Analysis Of Factors Affecting The Bioaccessibility Of ?-carotene In The Emulsions Stabilized By OSA-modified Starches

?-Carotene has highest provitamin-A activity and strong antioxidant activity,which can help to reduce the risk of developing diseases induced by shortage of vitamin A,as well as cancer and cardiovascular diseases.?-Carotene is considered as a promising lipophilic functional nutrient.However,its low water solubility,poor chemical stability,and low oral bioavailability limit its application in functional foods.Emulsion is one of the most efficient delivery systems to overcome these limitations.The bioaccessibility of a lipophilic nutrient refers to the fraction of the ingested nutrient that is incorporated into mixed micelles or vesicles and becomes accessible for intestinal absorption.Generally,the bioaccessibility is been used to predict the bioavailability in the in vitro digestion studies.In this study,?-carotene emulsion-based delivery systems were prepared with octenyl succinic anhydride?OSA?-modified starches as biopolymer emulsifiers.The factors affecting the bioaccessibility of?-carotene in the emulsions were analyzed.These factors include structural characteristics of the emulsifier,food environment conditions,gastrointestinal tract conditions and properties of carriers.Their influence on the structure and stability of emulsions were studied.Then the influence and mechanism of these factors on the bioaccessibility of?-carotene were further investigated by in vitro digestion.This study may provide theoretical guidance for the design of emulsions stabilized by OSA-modified starches with high bioaccessibility of lipophilic bioactive components.The main research contents and results are as follows:The effects of molecular weight?M_w?and degree of substitution?DS?of OSA-modified starch on its emulsifying activity and digestive properties were firstly investigated.The result of surface tension showed the emulsifying activity of OSA-modified starches increased with a decrease in M_w and increased with an increase in the DS.The digestion of OSA-modified starch in the starch aqueous solutions was not affected by the M_w,and the digestion rate and extent decreased with increasing DS.Then,?-carotene emulsions were prepared with OSA-modified starches.The effects of DS of OSA-modified starches on the digestion of emulsion and the bioaccessibility of?-carotene were studied.The particle size,zeta-potential and microstructure of the emulsion during digestion were determined by particle size analyzer and confocal scanning laser microscopy.It was found that all the emulsions remained stable during the mouth stage and their particle size increased significantly after incubation under gastric conditions,with the extent of increase in the particle size being negatively correlated to the DS.Under gastric conditions,emulsions stabilized by OSA-modified starch with higher DS values had greater magnitudes of the negative zeta-potential and less flocculation and coalescence.The D-Glucose assay kit and pH-stat were used to determine the hydrolysis of starch and lipid,respectively.The results showed that the digestion behaviours of starch in emulsions were almost the same to those in the starch aqueous solutions.The rate and extent of lipid digestion enhanced with an increase in DS.The bioaccessibility of?-carotene was expressed as the micellization efficiency of?-carotene.When the DS increased from 1.58%to4.16%,the bioaccessibility of?-carotene was enhanced from 8.01±0.88 to 20.39±1.93%.The interaction between calcium ions(Ca²⁺)and OSA-modified starch in aqueous solutions was determined using turbidity,light scattering and Ca²⁺binding isotherms.The results showed that the interaction between Ca²⁺and OSA-modified starch was affected by pH.At pH 7.0,the addition of Ca²⁺induced an increase in the turbidity and a decrease in the the magnitudes of the negative zeta-potential of OSA-modified starch in starch aqueous solutions,which indicated the aggregation of starch molecules.At pH 3.0,there was no interaction between Ca²⁺and OSA-modified starch.Then OSA modified starch-stabilized emulsions were prepared and the effects of Ca²⁺on structure and stability of emulsions were investigated.It was found that at neutral pH,Ca²⁺decreased the emulsifying capacity of OSA-modified starch,with the particle size of the emulsions increased and emulsion stability decreased with an increase in Ca²⁺concentration.At pH 3.0,flocculation and creaming occurred in the emulsions,but the structure of emulsions was not affected by the concentration of Ca²⁺.Through simulated digestion,the influence of Ca²⁺in and pH of emulsions on the digestion of emulsion and the bioaccessibility of?-carotene were further examined.The results showed that the extent of lipid digestion in the emulsion and the bioaccessibility of?-carotene were dependent on the concentration of Ca²⁺,but independent on the pH of emulsions.The addition of Ca²⁺in the emulsion promoted lipid digestion but reduced the bioaccessibility of?