| Sea buckthorn oil is one kind of functional oils extracted from sea buckthorn seeds, pulp or whole berries, there is much difference between oils from different parts of berry, but all of them contain a lot of nutrients such as PUFAs, VE, phytosterols, carotenoids and phenolic compounds. It also has many health effects for human to make it will be developed well. However, oil content of sea buckthorn seeds, pulp or berries is low, it is from 1.4% to 13.7% due to factors such as varieties and regional influence. Existed extraction methods are difficult to enrich nutrients of oil, and nutritional function has not been researched deeply, which will limit its development and utilization. Therefore, it is very important to study how to improve yield of sea buckthorn oil, promote more bioactive components dissolve in oil and evaluate its functional activity to improve the quality and development of sea buckthorn oil. In this paper, microwave pretreatment coupled with solvent extraction(MPE) and accelerated solvent extraction(ASE) were established according to oil yield, acid values and peroxide values, on this basis, the effect of solvent extraction(SE), ultrasound assisted solvent extraction(UAE) and two new methods on physicochemical properties, nutritional quality, thermal stability and antioxidant activity of sea buckthorn seed oil, pulp oil and whole berry oil was investigated. Finally, we chose pigmented rabbits which had similar retina structure with human being as light-induced damage model and discussed the retina protection effect of sea buckthorn seed oil, pulp oil and whole berry oil extracted by MPE on visual function, retina cell protection, antioxidant activity, inflammatory factor regulation and apoptosis protein expression. The main purpose of this paper was providing theoretical basis and technical support for increasing oil yield, improving quality of sea buckthorn oil and developing eyesight protection functional foods.The main research results were as follows:(1) New extraction methods for sea buckthorn oil were established: The new extraction method of microwave pretreatment coupled with solvent extraction was established for the yield and quality of oil. The optimal microwave pretreatment time was 2min for sea buckthorn seeds, 2 min for pulp and 3min for whole berries under 560W/120 g at frequency of 2450 MHz. Under the optimal conditions, oil yield was 7.94%, 5.73% and 6.91%, respectively. Compared to SE, MPE significantly increased yield of sea buckthorn seed oil, pulp oil and whole berry oil, the increasing proportion was 9.36%, 3.61% and 7.13%, respectively. Peroxide values were 1.77, 2.86 and 1.82 mmol/kg, respectively. Acid values were 7.55, 9.13 and 8.56 mg KOH/g, respectively. With orthogonal optimization of ASE, the optimum extraction parameters were 90℃, 5min and 2 times for seed oil; 90℃, 7min and 3 times for pulp oil; 100℃, 7min and 3 times for whole berry oil. Under the optimal conditions, oil yields were 7.21%, 6.12% and 6.92%, respectively. Yields of pulp oil and whole berry oil were increased by 10.67% and 7.29% with ASE. Acid values were 13.01, 13.38 and 11.57 mg KOH/g, respectively, peroxide values were 1.39, 2.87 and 1.58 mmol/kg, respectively. Physicochemical properties of oil from two new methods were superior to existing industrial standards.(2) Effect of four extraction methods on physicochemical properties and nutrients of sea buckthorn oil was investigated: Through the comparisons of three kinds of oil we found that the p-anisidine value of whole berry oil was higher than pulp oil and seed oil. Acid values, peroxide values, K232 and K270 of pulp oil was highest, while seed oil was lowest, which showed that physicochemical quality of seed oil was better than pulp oil and whole berry oil. There were huge differences of nutrients in oil from different parts of berry. In terms of α-linolenic acid and vaccenic acid(C18:1n-7), they were 30.92%-31.56% and undetected in seed oil, 7.92%-8.01% and 2.07%-2.17% in pulp oil, and 4.96%-5.37% and 11.77%-13.55% in whole berry oil, respectively. Phytosterols content of seed oil was 1617-1906 mg/100 g, which was higher than that of whole berry oil and pulp oil, and the main individual phytosterols was β-sitosterol. Vitamin E and carotenoids content of pulp oil were higher than that of seed oil and whole berry oil, total phenolic content of whole berry oil was higher than pulp oil and seed oil, the content of lutein, lycopene and β-carotene was 68.49-92.06, 45.94-68.07 and 85.49-104.82 mg/100 g, which was 4 times, 2 times and 7 times higher than seed oil, respectively, and the content of total phenols was 2 times higher than seed oil.The new methods had significant effect on enrichment of total phenols. Compared to SE, total phenols content of seed oil extracted from ASE, MPE and UAE increased by 2.37 times, 1.11 times and 0.26 times, that of pulp oil increased by 2.34 times, 0.25 times and 0.27 times and whole berry oil increased by 2.76 times, 0.31 times and 0.12 times, respectively. The content of carotenoids also increased obviously, the content of carotenoids in seed oil increased by 0.68 times, 0.67 times and 0.50 times, respectively, and for whole berry oil, it increased by 0.09 times, 0.27 times and 0.12 times, respectively, but there was no significant change of pulp oil. ASE method had the best effect for VE enrichment. MPE had the best effect for phytosterols enrichment, compared to SE, the content of phytosterols of seed oil, pulp oil and whole berry oil respectively increased by 3.9%, 23.9% and 22.9%. All above results indicated that there were different effects of different extraction methods on nutrients enrichment of sea buckthorn oil, and enrichment effect of ASE was best overall, but ASE would influence acid values, p-anisidine values, K232 and K270.