Vario max cube

All feed samples used in the experiment were dried at 60 °C for 48 h and ground in a cyclone mill (Foss, Hillerød, Denmark) fitted with a 1 mm screen. Dry matter (930.15), acid detergent fiber (973.18), ash (942.05), and ether extract (EE; 2003.05) were analyzed using the procedure reported by Horwitz and Latimer [12 ]. Neutral detergent fiber (NDF) was analyzed using a heat-stable amylase and expressed inclusive of residual ash (aNDF) [13 (link)]. CP was calculated as 6.25 times the nitrogen content, and total nitrogen was measured via the Dumas combustion method using an elemental combustor (Vario Max Cube, Elementar Gmbh, Frankfurt, Germany). The acid detergent-insoluble CP and neutral detergent-insoluble CP (NDICP) levels in each sample were determined according to the method described by Licitra et al. [14 (link)]. Non-fiber carbohydrates (NFC) were calculated as 100-ash-EE-CP-(aNDF-NDICP) based on the guidelines provided by the National Research Council (NRC) [15 ]. The experimental feed is described in Table 1.

In 1 mm of dried, sieved soil samples, the following parameters were determined: pH—potentiometrically and electrical conductivity (EC)—conductometrically. The sum of alkaline cations (S) and hydrolytic acidity (Hh) were determined by using the Kappen method. The total carbon (C_Tot), total nitrogen (N_Tot) and total sulphur contents were determined with a CNS analyser Vario MAX Cube (Elementar, Langenselbold, Germany) [35 ]. Bioavailable trace elements were extracted from the soil for 2 h with a 0.01 mol dm⁻³ solution of CaCl₂ (soil:solution = 1:10) [36 (link)]. The total macroelements and trace elements were measured after ashing the sample at a temperature of 550 °C for 12 h and mineralization in nitric and perchloric acids (3:2 v/v). The studied elements were determined by inductively coupled plasma optical emission spectrometry (ICP-OES, Perkin Elmer Optima 7300 DV, Waltham, MA, USA) [29 ].

Canary seeds, oat and wheat flours were prepared by grinding frozen dehulled seeds into fine powders using an analytical mill (IKA A11, IKA, Staufen, Germany). The flours were stored at room temperature in the dark until used for analysis. Total nitrogen content of canary seed, oat and wheat flours were determined using a Vario MAX Cube (Elementar, Langenselbold, Germany), following the Dumas combustion method [20 ] with EDTA as a nitrogen standard. Crude protein content of cereal flours was calculated as total nitrogen multiplied by a conversion factor of 5.7 for canary seed and 6.25 for oat and wheat. Fat content was determined using a Soxtec apparatus (Foss Tecator Soxtec System HT-6, 1043 extraction unit, Brampton, OT, Canada) according to AACC method 30–25.01 [21 ]. Ash content was determined according to AACC official method 08–03.01 [22 ], and moisture was determined by drying 2 g sample in a Fisher Isotemp vacuum oven (Fisher Scientific, Montreal, Qc, Canada) for 5 h at 100 °C [23 ]. Total carbohydrate comprising starch and fiber content was estimated by percent difference. All determinations were done at least in triplicate and average values were computed.

Bovine β-lactoglobulin (β-Lg) (A and B) was purified from whey protein isolate (WPI 895, Fonterra Co-operative Group, Auckland, New Zealand) by selective thermal aggregation with subsequent microfiltration/ultrafiltration and spray drying, as described in detail by [28 (link)]. The obtained powder had a total protein content of >99% on a dry matter basis (Vario MAX cube, Elementar Analysensysteme GmbH, Langenselbold, Germany; RP-HPLC), whereby >99% was β-Lg. Native β-Lg represented >86% of the total protein as determined by RP-HPLC (Toro-Sierra et al. 2013). Lactose (HPLC) and salts (Na, K, Ca) (ELEX 6361, Eppendorf AG, Hamburg, Germany) concentrations were <0.05% and 0.7%, respectively.

In order to determine chemical composition, the samples taken from initial materials and composts were subjected to incineration in a chamber furnace (Czylok, Jastrzębie-Zdrój, Poland) at a temperature of 800 °C for 12 h. Following digestion of the ash in diluted (1:2) HNO₃, the samples were transferred to flasks. Next, by adding redistilled water, the volume of the final sample was adjusted to 50 mL. The content of the examined elements (in the obtained solutions) was determined via the ICP-OES procedure. The tests were performed in duplicate.
The pH of the compost suspended in water (at a ratio of 1:10) was determined potentiometrically (pH-meter CP-505, Elmentron, Zabrze, Poland). This was performed analogously in the case of electrical conductivity (EC) by implementing the CCO-501 conductometer (Elmentron, Zabrze, Poland). The content of total carbon and nitrogen were established using the Vario MAX Cube analyzer, which is equipped with an IR sensor (Vario MAX Cube, ElementarAnalysensysteme, GmbH, Langenselbold, Germany). In order to determine the content of ash, the samples taken from the baseline materials were also subjected to incineration in a chamber furnace at a temperature of 800 °C for 12 h. The tests were performed in duplicate.

Total N was measured by dry combustion using an NC analyzer (Vario MAX cube, Elementar, Hanau, Germany). Aspartic acid was used as a standard in total N analysis.
Chlorophylls a and b were extracted from finely ground powder (5 mg) of freeze-dried leaf samples using N,N′-dimethylformamide (5 mL). After incubation for 24 h at 4 °C under dark conditions to allow complete decolorization, the samples were centrifuged at 2000×g for 30 min, and the absorbance of the supernatant was measured at 663.8 and 646.8 nm using a spectrophotometer (UV-1900, Shimadzu, Kyoto, Japan). Chlorophyll a and b contents were calculated using the equation of Porra et al. (1989)^{72 (link)}.