DOI: 10.23637/rothamsted.98883
title: Micro- and macro-elements in sheep excreta after consumption of feed supplements according to supplement formulation and level

description: In this experiment, sheep were given mineral supplements in their feed containing selenium (Se), zinc (Zn), manganese (Mn) and copper (Cu), with four treatments: two forms of the supplement (micronutrients in organic or inorganic compounds in the feed), and two levels of supplement dosing (higher or lower). The mass of excreta (faeces and urine) and the concentration of micro- and macro-nutrients in the excreta were measured over the course of the feeding trial. The trial was conducted over four weeks; the first two weeks were a baseline period and were followed by two weeks of feed that included the supplements.
Data presented include: sheep mass and body condition throughout the experiment; the mass of silage and concentrate feed consumed by sheep and its macro- and micro-nutrient contents; the volume of urine and the mass of faeces excreted and their macro- and micro-nutrient contents; and the results of a preliminary trial to assess whether drying of the sheep faeces affects the measurement of macro- and micro-nutrients by ICP analysis. In addition to the micronutrients in the supplements, data are presented for P, S, Fe, Mo, and Cd, plus Al, As, Ca, Co, Cr, K, Mg, Na, Ni, Pb, and Ti in the feed only. 
Animals were assessed daily for health and well-being, as determined by alertness, feed and water intake. All animal procedures and the care for the animals were carried out under strict regulations described in the Animals (Scientific Procedures) Act 1986 issued by the Home Office of Her Majesty’s Britannic Government under the Project License number P592D2677.
description: Twenty-four castrated male Charolais x Suffolk-Mule sheep were weighed, and body condition scored (see Feed_excreta_and_sheep_mass.csv), and the data used to allocate animals to one of four treatment groups to ensure comparable starting points. Sheep were housed in a specially designed facility with individual pens with ad-lib access to drinking water, and the volume consumed was not measured. Each pen had feed bins that measured the mass of feed given, or remaining unconsumed, for silage and concentrate feed separately. Sheep wore bespoke ‘nappies’ to allow the collection of faeces, while urine ran through the slatted floor and was collected in a tray underneath the pen. 
The sheep had been grazing pasture prior to the experiment. For the first seven days after entering the pens, sheep were offered silage as feed. For the following seven days, concentrate feed (sometimes known as pelleted feed or sheep nuts and containing grain and legume crops) was also offered to the sheep, with an additional ca 100 g day-1 offered to achieve a 40:60 dry matter (DM) ratio of concentrate to silage. This concentrate was a control, and had no supplemental micronutrients added to it. This 14-day period was defined as the baseline period, and day 0 of the experimental period immediately followed it.
During the experimental period (day 0 to day 14), the control concentrate feed was replaced with the concentrate feed that contained supplemental micronutrients. The organic treatment contained selenised yeast (Selplex®, Alltech Inc., USA) and Cu, Zn, Mn chelates of protein hydrolysate (Bioplex®, Alltech Inc., USA). The inorganic treatment contained selenite, zinc oxide, copper sulphate pentahydrate and manganese oxide. The higher rate of supplementation (final Se, Cu, Zn and Mn concentrations of 0.6, 17, 104 and 60 mg kg-1 in concentrate feed, respectively) was set at the level typically used by European industries based on the regulations set by the National Research Council of the USA. The lower doses were set at 0.2, 13, 84 and 48 mg kg-1 concentrate feed for Se, Cu, Zn and Mn, respectively. Treatments are referred to as organic-high (OH), organic-low (OL), inorganic-high (IH) and inorganic-low (IL). 
The mass of silage and concentrate feed consumed by each sheep was measured during both the baseline and experimental period (feed_excreta_and_sheep_mass.csv), and micro- and macro-element concentrations of the silage and control concentrate (dry matter basis) are given in Non_supplement_intake.csv. The mass of faeces and volume of urine excreted by each sheep was measured daily during the experimental period (feed_excreta_and_sheep_mass.csv). Excreta from a 24-hour period were bulked, per animal and per excreta type, and were recorded at ~8am on each experimental day. Therefore, excreta recorded on day 1 were collected from ~8am on day 0 (when feeding with concentrate containing the supplements commenced) until ~8am on day 1. On day 0, excreta samples were collected to represent the baseline period, but the total mass/volume of excreta was not recorded. On days 0, 1, 3, 7, 10 and 14, the micro- and macro-element concentrations of faeces and urine were measured (Faeces_concentrations.csv and Urine_concentrations.csv respectively). 
The moisture content of faeces was determined by drying faeces at 105 oC. Silage samples were freeze-dried and faecal samples were oven dried at 80 oC. Dry silage, faecal and concentrate feed were finely ground and a known mass (~0.25 g) was pre-digested in 3 mL nitric acid followed by a digestion in 3 mL ultrapure water and 2 mL hydrogen peroxide at 175 oC for 10 minutes in a microwave digestion system (MARS, CEM corporation, USA), before being diluted to 50 mL in ultrapure water. Drinking water samples, taken daily from individual water troughs and bulked, were acidified in 5 % (v/v) nitric acid. Urine samples were filtered through a Whatman No. 42 filter paper and diluted 20 times by nitric acid and methanol, final concentrations 0.5% and 1% respectively. Analysis of the prepared samples for total concentrations of macro- and micro-elements was by inductively coupled plasma – optical emission spectrometry (ICP-OES) or inductively coupled plasma – mass spectrometry (ICP-MS). 
In addition, a preliminary test to determine whether the method of drying faecal samples affects the concentration of elements measured by ICP, the main concern being the volatilisation of selenium, is presented in drying_method.csv. Sheep faeces from animals consuming silage and control concentrate feed from the main trial were collected and mixed to homogenise. The bulk sample was split and the faeces either air-dried (room temperature ~20 oC), oven-dried (80 oC) or freeze-dried, with 5 replicates of each method, until sample weight was constant. Samples were ground prior to analysis and analysed using the same digestion and analysis method as for the main trial. The results showed no significant difference between the drying methods in the concentration of elements measured and therefore oven-drying of samples was used for the main trial.