A carotenoid preparation based on red-fleshed navel oranges and a method of preparation

Abstract

The present application relates to the technical field of carotenoid enrichment, in particular to a carotenoid preparation based on red-fleshed navel orange and a preparation method thereof. The carotenoid preparation is rich in phytoene and phytofluene. The method comprises the following steps: during the planting period of the red-fleshed navel orange, the fruit is subjected to shading treatment; after the fruit is ripe, the juice sac residues in the fruit are picked and obtained; the juice sac residues are subjected to post-treatment to obtain the carotenoid preparation. The preparation method realizes directional enrichment of phytoene and phytofluene, significantly improves the content of the two in the juice sac residues and their in-vitro digestion bioavailability; the juice sac residues can be used as a preferred raw material for extraction and product development of natural linear carotenoids, not only high-quality carotenoid preparations can be obtained, but also a new technical path for high-value utilization of citrus juice processing by-products is provided.

Description

Technical Field

[0001] This invention relates to the field of carotenoid enrichment technology, and more specifically, to a carotenoid product based on red-fleshed navel oranges and its preparation method. Background Technology

[0002] Carotenoids are a class of fat-soluble natural products widely found in plants. They not only participate in plant photosynthesis, photosynthesis, photosynthesis, hormone synthesis, and fruit coloring, but are also important nutrients and functional components in the human diet. Current research and industrial development mainly focus on colored carotenoids such as lycopene, β-carotene, lutein, and zeaxanthin, while relatively little attention has been paid to linear colorless carotenoids such as phytoene and hexahydrolycopene.

[0003] Hydroxylenic acid (H2O) and phytohexylenic acid (H2O) are located upstream in the lycopene biosynthesis pathway and are key intermediates in the conversion of colorless carotenoid precursors into colored carotenoids in plants. Both exhibit characteristic absorption primarily in the ultraviolet (UV) region and have been shown to enter the human bloodstream through dietary intake, and can be detected in various tissues. Current research indicates that H2O and H2O have potential applications in skin photoprotection, oxidative stress regulation, and food and cosmetic development. Therefore, obtaining natural, stable, and high-content plant-based raw materials for H2O and H2O has significant industrial value.

[0004] The analytical content of carotenoids in raw materials is not equivalent to the proportion that can be absorbed and utilized by the human body. Fat-soluble carotenoids need to be released from plant tissues and their lipid-bound structures during digestion and enter a mixture of micelles formed by bile salts and lipid digestion products before they can be further absorbed by intestinal epithelial cells. The proportion of the target substance entering the micelle phase after digestion is called bioavailability, which reflects the efficiency of the target component's release from the food matrix and completion of micellization.

[0005] Currently, existing carotenoid products have complex and costly preparation processes, limited bioavailability, and limited commercial value. Summary of the Invention

[0006] The technical problem to be solved by the present invention is to provide a carotenoid product based on red navel orange and its preparation method.

[0007] The technical solution of the present invention to solve the above-mentioned technical problems is as follows: This invention provides a method for preparing a carotenoid product based on red-fleshed navel oranges, wherein the carotenoid preparation is rich in phytoene and lycopene. The preparation method includes the following steps: S1. During the planting period of red-fleshed navel oranges, the fruit is shaded. S2. After the fruit is ripe, harvest it and obtain the juice sac residue from the fruit; S3. Post-process the juice cell residue to obtain the carotenoid product.

[0008] Based on the above technical solution, the present invention can be further improved as follows.

[0009] Furthermore, the variety of red-fleshed navel orange is Cara Cara red-fleshed navel orange.

[0010] Furthermore, in step S1, the planting period is from the green ripening stage of the fruit to the mature harvesting stage, and the shading treatment is bagging.

[0011] Furthermore, the bagging begins on one of the days between 165 and 185 days after flowering.

[0012] Furthermore, the bagging method involves fixing the bag opening to the fruit stem, maintaining a gap between the bag body and the fruit surface, and keeping the bottom of the bag open.

[0013] Furthermore, in step S2, after peeling the fruit, it is crushed and squeezed to remove the juice, and the resulting non-liquid solid is the juice cell residue.

