Double-strain synergistic fermentation composite vegetable oil as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of cosmetic raw materials, and particularly relates to double-strain synergistic fermentation composite vegetable oil as well as a preparation method and application thereof. According to the invention, an integrated technical scheme of preparing the fermented composite vegetable oil through composite pretreatment-double-strain synergy-staged low-temperature fermentation is adopted, so that a plurality of links such as raw material treatment efficiency, strain synergy efficiency, fermentation process control and the like can form tight synergy; compared with the traditional crude oil processing or single fermentation process, the compound vegetable oil has the advantages that the purity, the stability, the functionality, the safety and the mildness of the product are obviously improved, and a brand-new technical path is provided for the development of natural fermentation cosmetics.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of cosmetic raw materials, and particularly relates to a double-strain synergistic fermentation compound plant oil and a preparation method and application thereof. BACKGROUND

[0002] Traditional fermentation processes mostly use single strains (such as lactic acid bacteria, yeast or bacillus) to ferment single plant oils (such as olive oil, coconut oil), which has the problems of single type of metabolic products and low conversion rate of active substances (usually less than 60%). For example, CN117821273A discloses a method for preparing cosmetics by fermenting green tea seed oil with Candida, but the hydrolysis rate of triglycerides in the fermentation product is still low compared with other single colony fermentation, and the problem of poor oxidative stability of the oil after fermentation is not solved.

[0003] In existing mixed strain fermentation technology, the strain combination is mostly selected based on experience (such as CN119523845A uses Escherichia coli, Staphylococcus aureus, Lactobacillus and yeast for co-fermentation), and the metabolic synergy mechanism between strains is not deeply analyzed, resulting in easy occurrence of competitive inhibition of strains (such as pH or nutrition competition) in the fermentation process, and large fluctuation of product yield.

[0004] In the existing technology, plant oils are mostly directly mixed by physical methods, and are not synergistically enhanced at the molecular level through fermentation. For example, CN118634176A discloses an anti-aging skincare product containing 12 kinds of plant oils, but the raw oil is not biologically modified, although the antioxidant effect of the fermented plant oil is improved to some extent, the plant oil raw material types are more and the formula is more complex, and the fermentation period is longer, the raw material and labor costs are higher. SUMMARY

[0005] The purpose of the present application is to overcome the shortcomings and deficiencies of the prior art and provide a preparation method of double-strain synergistic fermentation compound plant oil, which effectively improves the polyphenol content in the prepared fermented compound plant oil through the innovative design of double-strain synergistic metabolic regulation, compound plant oil directional modification and low-temperature phased fermentation process, significantly reduces the content of hexanal and fatty acids, and stably improves the DPPH clearance rate while significantly reducing the irritation, realizes the comprehensive advantages of significant effect, high safety and controllable cost, and provides a new technical path for the development of natural fermented cosmetics.

[0006] In order to achieve the above-mentioned application purposes, the technical scheme adopted by the present application is as follows:

[0007] In a first aspect, the present application provides a preparation method of fermented compound plant oil, comprising the following steps:

[0008] S1, cellulase is added to the composite vegetable oil for enzymatic hydrolysis, and then ultrasonic treatment is performed to obtain pretreated composite vegetable oil;

[0009] S2, the pretreated composite vegetable oil is mixed with a fermentation medium, and then inoculated with Lactobacillus plantarum and Rhodotorula mucilaginosa for co-fermentation culture to obtain a fermentation liquor;

[0010] S3, the fermentation liquor is centrifuged, and the upper oil phase is collected, and then the upper oil phase is subjected to demulsification treatment and ultrafiltration to remove impurities, so as to obtain an ultrafiltrate;

[0011] S4, the ultrafiltrate is evaporated and concentrated to obtain a concentrated oil phase product, and then an antioxidant is added to the concentrated oil phase product, so as to obtain the fermented composite vegetable oil;

[0012] The composite vegetable oil comprises olive oil, camellia seed oil and sea buckthorn oil;

[0013] In step S2, the specific method of fermentation culture is as follows: pre-fermentation is performed at 10-15 DEG C for 22-26 h, and then main fermentation is performed at 25-30 DEG C.

[0014] The integrated technical scheme of the present application adopts composite pretreatment-bacterial strain cooperation-phase low-temperature fermentation to prepare fermented composite vegetable oil, and can form close cooperation from multiple links such as raw material treatment efficiency, strain cooperation efficiency and fermentation process control, so that the final prepared composite vegetable oil realizes multiple goals of function enhancement and stimulation reduction, and the product purity, stability, functionality and safety mildness are significantly improved compared with traditional crude oil processing or single fermentation process.