-carotene.The results indicated that although the addition of Ca²⁺in the emulsions increased the particle size of OSA-modified starch emulsions,the promotion of Ca²⁺to lipid digestion was greater than its inhibition to lipid digestion through increasing particle size of emulsions.In order to explore the reason for the negative correlation between the extent of lipid digestion and the bioaccessibility of?-carotene at different Ca²⁺concentrations,the effects of Ca²⁺in the small intestinal juice on the digestion of emulsions and the bioaccessibility of?-carotene in OSA-modified starch emulsions were studied.The results showed that Ca²⁺had no effect on the structure of emulsions stabilized by high concentration of OSA-modified starches in the small intestinal juice without enzymes and bile salts.After incubation in the small intestine,it was found that higher concentrations of Ca²⁺resulted in more irregular aggregates and a greater rate and extent of lipid digestion.The bioaccessibility of?-carotene decreased from 30.84±8.66%to 0.48±0.18%when Ca²⁺concentration increased from 0 to20.0 mM.The lipolytic products and bile salts in the mixed micelles were analyzed by the GC and a bile acid kit.It was found that both of them decreased with an increase in the Ca²⁺concentration because of the formation of calcium soaps and calcium-bile acid complexes.A correlation analysis showed that the bioaccessibility of?-carotene had a highly statistically significant positive linear correlation with both lipolytic products?r=0.943?and bile salts?r=0.896?in mixed micelles,which suggestied that the lipolytic products and bile salts in the micelle phase play critical roles in the micellization of?-carotene.Emulsion gels were prepared by adding the gelling agent?whey protein isolate,WPI?to OSA-modified starch emulsions followed by heating.Human mouth,the human stomach simulator?HGS?and a static small intestinal digestion model were used to examine the effect of gel strength on the digestion of the emulsion and the bioaccessibility of?-carotene.Gel strength and mechanical properties of emulsion gels were determined by the rheological measurements,large deformation properties and fracture properties.It was found that higher ion strength and larger DS of OSA-modified starches contributed to higher strength of emulsion gels.The fracture force and fracture strain of emulsion gels decreased with an increase in ion strength.At low ion strength,the fracture force and fracture strain decreased with an increase in DS;at high ion strength,DS had little effect on the fracture properties of the gels?P<0.05?.The oral digestion results showed that the fragmentation of the gels was significantly linearly related to their mechanical properties.More chewing cycles were required when the gel hardness was greater,the Young's modulus was greater,or the fracture strain was higher,resulting in smaller median size of particles in masticated gels,i.e.higher extent of fragmentation of the gels.No emulsion was released from the protein network during oral digestion.The swelling ratio of emulsion gels in the simulated gastric juice without enzymes enhanced with a decrease in gel mechanical strength.During dynamic gastric digestion,the results of the changes in pH,the particle size and distribution of emptied digestas and emulsion droplets,and the microstructure of emptied digestas showed that pepsin has a stronger effect on the decomposition of emulsion gels with less strength.Smaller gel strength led to higher speed of proteolysis,stronger pH buffering capacity and faster release of emulsion droplets from the protein network matrix to the gastric juice.The digesta emptied in the early period?60 min?and late period?240 min?of gastric digestion were chosen for small intestinal digestion.The effects of the composition and structural properties of the emptied gastric digesta on the intestinal digestion of emulsions and the bioaccessibility of?-carotene were further investigated.The 60 min emptied gastric digesta of all gels only contained relatively large gel fragments.Smaller gel strength led to greater enzymatic decomposition of the gel fragments,earlier release of emulsion droplets from gel matrix and higher initial rate and extent of lipid digestion in the 60 min emptied gastric digestas.The 240min emptied gastric digesta of soft gels consisted of free emulsion droplets and small amount of gel fragments with small size,while those from hard gels had only gel fragments with small size but no free emulsion droplets.In the early period of intestinal digestion,the emulsions of former digestas had higher rates of lipid digestion than the latter ones;the extents of lipid digestion were close for the two types of emptied digestas,which could be attributed to their similar structure and composition obtained after early digestion in the small intestine phase.In addition,the extent of starch digestion,the rate and extent of lipid digestion and the bioaccessibility of?-carotene in the emulsions of the 240 min emptied gastric digestas were greater than those in the emulsions of the 60 min emptied gastric digestas.