[0014] Furthermore, in step S3, the post-processing methods include room temperature drying and / or freeze drying.

[0015] The present invention also provides a carotenoid product based on red navel orange prepared by the method described above, wherein the carotenoid product contains 250 μg / g dry weight of 8-hydrolycopene and 100 μg / g dry weight of 6-hydrolycopene.

[0016] Preferably, in the carotenoid products, the bioavailability of 8-hydrolycopene is greater than or equal to 35%, and the bioavailability of hexa-hydrolycopene is greater than or equal to 20%.

[0017] The present invention also provides a carotenoid food additive, including the carotenoid products based on red navel oranges as described above.

[0018] The beneficial effects of this invention are as follows: (1) Screening to obtain high-accumulation germplasm suitable for targeted regulation This invention compared 31 citrus germplasm materials. The results showed that there were significant differences in the accumulation levels of phytoene and hexahydrolycopene among different red-fleshed citrus varieties, and the red-fleshed phenotype itself is not necessarily accompanied by a large accumulation of these two linear colorless carotenoids.

[0019] In two samples of Cara Cara red navel oranges from different sources, the content of phytolycopene was approximately 212–259 μg / g DW, and the content of phytolycopene was approximately 63–110 μg / g DW, which was significantly higher than that of most of the tested citrus materials.

[0020] Therefore, a specific high-accumulation metabolic background is an important basis for the significant regulatory effect obtained by this invention.

[0021] (2) The main processing and utilization tissues of 8-hydrolycopene and 6-hydrolycopene were identified. This invention further separates the fruit into the yellow peel layer, the sac membrane, the juice sac residue, and the juice.

[0022] The results showed that 8-hydrolycopene and 6-hydrolycopene were mainly distributed in the cell membrane and juice sac residue, while their content in liquid juice was relatively low.

[0023] This indicates that the juice sac residue formed after juice processing is not simply a low-value byproduct, but rather an important natural enrichment site for phytoene and lycopene.

[0024] (3) Increase the content of phytoene in juice vesicle residue After bagging, the phytoene content in the juice cell residue of Cara Cara red navel oranges increased from about 241 μg / g DW to about 284 μg / g DW, an increase of about 43 μg / g DW.

[0025] (4) Increase the content of phytohexopene in juice vesicle residue After bagging, the phytohexene content in the juice cell residue of Cara Cara red navel oranges increased from about 66 μg / g DW to about 104 μg / g DW, an increase of about 38 μg / g DW.

[0026] (5) Simultaneously improve the bioavailability of phytoene and lycopene. An increase in the content of a target ingredient does not necessarily lead to an increase in its percentage of bioavailability.

[0027] The present invention further discovers that bagging treatment increases the in vitro digestibility of phytoene in juice cell residue from about 30% to about 38%, and the bioavailability of phytoene from about 18% to about 25%.

[0028] Therefore, the present invention not only increases the initial content of the target component, but also simultaneously improves its digestion, release and micellization efficiency.

[0029] (6) Achieve high-value utilization of by-products from citrus juice cell processing The juice sac residue obtained by the present invention can be directly processed into freeze-dried products, and can be further used as raw materials for the development of food, food ingredients, nutritional products or other natural products, providing a new technical route for the high-value utilization of citrus processing by-products. Attached Figure Description

[0030] Figure 1 This is a comparison chart of the content of phytoene and hexahydrolycopene in the pulp of different citrus varieties in Example 1 of the present invention. Figure 2 Phenotypic images of mature fruits from the unbagged control group and the bagged treatment group of Qingjia navel oranges and Cara Cara red-fleshed navel oranges in Embodiment 3 of the present invention; Figure 2 In the middle, A represents the unbagged control group. Figure 2 Group B is the bagging treatment group; Figure 3 This is a comparison chart of the contents of phytoene, lycopene and lycopene in the peel, sac, juice sac residue and juice of Qingjia navel oranges and Cara Cara red-fleshed navel oranges at three different time periods in the unbaked control group and the bagged treatment group. Figure 4 This is a comparison chart of the bioavailability of major carotenoids in the unbagged control group and the bagged treatment group of Cara Cara red navel orange juice cell residue in Example 5 of the present invention. Detailed Implementation

[0031] The principles and features of the present invention are described below. The examples given are only for explaining the present invention and are not intended to limit the scope of the present invention.

