensitivity compared to the original equipment was estimated under three physiological conditions: low carbohydrate diet feeding, low blood hemoglobin level, and carbohydrate supplementation during exercise. The new spout part improved uniformity and quick adjustment of surface current, yielding an increase of workload in a stepwise manner during swimming. Exercise workload was increased in proportion to surface current speed, as evidenced by cadence of kicks and serum lactic acid levels. The improved swimming pool showed higher reproducibility of swimming time until fatigue (p<0.0001). Correspondence between blood hemoglobin concentration and swimming time was improved in the swimming pool. The improved swimming pool yielded higher sensitivity for low carbohydrate diet feeding (p<0.0001) and carbohydrate supplementation during exercise (p<0.01) compared to the original swimming pool. The improvement of the swimming pool achieved higher sensitivity and reproducibility in assessing various diet and food components compared to the original swimming pool.

Elevated urinary Cr loss induces a reduction in renal Cr concentration and the negative Cr balance in streptozotocin-induced diabetic mice.

Chromium (Cr) is an essential trace element and is important for normal carbohydrate metabolism, and its deficiency in animals can cause a diabetic-like state. Human and experimental animal studies suggest that urinary Cr excretion is increased in diabetic populations. To investigate whether hyperglycemia-induced elevation of urinary Cr excretion reduces tissue Cr storage conditions, we assessed total Cr balance and Cr distribution in streptozotocin (STZ)-induced diabetic mice. Male C57BL mice were randomly assigned to STZ or control groups and their urine was collected 7, 14, 21 and 28 d after STZ injection. An inductively coupled plasma mass spectrometry instrument equipped with a dynamic reaction cell was used for determination of Cr in urine, plasma and tissues samples. The urinary excretions of Cr were 15.4+/-3.0 and 356+/-62 ng/d, and the renal Cr concentrations were 0.85+/-0.12 and 0.17+/-0.03 ng/mg for the control and diabetes groups, respectively (p<0.01), after 28 d. The Cr balance in STZ-treated mice was distinctly negative due to the increase in urinary Cr loss (p<0.01). These results suggest that in mice, STZ induces a reduction in renal Cr concentration and total negative Cr balance caused by an increase in urinary Cr output.

Zinc bioavailability is improved by the micronised dispersion of zinc oxide with the addition of L-histidine in zinc-deficient rats.

Zinc fortification of milk or soft drinks is usually used to combat zinc deficiencies in developing countries. Water-soluble zinc compounds, such as zinc sulfate or zinc citrate, are better absorbed but have an unacceptable taste. A micronised, dispersible zinc oxide (MDZnO), which does not have such a problem concerning taste, had higher solubility compared to ZnO (zinc oxide) in an artificial gastric solution. MDZnO was tested for its bioavailability using zinc-deficient Wistar rats. Prior to the experiment, rats were fed zinc-deficient diet for 3 wk and were orally administered control (distilled water) or zinc solutions (ZnO, ZnO+L-histidine (His), MDZnO, MDZnO+His, 1 mg zinc/kg or 3.2 mg His/kg body weight). Compared to ZnO, MDZnO showed a lag in peak time and a lengthy period of continued high plasma zinc concentration after the single oral administration of zinc compounds. Addition of His to MDZnO elevated serum zinc concentration. Serum zinc concentration (area under the curve) in rats administered MDZnO with His was significantly higher than in rats administered distilled water (p<0.05). Liver zinc level was significantly higher in rats administered MDZnO with His compared with control rats (p<0.05), although the level was not affected by the administration with ZnO alone, ZnO+His, or MDZnO alone. In conclusion, the solubility of ZnO was elevated by the micronised dispersion tecnique and an in vivo study using zinc-deficient rats confirmed that its bioavailability was significantly improved compared to ZnO and the coadministration of His additively enhanced the bioavailability of MDZnO.

Supplementation with chromium picolinate recovers renal Cr concentration and improves carbohydrate metabolism and renal function in type 2 diabetic mice.

To study the preventive effect of supplemented chromium picolinate (CrPic) on the development of diabetic nephropathy in mice, we analyzed the effects of CrPic supplementation on renal function and concentrations of serum glucose and tissue chromium (Cr). In experiment 1, male KK-Ay obese diabetic mice were fed either a control diet (control) or a diet supplemented with 2 mg/kg diet (Cr2) or 10 mg/kg diet (Cr10) of Cr for 12 wk. Cr10 significantly ameliorated hyperglycemia after a glucose load, creatinine clearance rates, and urinary microalbumin levels (p<0.05). In experiment 2, the Cr10 diet was fed to male KK-Ay obese diabetic mice and C57BL nondiabetic mice for 4 wk. The CrPic diet reduced urinary albumin excretion in the diabetic mice (p<0.05). Inductively coupled plasma-mass spectrometry analysis revealed that the renal Cr content and the recovery of renal Cr concentration after Cr supplementation were significantly lower in the diabetic mice than in the nondiabetic mice (p<0.01). These observations suggest that Cr supplementation of type 2 diabetic mice reduces the symptoms of hyperglycemia and improves the renal function by recovering renal Cr concentration.

Role of the carbohydrate chain and two phosphate moieties in the heat-induced aggregation of hen ovalbumin.

