Preparation Of High Loading Beta-carotene Microcapsules And Analysis Of Factors Affecting Its Bioefficacy

Vitamin A deficiency(VAD)is mainly caused by inadequate intake of VA and VA sources.VAD is widely distributed in the world,especially in low-income countries.?-carotene(BC)is one of the most effective VA sources.However,BC has poor chemical stability and low bioavailability in gastrointestinal tract,which greatly limits its application scope and effect of BC as a VA supplement.Embedding BC into a suitable carrier is a common method to improve its stability and bioavailability.However,due to the low solubility of BC in almost all solvents,the preparation of high load carrier of molten BC has always been the difficulty.Based on this,we studied the preparation of high loading BC microcapsules by high temperature melting method.By optimizing the formulation of microcapsules(core material,wall material and core wall ratio)and preparation process(melting,emulsification and spray drying),we explored the factors that affect the loading and physicochemical stability of microcapsules.At the same time,animal models suitable for evaluating the bioavailability of high loading microcapsules BC were constructed.Based on the in vitro and in vitro digestion models,the key factors affecting the bioavailability of BC in high temperature melt microcapsules(MO)and crystallographic microcapsules(CW)were analyzed systematically,including BC physical status,lipid status,particle size and dose.It provides theoretical guidance for the construction of BC delivery system with high load,high stability and high bioavailability.The main research contents and results are as follows:Preparation of BC microcapsules with high load by melting method: To achieve the goal of high load,high embedding efficiency and long-term storage stability of MO microcapsules BC,to improve the oil phase's own antioxidant capacity,the wall material's oxygen barrier capacity and the wall material's antioxidant capacity,optimize the formulation and preparation process of BC microcapsules with high load.The results show that,compared with the traditional reactor heating method,the BC in the oil phase can be transformed into an amorphous state by using the 180 ? coil instantaneous melting process,and the degradation and isomerization phenomenon in the melting process can be effectively reduced.The melting retention rate of BC can reach 80%;adding 5% tocopherol in the oil phase can significantly improve the oxidation stability of BC,while adding 1.5% tocopherol in the wall material based on OSA starch 5% ascorbic acid can effectively improve the antioxidant capacity of the wall material itself;the use of small molecular carbonhydrates sucrose can significantly improve the oxygen barrier capacity of the wall material.When the ratio of sucrose to OSA starch reaches or exceeds 3:7,DSC and Raman confocal test show that the wall material has phase separation phenomenon,and the formation of dense outer sucrose phase can significantly improve the oxygen barrier capacity of the wall material.The SEM image analysis showed that the collapse probability of the microcapsule powder under the high energy electron beam bombardment also decreased with the increase of sucrose ratio.When the ratio of sucrose to OSA starch increased to 2:1,the microcapsule was round grape cluster,with complete powder structure and dense wall material.With the optimized composite core and wall materials mixed in the ratio of 1:3,the BC loading of MO microcapsules can be 3.7%,the embedding efficiency can be 96%,and the retention rate can still be more than 99% after 30 days storage in 60 degree oven.Establishment of BC bioavailability evaluation model in high load BC microcapsules: Optimize the evaluation methods and evaluation indexes of the commonly used in vitro static simulation gastrointestinal digestive model and VAD animal experimental model,so as to accurately evaluate the VA supplement,cumulative efficiency and the prevention and treatment effect of VAD brought by BC intake.An inbred Babl/C mouse was used as the experimental animal,and the VAD model was constructed with VA deficient diet.The results showed that 24 hours after single administration,blood pharmacokinetic determination and acute/accumulated concentration of BC in liver could not effectively evaluate the VA supplement efficiency of BC;After continuous feeding with non VA source feed,the VA level in liver and blood of mice decreased with feeding time,and the average VA equivalent(RAE)concentration in serum decreased to 0.1 mg/L four weeks later,which can be regarded as VAD status in rodents;The intake of two kinds of high load BC microcapsules can effectively improve the accumulation level of VA