[0032] The present invention relates to a method for preparing carotenoid products based on red-fleshed navel oranges, wherein the carotenoid preparation is rich in phytoene and hexahydrolycopene. The preparation method includes the following steps: S1. During the planting period of red-fleshed navel oranges, the fruit is shaded. S2. After the fruit is ripe, harvest it and obtain the juice sac residue from the fruit; S3. Post-process the juice sac residue to obtain carotenoid products.

[0033] The present invention relates to a method for preparing carotenoid products based on red-fleshed navel oranges. Targeting red-fleshed navel oranges with high accumulation and metabolic characteristics of phytoene and hexahydrolycopene, the method employs shading treatment during the growing period to achieve targeted enrichment of phytoene and hexahydrolycopene, significantly increasing the content of both in the juice sac residue and their bioavailability in vitro. Although juice sac residue is usually produced as a byproduct during juice processing, it is actually an important enrichment site for phytoene, hexahydrolycopene, and lycopene, and can be used as a preferred raw material for the extraction and product development of natural linear carotenoids.

[0034] The method described above in this invention not only yields high-quality carotenoid products, but also provides a new technical path for the high-value utilization of by-products from citrus juice processing.

[0035] Preferably, the Cara Cara red-fleshed navel orange variety is used. Experiments have verified that Cara Cara red-fleshed navel oranges exhibit significant high accumulation of phytoene and hexahydrolycopene. Further analysis of the fruit into the pericarp, sclerotium, juice sac residue, and juice revealed that the two target components are not uniformly distributed throughout the fruit, but are mainly concentrated in the sclerotium and juice sac residue, with relatively low levels in the juice. Further research unexpectedly found that continuous shading during the green ripening stage not only increases the content of phytoene and hexahydrolycopene in the juice sac residue of mature Cara Cara red-fleshed navel oranges, but also simultaneously improves their bioavailability after simulated in vitro digestion.

[0036] In the preparation method of the present invention, the planting period in step S1 is specifically from the green ripening period of the fruit to the mature harvesting period, and the shading treatment is bagging.

[0037] Preferably, the specific processing procedure of step S1 is as follows: during the green ripening period of the fruit, select fruits that are normally developed on the outer periphery of the tree canopy, free from pests and diseases, free from mechanical damage, and of basically the same size for bagging; use shading fruit bags to cover each fruit one by one, with the bag opening fixed at the fruit stem, maintaining a gap between the bag and the fruit surface, and maintaining necessary gas and water vapor exchange; after bagging, keep the fruit continuously shaded until it is ripe and harvested.

[0038] Different production areas may adjust the specific bagging time according to local flowering period, climate conditions and fruit ripening period, but the fruit should be kept in shade continuously from the green ripening period until ripe harvest.

[0039] Preferably, the fruit bag is a double-layered bag with a yellow outer layer and a black inner layer.

[0040] Preferably, the fruit bags are inspected regularly during the bagging period, and any damaged, detached, or water-infiltrated fruit bags are replaced promptly to ensure continuous processing.

[0041] Preferably, the bagging period is 165–185 days after flowering, with the 175th day being the optimal time.

[0042] The above-mentioned step S1 of the present invention has clear processing conditions and is simple to operate in the field, which has the basis for large-scale application in red-fleshed navel orange producing areas.

[0043] In the preparation method of the present invention, step S2 involves peeling the fruit, crushing and squeezing it, and then removing the liquid juice through pressing, juicing, centrifugation or other juice separation treatments. The resulting non-liquid solids are the juice cell residue.

[0044] In the preparation method of the present invention, the post-processing in step S3 includes room temperature drying and / or freeze drying.