We investigated the effect of the carbohydrate chain and two phosphate moieties on heat-induced aggregation of hen ovalbumin. The dephosphorylated form of ovalbumin was obtained by treating the original protein with acid phosphatase. The single carbohydrate chain was removed by digestion of heat-denatured ovalbumin with glycopeptidase F, and the resulting polypeptide without this carbohydrate chain was correctly refolded to acquire protease-resistance. Thermal unfolding can be approximated by a mechanism involving a two-state transition between the folded and unfolded states with a midpoint temperature of 76 degrees C for the original form, of 74 degrees C for the dephosphorylated form, and of 71 degrees C for the carbohydrate-free form. The conformational stability of the original form was higher than that of the carbohydrate-free form. When the three forms of ovalbumin were heated to 80 degrees C and then cooled rapidly in an ice bath, the polypeptide chains were compactly collapsed to metastable intermediates with secondary structures whose properties were indistinguishable. Upon incubation at 60 degrees C, renaturation was possible for a large portion of the intermediates of the original form, but for only a small portion of those of the carbohydrate-free form. Light scattering experiments showed that in the presence of sulfate anions, the intermediates of the carbohydrate-free form aggregated to a greater extent than did those of the original form. The intermediates of the carbohydrate-free form bound to the chaperonin GroEL with about 10-fold higher affinity than those of the original form. It follows that the carbohydrate chain and the two phosphate moieties do not affect hydrophobic collapse in the kinetic refolding of hen ovalbumin but play an important role in the slow rearrangement. They block the off-pathway reaction that competes with correct refolding by effectively decreasing surface hydrophobicity.

Purification and characterization of aromatic amine dehydrogenase from Alcaligenes xylosoxidans.

Aromatic amine dehydrogenase was purified and characterized from Alcaligenes xylosoxidans IFO13495 grown on beta-phenylethylamine. The molecular mass of the enzyme was 95.5 kDa. The enzyme consisted of heterotetrameric subunits (alpha2beta2) with two different molecular masses of 42.3 kDa and 15.2 kDa. The N-terminal amino acid sequences of the alpha-subunit (42.3-kDa subunit) and the beta-subunit (15.2-kDa subunit) were DLPIEELXGGTRLPP and APAAGNKXPQMDDTA respectively. The enzyme had a quinone cofactor in the beta-subunit and showed a typical absorption spectrum of tryptophan tryptophylquinone-containing quinoprotein showing maxima at 435 nm in the oxidized form and 330 nm in the reduced form. The pH optima of the enzyme activity for histamine, tyramine, and beta-phenylethylamine were the same at 8.0. The enzyme retained full activity after incubation at 70 degrees C for 40 min. It readily oxidized various aromatic amines as well as some aliphatic amines. The Michaelis constants for phenazine methosulfate, beta-phenylethylamine, tyramine, and histamine were 48.1, 1.8, 6.9, and 171 microM respectively. The enzyme activity was strongly inhibited by carbonyl reagents. The enzyme could be stored without appreciable loss of enzyme activity at 4 degrees C for one month at least in phosphate buffer (pH 7.0).

A soybean peptide isolate diet promotes postprandial carbohydrate oxidation and energy expenditure in type II diabetic mice.

The aim of the present study was to determine the effects of dietary proteins on the oxidation of dietary carbohydrate and lipids in type II diabetic mice. KK-A(y) strain mice were provided free access to a high fat diet (30% of energy as fat) for an initial 4-wk period to induce diabetes. To reduce body weight gain, the mice were subsequently fed restrictive isoenergetic and isonitrogenous diets (35% of energy as protein and 5% as fat) based on either casein or soy protein isolate hydrolysate (SPI-H) for 4 wk. To measure exogenous carbohydrate and lipid oxidation, the mice were fed a diet containing (13)C-glucose or (13)C-triolein while they were in a respiratory chamber for 72 h. Postprandial energy expenditure was higher in the SPI-H than in the casein group; this difference was due to an increase in postprandial exogenous and endogenous carbohydrate oxidation. There were no differences in 24-h energy expenditure between dietary groups. Oxidation of exogenous carbohydrate tended to be higher (P = 0.054) in the SPI-H group during the 24 h of measurement. Fecal excretion of (13)C-glucose was lower but the excretion of lipid was higher in mice fed the SPI-H diet than in casein-fed mice. These results indicate that in type II diabetic mice, dietary SPI-H not only inhibits the absorption of dietary lipids and increases the absorption of dietary carbohydrates but also augments postprandial energy expenditure, which is accompanied by a postprandial increase in oxidation of dietary carbohydrates.

Amino acid composition of soybean protein increased postprandial carbohydrate oxidation in diabetic mice.

The effects of an amino acid mixture simulating dietary soybean protein on the postprandial energy metabolism was investigated using type II diabetic mice. KK-A(y) strain mice were fed restrictive isoenergetic and isonitrogenous diets (35% of energy as protein and 5% as fat) based on either casein, soybean protein isolate hydrolysate (SPI-H), SPI-HET (ethanol unsoluble fraction of SPI-H), SPI-AA and casein-AA (amino acid mixtures simulating SPI-H and casein). To measure dietary carbohydrate oxidation, the animals were fed a diet containing (13)C-glucose. Postprandial respiratory quotient and expired (13)CO(2) were higher in the SPI-AA than in the casein-AA group, as the differences were similarly observed in mice fed SPI-H and casein diet. No significant differences were observed in the postprandial respiratory quotient and expired (13)CO(2) between the SPI-H and SPI-HET group. In conclusion, this study on food-restricted mice indicates that the amino acid mixtures simulating SPI-H or casein could affect postprandial energy metabolism in diabetic mice, as observed in those fed SPI-H or casein in the form of peptide or protein.

Use of 13C-labeled glucose for measuring exogenous glucose oxidation in mice.

The author modified a respiratory gas analyzer to analyze the respiratory 13CO2 of 12 small laboratory animals all at once. To investigate the practical use of this system, mice were orally (OR) or intravenously (i.v.) given glucose solutions containing three different amounts of 13C-labeled glucose. Expired 13CO2 derived from exogenous glucose was detected within 10 minutes after administration in OR mice, but about 30 minutes in i.v. mice. The height of the peak of 13CO2 expiration was correlated with the administered 13C-glucose mass.