[0015] The plant oil composite pretreatment process (enzymatic hydrolysis-ultrasonic combined treatment) is designed, the residual plant cell wall structure is broken through cellulase pretreatment combined with ultrasonic process, and the release amount of polyphenols in oil is increased. The strain metabolic complementary system is developed, Lactobacillus plantarum (Lactobacillus plantarum) CGMCC 1.557 is combined with Rhodotorula mucilaginosa (Rhodotorula mucilaginosa) CICC 33013, the former secretes lipase (activity ≥120 U / mL) to hydrolyze triglyceride, and the latter synthesizes β-carotene (≥1.2 mg / g) to neutralize free radicals, so that the metabolic path is complementary, and the product quality can be greatly improved compared with single strain fermentation. The two-stage low-temperature fermentation process (10-15 DEG C pre-fermentation and 25-30 DEG C main fermentation) is established, the oxidation of oil is reduced, and the specific metabolic products of the strain are released in stages (lipase is generated in the pre-fermentation stage, and antioxidant is synthesized in the main fermentation stage), so that the metabolic efficiency and product specificity are effectively improved, and the antioxidant property and other functions of the final composite vegetable oil product are enhanced.

[0016] In addition, based on the nutritional complementation and process adaptation design, the present application selects olive oil, camellia seed oil and sea buckthorn oil as the base oil for the above process treatment. The olive oil is rich in monounsaturated fatty acids, and the fatty acid composition of the camellia seed oil is close to that of the olive oil, so the two provide a high-quality base fatty acid skeleton for the compound oil; the sea buckthorn oil is rich in antioxidants such as vitamin E and natural carotenoids, which can not only enrich the antioxidant species of the final product, but also reduce the nutritional consumption of the strain for synthesizing beta-carotene, thereby assisting the antioxidant while reducing the metabolic burden of the fermentation strain. Therefore, by compounding the three specific plant oils, the present application not only solves the nutritional shortcomings of single oil, but also adapts to the metabolic needs of the fermentation strain, thereby further improving the efficacy of the finally prepared compound plant oil.

[0017] Preferably, the volume ratio of the olive oil, camellia seed oil and sea buckthorn oil is (6-7):(4-5):(2-3).

[0018] More preferably, the volume ratio of the olive oil, camellia seed oil and sea buckthorn oil is 7:(4-5):3.

[0019] It is found through experimental exploration that the amount and ratio of the olive oil, camellia seed oil and sea buckthorn oil have an important influence on the polyphenol content of the finally prepared compound plant oil, thereby affecting the antioxidant, anti-inflammatory and other effects of the product. When the three plant oil components are compounded in the above optimal ratio, the polyphenol content of the prepared compound plant oil is the highest.

[0020] Preferably, in step S1, the volume percentage content of the cellulase in the compound plant oil is 0.8-1%; and the specific conditions of the enzymolysis are: enzymolysis at 45-55℃ for 1.5-2.5h.

[0021] Preferably, the specific conditions of the ultrasonic treatment are: ultrasonic treatment at 40-60kHz for 15-25min.

[0022] Preferably, the compound plant oil further comprises peony seed oil, and the volume ratio of the peony seed oil to the olive oil is (1-2):(6-7).

[0023] The present application can further add peony seed oil to the olive oil, camellia seed oil and sea buckthorn oil for fermentation treatment, and the peony seed oil is rich in alpha-linolenic acid, which can compensate for the low content of alpha-linolenic acid in the remaining plant oils, so that the finally prepared compound plant oil realizes multiple functions, and the efficacy value is further improved compared with traditional compound plant oil or single fermented oil.

[0024] Preferably, in step S2, the volume ratio of the pretreated compound plant oil to the fermentation medium is (3-5):(5-7).

[0025] Preferably, the volume ratio of the pre-processed compound vegetable oil and the fermentation medium is (3-4):(6-7).

[0026] More preferably, the volume ratio of the pre-processed compound vegetable oil and the fermentation medium is 3:7.

[0027] It is found through experiments that the volume ratio of the fermentation medium and the compound vegetable oil directly and significantly affects the DPPH scavenging rate of the prepared fermented compound vegetable oil, and the increase of the proportion of the vegetable oil cannot improve the DPPH scavenging rate of the fermented compound vegetable oil, so the volume ratio between the two is crucial to the antioxidant effect of the final product. When the fermentation treatment is carried out with the above optimal ratio, the DPPH scavenging rate of the prepared fermented compound vegetable oil reaches 89%, and the antioxidant effect is the best.