[0045] To minimize the exposure of sclerotium residue to room temperature, light, and air, the obtained sclerotium residue was not piled up but rapidly frozen in liquid nitrogen in small batches during the extrusion process. Each batch of sclerotium residue weighed approximately 3g, and the maximum time from extrusion to immersion in liquid nitrogen did not exceed 10 minutes. Each batch was ensured to be completely frozen before the next batch was rapidly frozen in liquid nitrogen. This prevented incomplete freezing of the sclerotium interior from affecting the efficiency of subsequent freeze-drying. After all batches were rapidly frozen, they were allowed to stand for 2 minutes, then transferred to a light-proof container and stored at -80°C. Before freeze-drying, the sclerotium residue was removed from the -80°C environment and pre-frozen again in liquid nitrogen under light-proof conditions until the entire sample was completely frozen. It was then immediately transferred to a SCIENTZ-10N / A freeze dryer for vacuum freeze-drying for 7 days. This second pre-freezing in liquid nitrogen was to reduce the rewarming or partial thawing of the sclerotium residue during removal, transfer, and loading, ensuring the sample remained fully frozen before entering the freeze-drying stage. The freeze-dried juice sac residue was immediately ground into powder under light-proof and low-temperature conditions, and then stored at -80℃ after being strictly sealed to obtain freeze-dried juice sac residue products rich in phytoene and hexahydrolycopene.

[0046] The carotenoid products of the present invention based on red-fleshed navel oranges have 250 μg / g DW of 8-hydrolycopene and 100 μg / g DW of hexahydrolycopene.

[0047] Preferably, the 8-hydrolycopene content is 283.96±34.97 μg / g dry weight, and the 6-hydrolycopene content is 103.79±13.08 μg / g dry weight.

[0048] In this type of carotenoid product, when measured using an in vitro simulated digestion method, the bioavailability of 8-hydrolycopene is greater than or equal to 35%, and the bioavailability of 6-hydrolycopene is greater than or equal to 20%.

[0049] Preferably, the bioavailability of 8-hydrolycopene is 37.70±0.08%, and the bioavailability of 6-hydrolycopene is 24.89±0.13%.

[0050] The carotenoid products based on red-fleshed navel oranges of the present invention can be used to prepare food, food ingredients, nutritional supplements, or cosmetic raw materials. During processing, light-protected, low-temperature, and low-oxygen exposure conditions are preferably used to reduce the oxidation, isomerization, and degradation of phytoene and hexahydrolycopene.

[0051] The carotenoid food additives of the present invention include carotenoid products based on red navel oranges as described above.

[0052] Preferably, this type of carotene food additive may also include other excipients acceptable in the food industry.

[0053] The effects of the present invention will be specifically described below through specific embodiments.

[0054] Example 1: Screening of citrus germplasm with high accumulation of phytoene and hexahydrolycopene In this embodiment, 31 citrus germplasm materials were selected, and the composition and content of carotenoids were extracted from the pulp of mature fruits. The specific extraction method is as follows: Accurately weigh 5 mg of freeze-dried citrus pulp sample powder from 31 different production areas, place them in 1.5 mL centrifuge tubes, add 250 μL of methanol, and vortex mix thoroughly for 1 min.

[0055] Then add 500 μL of chloroform, vortex for 1 min, and let stand or mix for 20 min under light-protected and low-temperature conditions. Add 250 μL of ultrapure water, vortex for 1 min, and centrifuge at 15000 r / min for 5 min to form an aqueous phase, a protein precipitate layer, and an organic phase. Aspirate the lower organic phase and transfer it to a new centrifuge tube, avoiding aspiration of the intermediate protein precipitate layer. Add another 500 μL of chloroform to the original centrifuge tube, centrifuge at 15000 r / min for 5 min, collect the lower organic phase, and combine it with the organic phase obtained in the first step. Concentrate the combined organic phase to dryness under vacuum under light-protected and low-temperature conditions.

[0056] Throughout the entire process of extracting carotenoids, light exposure, temperature rise, and oxygen exposure should be minimized.

[0057] Chromatographic analysis was performed on the carotenoid components and contents of each sample. The specific analytical methods are as follows: The concentrated sample was redissolved in 100 μL of ethyl acetate, filtered through a 0.22 μm organic phase filter membrane, and then transferred to a brown sample vial.