liver in mice and prevent mice from changing into VAD status.The bioavailability of BC microcapsules can be quantitatively evaluated with the comprehensive RAE accumulation efficiency of BC and retinol derivatives as indicators.Based on the results of bioavailability evaluation of animal model,the in vitro static digestion model was evaluated.The results showed that the bioavailability of MO microcapsules in vivo could be effectively predicted by using the traditional gastrointestinal digestion model with the BC micelle efficiency in small intestine phase as the index,but the bioavailability of crystal BC microcapsules CW(0.29%)determined by this evaluation method was far away lower than the results of animal experiments.By changing the shear conditions of digestion in vitro,applying additional shear and increasing the friction surface area to simulate the emulsification and shear force in gastrointestinal tract under physiological conditions,the transport efficiency of BC microcapsules prepared by melting and crystal method in vitro can match the bioavailability of MO and CW in vivo.The influence of BC physical status and coexistent oil on the bioavailability of BC in microcapsules: Besides MO and CW,two high loading BC microcapsules were introduced: unmelted emulsion based microcapsules(crystals dispersed in oil phase,CO)and crystal-emulsion microcapsules(crystals and shaped milk embedded in wall materials,CE).The results of animal experiments showed that: four kinds of BC microcapsules all showed a certain degree of improvement in the liver VA state of mice,the trend of liver VA accumulation efficiency was: CE > CW > MO > CO,BC accumulation bioavailability reached 68.6%,51.7%,46.7% and 15.5% respectively;At the same time,the in vitro bioaccessibility of the four kinds of BC microcapsules under the simulated shear condition had the same trend as that of the liver VA accumulation efficiency of animal experiments.The bioavailability of CE,MO and CO microcapsules was positively correlated with the extent of lipolysis in the simulated digestion system.When BC was present in the oil phase,the accumulation of VA in the liver of mice increased with the decrease of BC/oil ratio,the viscosity of oil phase increased rapidly with the increase of BC/oil ratio,and the particle size of oil droplet increased rapidly with the digestion of emulsified wall materials.The main inducement for the decrease of micelle efficiency and bioavailability of BC.When BC is independent of oil phase,the digestion of oil is not interfered by BC,and the higher extent of oil digestion improves the BC capacity of micelle phase,which is the main reason for the high bioavailability of CE.The micellization efficiency of CW microcapsules in vitro increases with the increase of shear strength in the system.The results of microscopic observation and particle size test show that under the presence of shear force,the crystal structure of BC is destroyed,the particle size is reduced,the specific surface area of BC particles is increased,and the solvation efficiency of BC in bile salt micelles is improved.Analysis of particle size and dose factors affecting BC bioavailability in high loaded BC microcapsules: Based on the proven correlation between BC micellization efficiency and bioavailability,three high loading BC microcapsules MO,CW and CE with high BC bioavailability were selected to study the effects of droplet size,crystal size and feeding dose on BC bioavailability.The results showed that the bioavailability of microcapsules could be effectively improved by reducing the particle size and decreasing the droplet size of the microcapsules.Among them,the low crystallite size and low droplet size CE microcapsules had the highest VA accumulation efficiency in mice liver.When compared with the positive control(retinol acetate)similar to VA intake equivalent,the VA liver accumulation efficiency(VA)was higher than that of the positive control group(19.7%,In BC).The bioavailability of BC in the three microcapsules was the same as that in the low dose.With the increase of gavage dose,the accumulation of VA in liver increased significantly,but the bioavailability decreased.The accumulation of VA in liver of mice could reach the level of healthy mice four weeks later.Based on the realization of the preparation of three kinds of BC microcapsules with high loading capacity and high bioavailability,the existing status of BC in the carrier and the mechanism of the effect of coexisting oil on the transport efficiency of BC were preliminarily clarified.It was proposed that the melting and small-size crystal BC microcapsules could be used as potential high-efficiency VA supplements under the condition of coexistence of low-size oil.