Interactions Among Phytate,Calcium/Magnesium And Proteins In Soymilk And Their Effects On Protein Aggregation

Phytic acid(phytate,PA),calcium,and magnesium ions are the important charged small molecular components that coexist with proteins in soybean seeds.They could interact with proteins in complicated manners thus affect the aggregation behaviors of proteins.This study focused on the interactions among the endogenous phytate,calcium/magnesium and proteins within the soymilk system and analyzed the existing forms of these small molecules in soybean as well as their changes in soymilk and tofu processing.In addition,how phytate affected the heat-induced protein aggregation and Ca2+-induced coagulation was examined and discussed in depth.The roles played by phytate in the soybean processing were made clear,which provided guidance to modulating the quality of soymilk and tofu products from the perspective of small molecular compounds.Detailed studies and results were stated as follows:1.The occurrence and distribution of endogenous phytate,calcium,and magnesium in raw soymilk and the complexation stability of the "protein-phytate/calcium/magnesium" complexes were analyzed by virtue of equilibrium dialysis.Results indicated that in raw soymilk,roughly half of the phytate,calcium,and magnesium formed dialyzable but unionized salts,with the rest of them being bound to the 7S protein,most probably as "PA-Ca/Mg-7S" and "7S-Ca/Mg" complexes.These electrostatically bound phytate and metal ions were heat-stable,but can be dissociated from proteins when subjected to lower pH,intensified ionic strength,and exogenous Ca2+.Chemical denaturants did not lead to the dissociation of the original "PA-Ca/Mg-7S" complexes but gave rise to the further binding of phytate to proteins in soymilk.2.The interactions between phytate and proteins when soymilk was subjected to heat treatment and their effects on the heat-induced soy protein aggregation were discussed.It was indicated that in comparison with the 7S protein,the 11S protein had a considerably higher phytate-binding capacity(ca.30 mg/g protein).Phytate exerted a significant salting-in effect on natural 11S protein as well as an inhibitory role on its heat-induced aggregation.When raw soymilk was heated to temperatures above 75?,the denaturation and dissociation of 11S proteins led to the electrostatic interactions between phytate and the positively charged basic polypeptides,which occurred at an "opening period" when the buried basic amino acids exposed to the surface of the basic polypeptides.The binding reaction made roughly 1/3 of the free phytate transferred to soymilk particulate protein fraction after heat treatment at 95?.Ionic strength higher than 500 mM and alkaline pHs could inhibit their binding.In addition,when heating soymilk,phytate competed with the ?,?' subunits of the 7S protein and the acidic polypeptides of the 11S protein,for the binding sites on the basic polypeptides,which led to a decrease in both the ratio of soymilk particulate proteins and the average particle size.3.The effects of phytate on the Ca2+-induced soymilk coagulation process as well as the gel properties were further investigated.It was found that the coagulation rate constant(k)was significantly negatively correlated with soybean phosphorus content.Both free and bound phytate exerted a buffering effect to decelerate the protein coagulation process.The removal of free phytate led to a significantly lower activation energy for the protein aggregation reaction,increasingly accelerated coagulation,coarser gel network,as well as poorer water-holding capacity of the resultant gels.4.The mechanism of the buffering effect phytate exerted in the coagulation of soymilk was discussed.It was found that when the coagulant(Ca2+)concentration in soymilk was low(1-4 mM),the endogenous phytate preferably consumed Ca2+ to form stable calcium-phytate complexes,so as to lower the odds of direct interaction between Ca2+ and proteins,which led to the occurrence of a "buffering period" as the first stage of the Ca2+-induced coagulation.When the phytate concentration in soymilk decreased,its buffering effect was weakened or lost,which led to the easier neutralization of the surface negative charges and the intensified protein aggregation at lower Ca2+ concentration.5.Aiming at explicating the relationship between coagulation rate and gel formation and properties in general,the Ca2+-induced coagulation rate was controlled by altering coagulation variables and the chemical components in soymilk,with the gel properties compared.Results indicated that when Ca2+ or Mg2+ was used as the coagulant,protein coagulation rate played a predominant role in determining the strength and water-holding capacity of the gel network.Within a proper range(k?0.5-2.5*10-3s-1),the coagulation rate was highly positively correlated with the gel strength.However,either too quick or too slow coagulation was unfavorable to the formation of soymilk gels.