[0058] The analysis was performed using an Ultimate 3000 ultra-high performance liquid chromatography system equipped with a diode array detector. The chromatographic column was a YMC C30 carotenoid-specific column with dimensions of 150 mm × 3.0 mm and a particle size of 3 μm.

[0059] The injection volume was 10 μL, the column temperature was 35 ℃, and the flow rate was 0.8 mL / min.

[0060] Mobile phase A is a mixed solution of methanol and methyl tert-butyl ether in a volume ratio of 50:50; mobile phase B is a mixed solution of methanol, water and methyl tert-butyl ether in a volume ratio of 6:3:1.

[0061] The elution program was as follows: 0–19 min, the mobile phase was linearly changed from 30% A and 70% B to 100% A; 19–24 min, 100% A was maintained; 24–25 min, the mobile phase was restored to 30% A and 70% B; 25–29 min, the initial ratio was maintained for equilibration.

[0062] The phytoene octahydrolycopene and phytoene hexahydrolycopene were qualitatively identified based on the retention time, UV-Vis absorption spectral characteristics, and peak order of the corresponding standards.

[0063] Quantification is performed by establishing an external standard curve using corresponding reference standards; for target substances for which corresponding reference standards are temporarily unavailable, equivalent quantification can be performed using carotenoid reference standards with similar structures, and this should be clearly indicated in the test results.

[0064] like Figure 1 As shown, most of the tested materials had low levels of phytoene and lycopene. In the two Cara Cara red navel orange samples from Huazhong Agricultural University and the Zhijiang experimental site, the phytoene content was approximately 212–259 μg / g DW, and the lycopene content was approximately 63–110 μg / g DW, both significantly higher than in the other tested materials.

[0065] Compared to other red-fleshed citrus fruits such as Starbucks grapefruit, Huazhong Agricultural University red grapefruit, Red Willow, Majia grapefruit, and Flame grapefruit, Cara Cara red navel oranges contain at least 15 times more phytolysin and 10 times more phytolysin.

[0066] Cara Cara red-fleshed navel oranges from two different sources showed similar high accumulation characteristics, indicating that the high accumulation of phytoene and hexahydrolycopene is a relatively stable metabolic characteristic of this variety, rather than an accidental change in a single sample.

[0067] In contrast, the phytoene content in the pulp of Qingjia navel oranges is relatively low, and phytoene was not detected under the detection conditions of this example. It mainly accumulates esterified lutein pigments, which is significantly different from the linear carotenoid accumulation type of Cara Cara red navel oranges.

[0068] Therefore, in subsequent embodiments, Cara Cara red navel oranges were selected as the bagging treatment material, and Qingjia navel oranges were used as the comparative material for different types of carotenoid accumulation.

[0069] Example 2: Bagging treatment of red-fleshed navel oranges during the growing period The experiment in this embodiment was conducted at the Citrus Breeding Center of Huazhong Agricultural University, and the specific steps are as follows: Cara Cara red-fleshed navel oranges were used as the target material, and Qingjia navel oranges were used as the comparison material. Four trees of each variety with basically the same growth were selected and the same water, fertilizer, pruning and pest and disease management measures were adopted.

[0070] At the green ripening stage of the fruit, 175 days after flowering, randomly select normally developed fruits that are free from pests and diseases, have no obvious mechanical damage, and are of roughly the same size from the outer edge of the tree canopy. Cover each fruit with a double-layered fruit bag, yellow on the outside and black on the inside. Fix the opening of the fruit bag to the fruit stem, maintain a gap between the bag and the fruit surface, and keep the bottom of the bag open to allow for gas and water vapor exchange.

[0071] Meanwhile, unbagged fruits of the same variety from the same tree were used as a control.

[0072] During the bagging period, regularly inspect the fruit bags and replace any damaged, fallen, or water-infiltrated bags in a timely manner to keep the fruit continuously shaded from the green ripening stage to the mature harvest.

[0073] Samples were taken at 200, 230, and 270 days after flowering. At each time point, 12-15 fruits were randomly collected from both the bagged group and the unbagged control group around the outer edge of the tree canopy.

