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Clinical Trial
, 10 (10)

A Pilot Study for the Detection of Cyclic Prolyl-Hydroxyproline (Pro-Hyp) in Human Blood After Ingestion of Collagen Hydrolysate

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Clinical Trial

A Pilot Study for the Detection of Cyclic Prolyl-Hydroxyproline (Pro-Hyp) in Human Blood After Ingestion of Collagen Hydrolysate

Yasutaka Shigemura et al. Nutrients.

Abstract

Levels of short linear hydroxyproline (Hyp)-containing peptides, such as prolyl-hydroxyproline (Pro-Hyp), increase in human blood after the ingestion of collagen hydrolysate, which has been associated with beneficial effects for human skin and joints. The present study demonstrates the presence of a novel food-derived collagen peptide, cyclic Pro-Hyp, in human blood after the ingestion of collagen hydrolysate. The cyclic Pro-Hyp levels in plasma samples were estimated by liquid chromatography mass spectrometry (LC-MS). Cyclic Pro-Hyp levels significantly increased in the plasma after ingestion of collagen hydrolysate, reaching a maximum level after 2 h and then decreasing. The maximum level of cyclic Pro-Hyp in plasma ranged from 0.1413 to 0.3443 nmol/mL, representing approximately 5% of linear Pro-Hyp in plasma after ingestion of collagen hydrolysate. Addition of cyclic Pro-Hyp in medium at 7 nmol/mL significantly enhanced the growth rate of mouse skin fibroblasts on collagen gel more extensively compared to linear Pro-Hyp.

Keywords: Pro-Hyp; collagen hydrolysate; collagen peptides; cyclic Pro-Hy; fibroblasts; mouse skin.

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
High-performance liquid chromatography (HPLC) and electrospray ionization mass spectrometry (ESI-MS) of linear and cyclic prolyl-hydroxyproline (Pro-Hyp). (A) The HPLC chromatogram peaks of (1) the linear Pro-Hyp and (2) after heating at 50 °C in 0.02 M ammonia solution. (B) Electrospray ionization mass spectra of peaks 1 and (C) 2 recovered from HPLC.
Figure 2
Figure 2
Design of the human study. Blood samples were collected before and after the ingestion of collagen hydrolysate.
Figure 3
Figure 3
Diagram of the separation procedure for cyclic and linear peptide fractions from human plasma before and after ingestion of collagen hydrolysate.
Figure 4
Figure 4
Amino acid analysis of the cyclic peptide fraction of mixed plasma samples from five volunteers. Cyclic peptide fractions prepared before (A) and at 1 (B), 2 (C), and 4 h (D) after ingestion of collagen hydrolysate were resolved by HPLC. Hydroxyproline peaks (Hyp) were observed in chromatograms of samples.
Figure 5
Figure 5
Changes in the concentration of cyclic Pro-Hyp in human plasma of five volunteers.
Figure 6
Figure 6
Changes in the average concentrations of linear (A) and cyclic (B) Pro-Hyp in human plasma of five volunteers. Data are shown as means ± SDs (n = 5). Different letters adjacent to data points indicate significant differences (P < 0.05).
Figure 7
Figure 7
Growth rates of mouse skin fibroblasts on collagen gel. Growth rates were estimated after 0, 1, 2, and 4 days of incubation in the presence of linear () and cyclic () Pro-Hyp or in the absence () of both peptides. ** and * indicate significant differences (P < 0.01 and P < 0.05, respectively).

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References

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