Process optimisation of haemoglobin hydrolysis by complex proteases to produce haem‐enriched peptides and its iron uptake property evaluation by Caco‐2 cell model

Abstract

In this study, haemoglobin was hydrolysed by compound protease to produce haem-enriched peptides in order to improve the bio-absorptivity of haem iron. The hydrolysis process was optimised through an orthogonal experiment. The molecular weight, solubility and absorptivity of haem-enriched peptides were also investigated. The optimal hydrolysis process was papain/alkaline protease ratio 6:4, enzyme content 9000 U g −1 , hydrolysis at 50°C and pH 8 conditions for 4 h. After hydrolysis at this condition, the haem released amount and haem/peptide ratio were 89.6% and 28.2%, respectively. The molecular weight of the haem-enriched peptides was mainly distributed in 3-14.4 kD, and the solubility increased significantly with pH increasing. Caco-2 cell absorption experiment demonstrated that the iron absorption of haem-enriched peptides was significantly higher (50.25 ng ferritin per mg protein) than haemin chloride, haemoglobin and collagen plus haemin chloride complex. This study provides a very promising way for the development of iron supplement products.

Production and Characterization of Heme Iron Polypeptide from the Blood of Skipjack Tuna (Katsuwonus pelamis) Using Enzymatic Hydrolysis for Food Supplement Application

“…On the other hand, studies have showed that the resulting heme iron molecules from digested hemoglobin under stomach condition were still poorly soluble at low pH values of the stomach. This subsequently could reduce their absorption in the intestine [8,30,31]. However, our study could yield tuna-HIP with high solubility of around 60% at pH 2 that might possibly promote better iron absorption under physiological conditions.…”

“…This finding might be useful for the commercial production of high-solubility HIP for iron supplementation in the future due to the short hydrolysis time when compared to previous studies (Table 4). Wide range pH (N/A specific pH) [33] Alcalase + Flavourzyme, pH 7.5 8-12% 1-7.5 kDa pH 2 [7] Neutrase, acid enzyme, alkaline enzyme, and Papain 6-12% 3-14.4 kDa pH > 8 [8] Yellowtail fish Alcalase, pH 10, 24 h N/A 2-3.6 kDa pH < 4, pH > 6 [5] N/A: not available (data were not mentioned by the authors).…”

“…Previous studies have illustrated that HIP exhibit good solubility at pH 2 but that the MW of the associated peptides varies depending on the sources and preparation. These include 2-3.6 kDa in fish HIP [5], 4-5 kDa in bovine HIP [6], 1-7.5 kDa in porcine HIP [7], and 3-14.4 kDa in porcine HIP [8]. Compared to pure heme, HIP could lead to 10 times higher heme iron absorption [5,6].…”