Studies On Safety Assessment Of Carotenoids And Their Accumulation Mechanism In Yolk

Most citrus is used as raw materials in conventional food processing. As the result of processing, the peel is the major by-product. In fact, citrus peel contains abundant carotenodis. Therefore, citrus peel could be used as source material for carotenoids extraction. Ultrasound extraction was applied to extract carotenoids form citrus peel and their stability was studied under ultrasound treatment. Meanwhile, a safety assessment for citrus peel as the source of carotenoid was performed. Furthmore, in order to study the accumulation mechanism of carotenodis in yolk, carotenoid-binding proteins were isolated from yolk and their composition and structure were investigated. Research results were concluded as follows.1. Ultrasound extraction was used for the extraction of carotenoids from citrus peel. The effects of different factors, including particle size of the material, the extraction solvent, solid/solvent ratio, extraction time, temperature and electrical acoustic intensity on the extraction yield of lutein from citrus peels by ultrasound extraction were investigated. The extraction yield of lutein was significantly affected by particle size. Dichloromethane caused the degradation of lutein during ultrasound extraction. In comparison with classical extraction, acetone showed a pronounced higher extraction yield of lutein during ultrasound extraction, higher extraction yield was obtained at lower solvent consuming, and the extraction yield of ultrasound extraction did not easily arrive at equilibrium. Because of the thermal effect of ultrasound, the extraction yield of ultrasound extraction was not affected when the extraction temperature was higher than 15℃. The extraction yield of ultrasound extraction had a peak value at 484.08 W/cm2.2. The mixture of carotenoids extracted from citrus peel (Nanfengmiju, Citrus reticulata Blanco) was tested for subacute oral toxicity. In this study, dose levels of 0, 200,500 and 2000 mg/kg body weight/day were administered by gavage to 10 Wistar rats/sex/group for 28 days. During the study, detailed clinical observations were subjected outside the cage before and after treatment. Body weights were determined. Food consumption was measured and mean food consumption was calculated. Ophthalmologic examinations were performed twice, once before rats were administered to the test substance and again immediately before they were sacrificed. For the last 16 h before sacrifice, urine of rats was collected and analyzed. Blood samples were collected from the abdominal aorta for clinical chemistry and hematological evaluations. All rats at final sacrifice were subjected to necropsy and histopathology. No statistically significant, dose-related effect on food consumption, food efficiency, body weight gain, clinical signs or ophthalmoscopic parameters was observed in any treatment group. Urinalysis, hematological, blood coagulation and serum biochemical examination as well as necropsy or histopathology showed that no observed adverse effect was found. These findings suggested that the No-Observed-Adverse-Effect Level for the mixture of carotenoids extracted from citrus peel was at least 2000 mg/kg body weight/day.3. The mechanism of selective accumulation of carptenoids in yolk was investigated. In general, maize and maize products were main dietary sources of carotenoids in poultry industry and oxycarotenoids were the major carotenoids in those products. Therefore, poultry products were lack of hydrocarbon carotenoids. P-carotene and lycopene were used as source of natural pigment in poultry standard diet of hens, respectively. Supplementation with β-carotene or lycopene significantly increased the daily feed intake after one week. The egg laying rate, egg weight and feed conversion were not significantly affected by β-carotene or lycopene supplement. Supplementation with β-carotene or lycopene resulted in a significant effect on Roche colour fan scores and yolk colour a*. Furthermore, dietary lycopene supplementation significantly decreased malondialdehyde content in egg yolk. Both β-carotene and lycopene were able to be transferred form the feed to yolk. However, the doposition efficiency of β-carotene and lycopene were much lower in comparison with oxycarotenoids. In order to study the selective accumulation mechanism of carotenoids in yolk, the low density lipoprotein (LDL) binding different carotenoids in yolk was isolated. The composition and conformation of LDL were not affected by binding different carotenoids. In fact, LDL is structured such that the more polar constituents, such as proteins, phospholipids and cholesterol, are situated toward the outside of the LDL. The more nonpolar components, such as triglycerides and cholesterol esters, are located toward the inner core of the LDL. The more polar carotenoids are easier to be located in the LDL than those hydrocarbon carotenoids. This might be the reason why hydrocarbon carotenoids showed lower deposition efficiency in yolk than more polar carotenoids.4. Dry-heated Maillard reaction was applied in the preparation of protein-polysaccharide conjugates. Reaction mixtures containing soy protein isolate (SPI) and maltodextrin (1:1 weight ratio) were dry-heated at 60 ℃ and 79% relative humidity for three days.The mixtures of SPI and gum acacia (GA) were dry-heated at the same condition for one week. The conjugate of SPI-MD showed lower levels of free amino groups and higher degree of graft, which indicated that reaction between SPI and MD developed much faster than reaction between SPI and GA. The solubility of SPI at isoelectric point was improved remarkably after grafting with MD or GA. The grafted SPI showed significantly higher levels of emulsifying properties than SPI and the emulsifying properties of SPI-GA conjugate were much better than SPI-MD. Decreases of lysine and arginine contents after the graft reaction indicated that these two amino acid residues attended the covalent linkage between SPI and MD or GA. The graft reaction reduced surface hydrophobicity and fluorescence emission maximum value because of a shielding effect of the polysaccharide chain bound to proteins. The results of secondary structure suggested that grafted SPI had decreased the levels of a-helix, β-sheet and β-turn and increased unordered coils level. SPI-GA conjugate was applied to encapsulate the lycopene. X-ray powder diffraction (XRD) measurements were performed to study the structure of lycopene in the particles. For lycopene particles, diffraction peaks of pure lycopene disappeared. The interaction of lycopene with SPI-GA conjugate might decrease the crystalline order and crystallite size of lycopene effectively. The first-order reaction was used to determine the degradation of lycopene in the particles.