Recovery of intravenously infused chromium EDTA and lithium sulphate in the urine of cattle and their use as markers to measure urine volume

A series of metabolism experiments investigated the recovery of continuous-, intravenously infused chromium complexed with ethylenediamine tetra-acetic acid (CrEDTA) and lithium sulphate in the urine of cattle with a view to using the markers to estimate urine and metabolite output in grazing cattle. The recovery of Cr in urine from these infusions was similar (90%) in metabolism trials when cattle consumed three very contrasting diets: high-grain formulated pellet, lucerne hay (Medicago sativa) or low-quality native grass hay (predominantly Heteropogon contortus). By contrast, Li recovery in urine averaged 46.3 +/- 0.40% and 72.6 +/- 0.43% for native pasture and lucerne hays, respectively, but was not constant across days. There was negligible transfer of Cr from CrEDTA in blood serum to the rumen or faeces, whereas appreciable quantities of infused Li were found in both. The ratio of urine volume estimated by spot samples and marker dilution of Cr, to urine volume measured gravimetrically, was 1.05. In grazing studies using rumen-fistulated (RF) steers grazing seven different tropical and temperate grass and legume pastures, the ratio of concentrations of purine derivatives (PD) to Cr in spot samples of urine was shown to vary diurnally in the range of 49% to 157% of the average 24 h value. This finding indicated the need for regular sampling of urine to achieve an accurate average value for the PD: Cr ratio in urine for use in estimating urinary PD excretion and hence microbial protein production in the rumen. It was concluded that continuous, intravenous infusion of CrEDTA resulted in a constant recovery of Cr in the urine of cattle across diets and, provided an intensive sampling regime was followed to account for diurnal variation, it would be suitable as a marker to estimate urine volume and urinary output of PD in grazing cattle.

Disruption of iron homeostasis increases phosphine toxicity in caenorhabditis elegans

The aim of this study is to identify the biochemical mechanism of phosphine toxicity and resistance, using Caenorhabditis elegans as a model organism. To date, the precise mode of phosphine action is unclear. In this report, we demonstrate the following dose-dependent actions of phosphine, in vitro: (1) reduction of ferric iron (Fe3+) to ferrous iron (Fe2+), (2) release of iron from horse ferritin, (3) and the peroxidation of lipid as a result of iron release from ferritin. Using in situ hybridization, we show that the ferritin genes of C. elegans, both ferritin-1 and ferritin-2, are expressed along the digestive tract with greatest expression at the proximal and distal ends. Basal expression of the ferritin-2 gene, as determined by quantitative PCR, is approximately 80 times that of ferritin-1. However, transcript levels of ferritin-1 are induced at least 20-fold in response to phosphine, whereas there is no change in the level of ferritin-2. This resembles the reported pattern of ferritin gene regulation by iron, suggesting that phosphine toxicity may be related to an increase in the level of free iron. Indeed, iron overload increases phosphine toxicity in C. elegans at least threefold. Moreover, we demonstrate that suppression of ferritin-2 gene expression by RNAi, significantly increases sensitivity to phosphine. This study identifies similarities between phosphine toxicity and iron overload and demonstrates that phosphine can trigger iron release from storage proteins, increasing lipid peroxidation, leading to cell injury and/or cell death.

Purification of immunoglobulins from chicken sera by thiophilic gel chromatography

Immunoglobulin Y is different from most of the other immunoglobulins because it does not bind protein A or protein G. Thiophilic gel chromatography has been successfully used to purify IgY from chicken egg yolk, but the technology has not previously been used to purify IgY from serum. In this research note, we describe the optimization of T-gel chromatography for purification of IgY from serum. Data are provided on the recovery and purity of IgY obtained using potassium sulfate buffers of different concentrations. Decreasing the strength of potassium sulfate buffer from 0.5 to 0.3 M did not alter the amount of IgY recovered but increased the purity. Using 0.3 M potassium sulphate, we recovered approximately 63.7% of the serum Ig as almost pure IgY.

Key Role of Fe2+ in Epithiospecifier Protein Activity.

The chemical nature of the hydrolysis products from the glucosinolate-myrosinase system depends on the presence or absence of supplementary proteins such as epithiospecifier proteins (ESPs). ESPs promote the formation of epithionitriles from terminal alkenyl glucosinolates and, as recent evidence suggests, simple nitriles at the expense of isothiocyanates. From a human health perspective isothiocyanates are the most important because they are major inducers of carcinogen-detoxifying enzymes. Fe2+ is an essential factor in ESP activity, although several recent studies have highlighted discrepancies in the understanding of the ESP-iron interaction. To investigate further the role iron species play in regulating ESP activity, four ESP-containing seedpowders were analyzed for ESP and myrosinase activities, endogenous iron content, and glucosinolate degradation products after the addition of iron species, specific chelators, and reducing agents. For the first time this paper shows the effect of these additions on the hydrolysis of individual glucosinolates that constitute the total pool. Aged seeds and 3-day seedlings were also tested to investigate the effects of seed storage and early plant development on iron levels and ESP activity. The four ESP-containing plant systems tested gave two distinctive responses, thus providing strong evidence that ESPs vary markedly in their Fe2+ requirement for activity. The results also indicated that reduction of ferric to ferrous iron drives variations in ESP activity during early plant development. The reverse oxidation reaction provided a convincing explanation for the loss of ESP activity during seed storage. Aged seeds produced seedlings with substantially lower ESP activity, and there was a concomitant loss in germination rate. It was concluded that manipulation of endogenous iron levels of ESP-containing plants could increase the conversion of glucosinolates to isothiocyanates and enhance potential health benefits.