[0028] Preferably, in step S2, the inoculation amount ratio of the Lactobacillus plantarum and the Rhodotorula mucilaginosa is (1-2):1.

[0029] More preferably, the inoculation amount ratio of the Lactobacillus plantarum and the Rhodotorula mucilaginosa is 2:1.

[0030] It is found through experiments that the inoculation amount ratio of the Lactobacillus plantarum and the Rhodotorula mucilaginosa also has an important influence on the DPPH scavenging rate of the final fermented compound vegetable oil. The Rhodotorula mucilaginosa needs trace oxygen to synthesize carotenoids, but too much oxygen will accelerate the oxidation of oil, and when the Lactobacillus plantarum and the Rhodotorula mucilaginosa are inoculated at a ratio of 2:1 (v / v), the micro-anaerobic environment created by the Lactobacillus plantarum is most conducive to the synthesis of antioxidant substances such as β-carotene by the Rhodotorula mucilaginosa, thereby improving the DPPH scavenging rate of the fermented vegetable oil.

[0031] Preferably, the total inoculation amount of the Lactobacillus plantarum and the Rhodotorula mucilaginosa is 6-8%.

[0032] Preferably, in step S4, the antioxidant includes rosemary extract.

[0033] Preferably, the mass percentage content of the rosemary extract in the fermented compound vegetable oil is 0.02-0.08%.

[0034] Preferably, the fermentation medium includes the following components: 15-25 g / L glucose, 5-10 g / L peptone, 5-15 g / L yeast extract, 1-3 g / L dipotassium hydrogen phosphate, 0.5-1.5 g / L magnesium sulfate, 0.5-1.5 g / L sodium chloride, 1-3 g / L Tween 80, 0.1-0.5 g / L FeSO4·7H2O, and 0.05-0.15 g / L MnSO4·H2O.

[0035] In a second aspect, the present application provides a fermented compound vegetable oil prepared by the preparation method.

[0036] The fermentation compound vegetable oil prepared by the preparation method has an acid value of ≤1.2 mg KOH / g, a free fatty acid content of ≤2.7%, a polyphenol content of ≥2.6 mg / g, a hexanal content of ≤0.22 ppm, and a DPPH clearance rate of ≥89%.

[0037] In a third aspect, the present application provides the use of the fermentation compound vegetable oil in cosmetics.

[0038] The cosmetic in the present application can be common cosmetics such as skin toner, emulsion, essence, cream, mask, and eye mask.

[0039] Compared with the prior art, the present application has the following beneficial effects:

[0040] Compared with the traditional fermentation cosmetic process raw material, the active ingredient polyphenol content of the plant oil after pretreatment is increased by 133%; the hexanal content of the fermentation plant oil prepared by the two-stage fermentation process is reduced by 3.8-5.7 times compared with the traditional single-strain fermentation, the acid value is reduced by 121%-164%, and the irritation is significantly reduced; the DPPH clearance rate of the plant oil prepared by the double-strain synergistic fermentation is increased by about 30% compared with the single-strain fermentation, and the plant oil has the advantages of significant efficacy, high safety, controllable cost, and the like, thereby providing a new technical path for the development of natural fermentation cosmetics. BRIEF DESCRIPTION OF DRAWINGS

[0041] Figure 1 The preparation process flow chart of the fermentation compound vegetable oil of the present application is shown in the figure;

[0042] Figure 2 The product separation diagram of the two-stage fermentation and constant temperature fermentation is shown in the figure;

[0043] Figure 3 The peak diagram of the fatty acid content detection of the fermentation compound vegetable oil of the present application is shown in the figure. DETAILED DESCRIPTION

[0044] The above content of the present application will be further described in detail through the specific implementation methods in the form of examples. However, this should not be understood as the scope of the above subject matter of the present application being limited to the following examples.

[0045] Unless otherwise specified, the reagents used in the examples and comparative examples are conventional reagents in the art, and can be obtained through commercial channels. The experimental operations not specifically described in the examples and comparative examples are conventional operations in the art or can be understood or known by those skilled in the art according to the existing technology or common knowledge mastered by them.

[0046] The Lactobacillus plantarum CGMCC 1.557 and Rhodotorula mucilaginosa CICC 33013 were purchased from Beina Biological Technology Co., Ltd.; the cellulase was purchased from Novozymes; the olive oil, camellia seed oil, sea buckthorn oil and peony seed oil were all purchased from Shanghai Yuanye Bio-Technology Co., Ltd., and were all crude plant oils.

