| Calcium is an essential nutrient element and has important physiological andbiochemical functions, such as bone formation, muscle contraction, cell phagocytosisand enzyme activation. Calcium deficiency has become one of global nutritionproblems due to the poor absorption. So it has brought to the attention to develop newtypes of calcium supplements. Peptides have been cited as mineral carriers, thuscarrying calcium ions into mucous cells and promoting calcium absorption. We chosecalcium-binding peptide from tilapia fish scales and prepared peptide-calciumcomplex,then studied the stability and biological availability through building Caco-2model and SD rats model. The detailed works were as follows:1. Preparation of peptide-calcium complexThe calcium-binding peptide was prepared by pepsin (E/S2%4.5h), trypsin(E/S2.5%6h), flavourzyme (E/S1%5h) in optimum temperature and pH conditions.Peptide-calcium complex was prepared according to the ratio5:1(W/W) for peptide toCaCl2on conditions: temperature50℃, pH6.0,30min, the calcium amounts inpeptide-calcium complex was93.6mg/g.2. Study on the stability of peptide-calcium complexThe in vitro digestion enzymes, small intestinal brush border membrane enzymes,different pH value, heat treatment and possibly coexisting food ingredients such asNaCl, lactose were examined to estimate the stability of the peptide-calcium complex.Results showed that peptide-calcium complex could loss calcium under acid conditionand easily combine calcium when the pH was7or9; In vitro digestion experimentshowed peptide-calcium complex could mantain68.16%of calcium after treated bydigestion enzymes for2h; Heat treatment experiments showed that peptide-calciumcomplex was unstable above50℃;The coexisting food ingredients: NaCl, lactose, did not affect the stability of peptide-calcium complex.3. Study on the physical and chemical properties of peptide-calcium complexResults showed that the component with molecular weight of500-1000Dacombined the most calcium (196.3mg/g). Glu and Asp residues might be the siteswhich calcium combined with; UV scanning, Fourier transform infrared spectroscopy,ultrastructure and X-ray diffraction all showed that the COO-, N-H might be involvedin the combination. Compared with the original protein peptide, the microstructureand diffraction spectra of peptide-calcium complex changed obviously, confirmingthat the new substance generated.4. Effects of food components on peptide-complex absorption in Caco-2cellThe effects of phosphorus, vitamin D and phytic acid on the transport and uptakefor CaCl2and peptide-calcium complex were studied using Caco-2cells. Resultsshowed that the retention rate, transfer rate and absorption rate of peptide-calciumcomplex were significantly higher than that of CaCl2(P <0.05). The retention rateand absorption rate for CaCl2and peptide-calcium complex were significantlyincreased when increasing the phosphorus content; Adding vitamin D couldsignificantly improve the absorption of CaCl2, but had no effect on the absorption ofpeptide-calcium complex; Adding phytic acid could inhibit the absorption of bothCaCl2and peptide-calcium complex, but the inhibitory effect on peptide-calciumcomplex was significantly lower than that of CaCl2(P <0.05).5. Effects of food components on peptide-complex absorption in SD ratsThe peptide-calcium complex absorption in SD rats was studied as well as theeffects of dietary phosphorus, vitamin D and phytic acid content. Results showed thatcalcium apparent absorption rate, blood calcium, blood phosphorus levels inphosphorus deficiency group were significantly lower than control group (P<0.05);The vitamin D deficiency group and phytic acid group did not significantly affectcalcium apparent absorption rate; The peptide-calcium complex absorption wasaffected by dietary phosphorus level, but was not regulated by the calcium channelsand vitamin D, Peptide-calcium complex could delay the generation of phyticacid-calcium precipitation, thus promoting calcium absorption. Briefly, fish scale peptide-calcium complex was sensitive to pH value and heattreatment, but had a certain tolerance to the gastrointestinal tract digestive enzymesand intestinal rush border membrane enzymes, its stability was not affected by thecoexistence of food ingredients; Caco-2cell model and animal experiments showedthat dietary phosphorus level could affect the absorption of peptide-calcium complex,but its bioavailability did not rely on vitamin D intake; fish scale protein peptide couldreduce the inhibition of phytic acid on a certain degree. |
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