2 ± 1.4% (mean ± SD in triplicates) of wet weight. The concentrations of protein, hydroxyproline, sialic acid and uronic acid, expressed as milligrams per gram of dry tissue, were 724.8 ± 9.3, 35.5 ± 1.2, 6.7 ± 0.2 and 41.2 ± 0.9, respectively. Fig. 1 illustrates that the content of uronic acid liberated from antler cartilaginous tissues with papain under the fixed conditions of Veliparib pH 6.0, 50 °C and 4 h incubation time was dependent on increased hydrostatic pressure. Increased pressure, by increasing the solubility of CS, was one of the most important variables in the HHP-EH process.

The content of released uronic acid was highest at 75 MPa (94.4 ± 2.9% of total uronic acid recovered) and at 100 MPa (95.1 ± 2.5% of total uronic acid recovered). This value was 2 and 5 times higher (P < 0.05) than values obtained at 50 MPa (53.5 ± 3.0%) and 25 MPa (21.6 ± 1.1%), respectively. The extractability of uronic acid was less than 19 ± 1.1% at ambient pressure (0.1 MPa). As a result, higher pressure at 100 MPa led to a higher extraction yield. Fig. 2 illustrates that the content of uronic acid liberated from antler cartilaginous tissues with papain under the fixed conditions of pH 6.0, 50 °C and 100 MPa was dependent on the incubation time. The liquid mixtures of antler tissue and papain were hydrolysed in the high-pressure chamber machine for 1–4 and 8 h. The results show that the

yield of total uronic acid significantly increased Y-27632 chemical structure (P < 0.05) between 1 and 3 h incubation time and then increased slightly from 3 to 4 h. Papain demonstrated

significant increases in the uronic Phosphoprotein phosphatase acid yield during the initial 3 h incubation. However, the effect of the incubation time between 4 h and 8 h was not significantly different in papain treatment (P > 0.05). The result indicated that incubating for longer than 4 h was likely unnecessary because the yield did not significantly increase thereafter. The effect of different temperatures is illustrated in Fig. 3, when conditions are fixed at a constant pressure of 100 MPa for 4 h incubation time. The result showed that the HHP-EH demonstrated significant increases (P < 0.05) in total uronic acid yield from 20 to 30 °C, and then again significantly increased from 30 to 40 °C. However, the effect of the temperature between 40 and 50 °C was not significantly different in the HHP-EH treatment (P > 0.05). The results indicated that incubating at below 40 °C was not fully activating the papain to liberate CS from the samples. The CS uronic acid extracted from antler cartilaginous tissues hydrolysed with papain at 50 °C for 4 h in 100 MPa accounted for ∼94% of total uronic acid recovered (Fig. 1). The hydrolysed antler papain extracts were applied to the Sephacryl S-300 chromatography column to isolate antler CS fractions. The majority (94%) of antler CS fractions eluted at peaks of Kav, 0.15 in a single fraction ( Fig. 4).