[0074] Example 3: Fruit tissue separation and juice vesicle residue acquisition Cara Cara red-fleshed navel oranges obtained in Example 2 at various stages and Qingjia navel oranges were brought back to the laboratory on the day of harvest. The appearance characteristics of the fruits were recorded and cross-sections were photographed. The photographs are shown below. Figure 2 As shown in the figure, the cross-sectional observation results of mature fruit indicate that the Cara Cara red-fleshed navel orange flesh becomes darker red after bagging treatment.

[0075] The pericarp, pericarp, juice vesicle residue, and juice of each fruit were separated separately. After separating the juice vesicles from the pericarp, pericarp, and other tissues, the liquid juice was removed by squeezing or pressing the juice vesicles; the remaining non-liquid juice vesicle solids were defined as juice vesicle residue. The resulting tissue samples were immediately subjected to low-temperature treatment.

[0076] Samples used for component analysis were rapidly frozen in liquid nitrogen, stored in the dark at -80°C, and then freeze-dried under vacuum and ground into powder.

[0077] Example 4: Determination of Carotenoid Content in Different Fruit Tissues of Two Samples After Bagging Treatment The freeze-dried sample obtained in Example 3 was extracted according to the method described in Example 1.

[0078] The concentrated carotenoid sample was reconstituted with 100 μL of ethyl acetate, filtered through a 0.22 μm organic phase filter membrane, and then transferred to a brown sample vial.

[0079] Analysis was performed using an Ultimate 3000 ultra-high performance liquid chromatography system equipped with a diode array detector.

[0080] The YMC C30 carotenoid-specific chromatographic column with dimensions of 150 mm × 3.0 mm and a particle size of 3 μm was used.

[0081] The injection volume was 10 μL, the column temperature was 35℃, and the flow rate was 0.8 mL / min.

[0082] Mobile phase A is a mixture of methanol and methyl tert-butyl ether in a volume ratio of 50:50; Mobile phase B is a mixture of methanol, water and methyl tert-butyl ether in a volume ratio of 6:3:1.

[0083] The elution procedure is as follows: From 0 to 19 min, the ratio of 30%A / 70%B changed linearly to 100%A; 19–24 min, maintain 100% A; After 24–25 minutes, the body recovers to 30%A / 70%B. Set the initial conditions for 25–29 minutes to achieve equilibrium.

[0084] The target carotenoids were qualitatively identified based on the retention time, UV-Vis absorption spectral characteristics, and peak order of the standards, and quantitatively identified using a standard curve.

[0085] Example 5: Effect of bagging treatment on the content of linear carotenoids in different fruit tissues The levels of phytoene, lycopene, and lycopene in the peel, sac, juice residue, and juice of Cara Cara red navel oranges and Qingjia navel oranges in both bagged and unbagged groups were determined.

[0086] The results showed that there were significant differences in the response of the two types of navel oranges to bagging treatment. Figure 3 ).

[0087] No significant accumulation of phytoene and hexahydrolycopene was observed in Qingjia navel oranges before and after bagging, indicating that the metabolic effect of bagging is constrained by the variety's own carotenoid metabolic background.

[0088] In Cara Cara red navel oranges, phytoene and lycopene are mainly distributed in the peel and juice sac residue, while their content in the peel and juice is relatively low.

[0089] After bagging treatment at maturity, the phytoene content in the juice vesicle residue increased from approximately 241 μg / g DW to approximately 284 μg / g DW, an increase of approximately 43 μg / g DW.

[0090] The amount of phytohexopene increased from about 66 μg / g DW to about 104 μg / g DW, an increase of about 38 μg / g DW.

[0091] The above results indicate that the combination of a high-accumulation germplasm background and pre-harvest shading treatment can promote the accumulation of phytoene and hexahydrolycopene in juice vesicle residue.

[0092] In the mid-to-late stages of fruit development, the content of phytoene and hexahydrolycopene in the fruit capsule decreased after bagging, while the content of the two target components in the juice vesicle residue increased accordingly. This indicates that bagging altered the accumulation and distribution of linear colorless carotenoids in different fruit tissues, causing them to accumulate more in the juice vesicle residue, which is the main material that can be processed and utilized.