[0047] The MRS culture medium comprises the following components: 10 g / L peptone, 8 g / L beef powder, 4 g / L yeast powder, 20 g / L glucose, 2 g / L potassium phosphate dibasic, 2 g / L diammonium hydrogen citrate, 5 g / L sodium acetate, 0.2 g / L magnesium sulfate, 0.04 g / L manganese sulfate and 1 g / L Tween 80.

[0048] The YPD culture medium comprises the following components: 10 g / L yeast extract, 20 g / L peptone and 20 g / L glucose.

[0049] The fermentation culture medium comprises the following components: 20 g / L glucose, 8 g / L peptone, 10 g / L yeast extract, 2 g / L potassium phosphate dibasic, 1 g / L magnesium sulfate, 1 g / L sodium chloride, 2 g / L Tween 80, 0.2 g / L FeSO4·7H2O and 0.1 g / L MnSO4·H2O.

[0050] Fatty acid content detection: 50 mg of the centrifuged and filtered fermentation oil was weighed, dissolved with methanol, diluted to 0.5 mg / mL, filtered with a 0.45 μm organic filter membrane to prepare a detection sample. An Xb-C18 column was selected, the mobile phase was 0.1% phosphoric acid aqueous solution: acetonitrile (15:85), the flow rate was 1 mL / min, the detection wavelength was 203 nm, the column temperature was 5°C, and the injection volume was 20 μL.

[0051] Polyphenol detection: the detection standard LS / T 6119-2017 was followed. Acid value detection: the detection standard LS / T 6107-2012 was followed. Hexanal detection: the detection standards T / SDYZC 013-2024 and GB 1886.149-2015 were followed. Antioxidant performance test (DPPH clearance rate detection): the detection standard GB / T 39100-2020 was followed.

[0052] Example 1

[0053] The present embodiment provides a preparation method of a fermentation composite plant oil, comprising the following steps:

[0054] (1) Composite plant oil pretreatment

[0055] The olive oil, camellia seed oil, and sea buckthorn oil were mixed by magnetic stirring at a volume ratio of 6:4:2 to prepare a composite base oil. After adding 0.8% cellulase, the volume ratio of peony seed oil to olive oil was 1:6, and the enzyme hydrolysis was carried out at 45°C for 2.5h by magnetic stirring. Then, the mixture was transferred into an ultrasonic reactor and treated at 40kHz for 25min to obtain the pretreated composite vegetable oil.

[0056] (2) Construction of fermentation co-culture system

[0057] The Lactobacillus plantarum was inoculated into a pH 6.5 MRS medium at an inoculation amount of 1%, and incubated at 37°C and 220rpm for 12-14h. The Rhodotorula mucilaginosa glycerol strain was inoculated into a pH 7.0 YPD medium at an inoculation amount of 2%, and incubated at 25°C and 220rpm for 48h. The sterilized fermentation medium was mixed with the pretreated composite vegetable oil at a ratio of 6:4 (v / v). The Lactobacillus plantarum and Rhodotorula mucilaginosa were mixed at a ratio of 2:1 (v / v), and inoculated into the fermentation co-culture system at an inoculation amount of 6%. In the pre-fermentation stage, the temperature was set to 10°C for static culture for 26h, and the aeration amount was 0.5VVM to induce the secretion of low-temperature lipase by Lactobacillus plantarum and hydrolysis of triglyceride. In the main fermentation stage, the temperature was increased to 25°C, the rotation speed was 300rpm, and the aeration amount was 1VVM for 48h to synthesize β-carotene and promote the accumulation of Rhodotorula mucilaginosa to inhibit the accumulation of free fatty acids.

[0058] (3) Centrifugal ultrafiltration treatment

[0059] The fermentation broth after two-stage fermentation was transferred to a tubular centrifuge, and the rotation speed was set to 8000rpm for 15min to separate the bacterial cells and the fermentation supernatant. The upper oil phase (containing emulsified oil) and the lower water phase (containing soluble metabolites) were collected. 1% sodium chloride solution (w / v) was added to the upper emulsified oil phase, heated in a water bath at 60°C for 10min, and mechanically stirred at 300rpm to destroy the oil-water interface film. The oil phase was subjected to cross-flow filtration using a 50kDa ceramic membrane at an operating pressure of 0.3MPa and a temperature of 25°C to remove residual bacterial cells and macromolecular proteins by ultrafiltration.

