Isolation and characterization of collagen and antioxidant collagen peptides from scales of croceine croaker (Pseudosciaena crocea)

Abstract

Acid soluble collagen (ASC) from scales of croceine croaker (ASC-C) was successfully isolated with the yield of 0.37% ± 0.08% (dry weight basis), and characterized as type I collagen on the basis of amino acid analysis and electrophoretic pattern. The antioxidant hydrolysate of ASC-C (ACH) was prepared through a two-stage in vitro digestion (4-h trypsin followed by 4-h pepsin), and three antioxidant peptides (ACH-P1, ACH-P2, and ACH-P3) were further isolated from ACH using ultrafiltration, gel chromatography, and RP-HPLC, and their amino acid sequences were identified as GFRGTIGLVG (ACH-P1), GPAGPAG (ACH-P2), and GFPSG (ACH-P3). ACH-P1, ACH-P2, and ACH-P3 showed good scavenging activities on hydroxyl radical (IC50 0.293, 0.240, and 0.107 mg/mL, respectively), DPPH radical (IC₅₀ 1.271, 0.675, and 0.283 mg/mL, respectively), superoxide radical (IC₅₀ 0.463, 0.099, and 0.151 mg/mL, respectively), and ABTS radical (IC₅₀ 0.421, 0.309, and 0.210 mg/mL, respectively). ACH-P3 was also effectively against lipid peroxidation in the model system. The antioxidant activities of three collagen peptides were due to the presence of hydrophobic amino acid residues within the peptide sequences. The collagen peptides might be used as antioxidant for the therapy of diseases associated with oxidative stress, or reducing oxidative changes during storage.

Figure 1

SDS-PAGE pattern ASC. Lane 1, protein markers; lane 2, ASC; lane 3, CSC.

Figure 2

Hydroxyl radical scavenging activity of ACH and its three fractions by ultrafiltration. All the results were triplicates of mean ± SD; SD: Standard deviation. (a–c) Values with same letters indicated no significant difference of same sample at different concentrations (p > 0.05). (A–D) Values with same letters indicated no significant difference of different sample at same concentrations (p > 0.05).

Figure 3

Gel filtration chromatography of ACH-III on a Sephadex G-15 column (A) and hydroxyl radical scavenging activity of its four subfractions (B). All the results were triplicates of mean ± SD; SD: Standard deviation. (a–c) Values with same letters indicated no significant difference of same sample at different concentrations (p > 0.05). (A–E) Values with same letters indicated no significant difference of different sample at same concentrations (p > 0.05).

Figure 4

Chromatography of ACH-III-3 separated by RP-HPLC. Elution was performed with the 30% acetonitrile containing 0.1% TFA and monitored at 280 nm.

Figure 5

Hydroxyl radical (A), DPPH radical (B), superoxide radical (C), and ABTS radical (D) scavenging activities of ACH-P1, ACH-P2, and AC-P3. All the values were mean ± SD; SD: Standard deviation. (a–e) Values with same letters indicated no significant difference of same sample at different concentrations (p > 0.05). (A–D) Values with same letters indicated no significant difference of different sample at same concentrations (p > 0.05).

Figure 6

Lipid peroxidation inhibition of ACH-P1, ACH-P2, and ACH-P3. All the values were mean ± SD; SD: Standard deviation. (a–e) Values with same letters indicated no significant difference of the same sample at different concentrations (p > 0.05); (A–E) Values with same letters indicated no significant difference of different sample at same concentrations (p > 0.05).