[0093] The above results demonstrate that the selection of highly accumulating germplasm is fundamental to achieving significant improvements in bagging efficiency. This invention, by combining suitable materials with continuous bagging at specific times, achieves the targeted enrichment of phytoene and lycopene in juice vesicle residue.

[0094] Example 6: Low-temperature stabilization of juice cell residue and preparation of carotenoid products Juice is extracted or pressed from ripe Cara Cara red navel oranges, and the pulp is collected after removing the liquid juice.

[0095] To reduce the oxidation, isomerization, and degradation of phytoene and hexahydrolycopene during processing, the resulting juice cell residue is rapidly frozen under light-proof and low-temperature conditions. In this embodiment, liquid nitrogen is used for rapid freezing of the juice cell residue.

[0096] The frozen sac residue was stored at low temperature and then freeze-dried under vacuum while remaining fully frozen. The freeze-dried sac residue was then ground into powder under light-proof and low-temperature conditions and sealed for storage to obtain a carotenoid product rich in phytoene and hexahydrolycopene.

[0097] The specific liquid nitrogen treatment batches, freeze-drying times, equipment models, and storage conditions used in this embodiment are used to illustrate the feasibility of the present invention and do not constitute a limitation on the scope of protection of the present invention.

[0098] Example 7: In vitro simulated digestion and bioaccessibility assay This embodiment uses the INFOGEST static in vitro digestion simulation framework, and adjusts it according to the characteristics of the juice cell residue sample of red navel orange. It simulates digestion in the oral cavity, stomach and small intestine stages in sequence. The specific experimental process is as follows: (1) Sample preparation: The freeze-dried juice cell residue obtained in Example 3 was ground into powder, 0.5 g of sample was accurately weighed, 5 mL of water was added, and the mixture was thoroughly mixed to prepare a sample homogenate.

[0099] (2) Preparation of simulated digestive solutions: As shown in Table 1, simulated saliva, simulated gastric juice, and simulated intestinal juice were prepared respectively. The composition of the simulated digestive solutions is shown in the table below. The final volume of each simulated digestive solution was 500 mL. Except for CaCl2·2H2O, the other components could be prepared in advance as a 1.25-fold concentrated solution and stored at -20 ℃. CaCl2·2H2O was added before the start of the digestion experiment to reduce calcium salt precipitation.

[0100] Table 1. Preparation of simulated digestive fluid (3) Simulated oral digestion: 68.18 mg of α-salivary amylase was mixed with 4 mL of simulated saliva concentrate, 0.025 mL of 0.3 mol / L CaCl2·2H2O solution was added, and the volume was adjusted to 5 mL with ultrapure water.

[0101] The above mixture was preheated at 37 °C for 5 min and then added to the sample homogenate. The pH was adjusted to 7.0 with 1 mol / L NaOH solution. The resulting mixture was then shaken and digested at 37 °C and 150 r / min for 2 min to obtain the oral digested sample.

[0102] (4) Simulating gastric digestion: Mix 66.78 mg of pepsin with 8 mL of simulated gastric juice concentrate, add 0.005 mL of 0.3 mol / L CaCl2·2H2O solution, and bring the volume to 10 mL with ultrapure water.

[0103] The above mixture was combined with the oral digestion sample, and the pH was adjusted to 3.0 with 1 mol / L HCl solution. The resulting mixture was then digested by shaking at 37 °C and 150 r / min for 2 h to obtain the gastric digestion sample.

[0104] (5) Simulating small intestinal digestion: Mix 20 mg of pancreatic enzyme, 20.62 mg of pancreatic lipase and 0.41 g of bovine bile powder with 16 mL of simulated intestinal fluid concentrate, add 0.04 mL of 0.3 mol / L CaCl2·2H2O solution, and bring the volume to 20 mL with ultrapure water.

[0105] The above mixture was combined with the gastric digested sample, and the pH was adjusted to 7.0 with 1 mol / L NaOH solution. The resulting mixture was then digested by shaking at 37 °C and 150 r / min for 2 h to obtain the small intestinal digested sample.