(3) The effect of different extraction methods on thermal stability and antioxidant activity of sea buckthorn oil was investigated: Compared with three kinds of oil, the sequence of oxidation induction time and initial oxidation temperature were pulp oil>whole berry oil>seed oil, which showed that thermal stability of pulp oil was best, followed by whole berry oil and seed oil. Antioxidant activity of pulp oil and whole berry oil was obviously higher than seed oil in DPPH assay, ABTS assay and FRAP assay, while antioxidant activity of pulp oil and whole berry oil was lower than seed oil in FIC assay. Compare with four extraction methods, the ability of improving thermal stability of sea buckthorn oil of ASE was highest, followed by MPE, UAE, and SE. In the above four assays, antioxidant activity of ASE was highest, followed by MPE, UAE and SE. In terms of seed oil, antioxidant activity of seed oil extracted by ASE, MPE and UAE in DPPH assay was 2.77 times, 1.45 times and 1.18 times higher than SE. Antioxidant activity of seed oil extracted by ASE, MPE and UAE in ABTS assay was 4.28 times, 2.21 times and 1.21 times. Antioxidant activity of seed oil extracted by ASE, MPE and UAE in FRAP assay was 3.11 times, 1.76 times and 1.12 times. Antioxidant activity of seed oil extracted by ASE, MPE and UAE in FIC assay was 2.36 times, 1.66 times and 1.12 times. And the trends of antioxidant activity of pulp oil and whole berry oil were the same as seed oil. The results showed that two new methods of ASE and MPE could be used as effective means of improving thermal stability and antioxidant activity of sea buckthorn oil.As indicated from correlation analysis, the correlation coefficient of unsaturated fatty acids and oxidation induction time or initial oxidation temperature was-0.814 and-0.910, respectively. The correlation coefficient of VE, carotenoids and total phenols with initial oxidation temperature was 0.962, 0.861 and 0.714, respectively, and for oxidation induction time was 0.858, 0.686 and 0.626, respectively. The correlation coefficient of DPPH, ABTS and FRAP with total phenols content was 0.975, 0.995 and 0.998, respectively. The above results showed that unsaturated fatty acids was the dominant factor of decreasing of oxidation stability of oil, and enrichment of antioxidants in oil could significantly improve oxidation stability and antioxidant activity of oil.(4) The preventive effect of sea buckthorn oil on light-induced retinal damage in pigmented rabbits was investigated. The results showed that sea buckthorn seed oil, pulp oil and whole berry oil had protective effect on light induced retinal rods and cones injuries, and could increase the thickness of the retina ONL. Compare to model group, ONL thickness of seed oil group, pulp oil group and whole berry oil group increased by 15.09%, 12.54% and 30.31%, respectively. They could also increase the content of SOD and CAT under the oxidative stress and reduce 3-nitro tyrosine level to adjust the oxidative stress ability, CAT increased by 27.31%, 17.10% and 21.06%, SOD increased by 39.37%, 37.63% and 40.77%, while 3-nitrotyrosine decreased by 23.83%, 2.59% and 5.70%, respectively. The expression of HO-1 mRNA and Nrf-2 mRNA was increased by sea buckthorn oil, which resulted in raising level of antioxidant gene regulation, HO-1 mRNA and Nrf-2 mRNA expression were increased by 103.16% and 86.73%, 51.58% and 9.73%, 91.58% and 56.64%, respectively. Sea buckthorn oil had effect of suppressing Bax apoptosis protein expression, promoting Bcl-2 antiapoptotic proteins expression, significantly inhibiting retinal Caspase-3 expression and photoreceptor cells apoptosis and making it to return to normal levels, retinal Caspase-3 expression decreased by 46.0%, 34.0% and 6.0%, respectively. The results also revealed that the interleukin-1β mRNA expression of seed oil group, pulp oil group and whole berry oil group respectively decreased by 18.13%, 31.36% and 33.3% so as to inhibit the production of inflammatory factors. In addition, vascular endothelial growth factor content in retinal tissue membrane was reduced, which could improve the retinal microcirculation. All above results indicated that sea buckthorn oil had the certain effect of healing light-induced retinal damage of pigmented rabbits, and it could be used as the material of development of eye protection functional foods. |
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