[0106] (6) Micellar phase separation and target analyte determination: After small intestinal digestion, the digestive fluid was centrifuged at 4 ℃ and 10000 r / min for 1 h. After centrifugation, the sample separated into an insoluble precipitate and an upper micelle phase. The micelle phase was aspirated, filtered through a 0.45 μm microporous membrane, and then subjected to carotenoid extraction and chromatographic detection.

[0107] The bioavailability of the target carotenoid is calculated using the following formula: Bioaccessibility (%) = Total amount of target carotenoids in micelle phase ÷ Total amount of target carotenoids in sample before digestion × 100%.

[0108] The content of the target substance in the micelle phase and the sample before digestion was calculated after being converted into the total amount according to the corresponding sample volume, mass and dilution factor.

[0109] Each treatment was performed with independent biological replicates. Data are expressed as mean ± standard deviation. Significance was tested using independent samples tests or analysis of variance, depending on the experimental design. *, **, and *** indicate P < 0.05, P < 0.01, and P < 0.001, respectively.

[0110] like Figure 4 As shown, bagging treatment increases the proportion of various carotenoids entering the mixed micelle phase after in vitro digestion.

[0111] Among them, the bioavailability of phytoene increased from about 30% to about 38%, and the bioavailability of hexahydrolycopene increased from about 18% to about 25%. The bioavailability of β-carotene, β-cryptoxanthin and lycopene also increased to varying degrees.

[0112] The above results demonstrate that the present invention not only increases the initial content of phytoene and hexahydrolycopene in juice cell residue, but also improves their release and micellization efficiency during digestion, achieving a simultaneous increase in the content of the target components and their bioavailability.

[0113] Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention. Those skilled in the art can make changes, modifications, substitutions and variations to the above embodiments within the scope of the present invention.

Claims

1. A method for preparing carotenoid products based on red-fleshed navel oranges, characterized in that, The carotenoid preparation is rich in phytoene and hexahydrolycopene; The preparation method includes the following steps: S1. During the planting period of red-fleshed navel oranges, the fruit is shaded. S2. After the fruit is ripe, harvest it and obtain the juice sac residue from the fruit; S3. The juice cell residue is post-processed to obtain the carotenoid product.

2. The method for preparing carotenoid products based on red-fleshed navel oranges according to claim 1, characterized in that, The variety of the red-fleshed navel orange is Cara Cara red-fleshed navel orange.

3. The method for preparing carotenoid products based on red-fleshed navel oranges according to claim 1, characterized in that, In step S1, the planting period is from the green ripening stage of the fruit to the mature harvesting stage, and the shading treatment is bagging.

4. The method for preparing a carotenoid product based on red-fleshed navel oranges according to claim 3, characterized in that, The bagging should begin on one of the days between 165 and 185 days after flowering.

5. The method for preparing a carotenoid product based on red-fleshed navel oranges according to claim 3, characterized in that, The bagging method involves fixing the bag opening to the fruit stem, maintaining a gap between the bag body and the fruit surface, and keeping the bottom of the bag open.

6. The method for preparing a carotenoid product based on red-fleshed navel oranges according to claim 1, characterized in that, In step S2, the fruit is peeled, crushed, and squeezed to remove the juice. The resulting non-liquid solid is the juice cell residue.

7. The method for preparing a carotenoid product based on red-fleshed navel oranges according to claim 1, characterized in that, In step S3, the post-processing methods include room temperature drying and / or freeze drying.

8. A carotenoid product based on red-fleshed navel orange prepared by the method according to any one of claims 1-7, characterized in that, In the carotenoid products, the mass of 8-hydrolycopene is greater than or equal to 250 μg / g dry weight, and the mass of 6-hydrolycopene is greater than or equal to 100 μg / g dry weight.

9. A carotenoid product based on red-fleshed navel oranges according to claim 8, characterized in that, Of the carotenoid products, the bioavailability of 8-hydrolycopene is greater than or equal to 35%, and the bioavailability of 6-hydrolycopene is greater than or equal to 20%.

10. A carotenoid food additive, characterized in that, Including carotenoid products based on red navel oranges as described in claim 8 or 9.

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