[0060] (4) Concentration treatment

[0061] The ultrafiltration concentrated liquid was placed in a rotary evaporator, and the temperature was set to 40°C, the vacuum degree was-0.09MPa, and the solvent (mainly water phase residue) was evaporated to obtain the concentrated oil phase product. Then, 0.05% rosemary extract (natural antioxidant) was added to obtain the fermentation composite vegetable oil.

[0062] Example 2

[0063] The present embodiment provides a preparation method of fermentation composite vegetable oil, which comprises the following steps:

[0064] (1) Pretreatment of composite vegetable oil

[0065] The olive oil, camellia seed oil, and sea buckthorn oil were mixed by magnetic stirring at a volume ratio of 7:4:3 to prepare a composite base oil. After adding 1% cellulase, the volume ratio of peony seed oil to olive oil was 2:7. The mixture was subjected to enzymatic hydrolysis at 55°C for 1.5 h under magnetic stirring. Then, the mixture was transferred into an ultrasonic reactor and treated at 60 kHz for 15 min to obtain a pretreated composite vegetable oil.

[0066] (2) Construction of fermentation co-culture system

[0067] Lactobacillus plantarum was inoculated into a pH 6.5 MRS medium at an inoculation amount of 1% and incubated at 37°C and 220 rpm for 12-14 h. Rhodotorula glycerolica was inoculated into a pH 7.0 YPD medium at an inoculation amount of 2% and incubated at 25°C and 220 rpm for 48 h. The sterilized fermentation medium was mixed with the pretreated composite vegetable oil at a ratio of 7:3 (v / v). Lactobacillus plantarum and Rhodotorula glycerolica were mixed at a ratio of 2:1 (v / v) and inoculated into the fermentation co-culture system at an inoculation amount of 8%. In the pre-fermentation stage, the temperature was set to 15°C and the culture was incubated for 22 h under aeration at a rate of 0.5 VVM. Lactobacillus plantarum was induced to secrete low-temperature lipase and hydrolyze triglycerides. In the main fermentation stage, the temperature was increased to 30°C, the rotation speed was 500 rpm, and the aeration rate was 1 VVM. Rhodotorula glycerolica was induced to synthesize β-carotene, thereby inhibiting the accumulation of free fatty acids.

[0068] (3) Centrifugation and ultrafiltration treatment

[0069] The fermentation broth after two-stage fermentation was transferred to a tubular centrifuge, and the rotation speed was set to 8000 rpm. The centrifugation time was 15 min to separate the bacterial cells and the fermentation supernatant. The upper oil phase (containing emulsified oil) and the lower aqueous phase (containing soluble metabolites) were collected. 1% sodium chloride solution (w / v) was added to the upper emulsified oil phase, and the mixture was heated in a water bath at 60°C for 10 min and mechanically stirred at 300 rpm to destroy the oil-water interface film. The oil phase was subjected to cross-flow filtration using a 50 kDa ceramic membrane at an operating pressure of 0.3 MPa and a temperature of 25°C. Ultrafiltration was performed to remove residual bacterial cells and macromolecular proteins.

[0070] (4) Concentration treatment

[0071] The ultrafiltration concentrated liquid was placed in a rotary evaporator, and the temperature was set to 40°C and the vacuum degree was -0.09 MPa. The solvent (mainly water phase residue) was evaporated to obtain a concentrated oil phase product. Then, 0.05% rosemary extract (natural antioxidant) was added to obtain the fermentation composite vegetable oil.

[0072] Example 3

[0073] The present example provides a preparation method of a fermented compound vegetable oil, which is only different from the example 2 in that in step (1), the olive oil, the camellia seed oil and the sea buckthorn oil are mixed in a volume ratio of 7:4:2.

[0074] Example 4

[0075] The present example provides a preparation method of a fermented compound vegetable oil, which is only different from the example 2 in that in step (1), the olive oil, the camellia seed oil and the sea buckthorn oil are mixed in a volume ratio of 7:5:3.

[0076] Example 5

[0077] The present example provides a preparation method of a fermented compound vegetable oil, which is only different from the example 2 in that in step (2), the Lactobacillus plantarum and the Rhodotorula mucilaginosa are mixed in a ratio of 1:1 (v / v).

[0078] Example 6

[0079] The present example provides a preparation method of a fermented compound vegetable oil, which is only different from the example 2 in that in step (2), the sterilized fermentation medium and the pretreated compound vegetable oil are mixed in a ratio of 6:4 (v / v).

[0080] Example 7

[0081] The present example provides a preparation method of a fermented compound vegetable oil, which is only different from the example 2 in that in step (2), the sterilized fermentation medium and the pretreated compound vegetable oil are mixed in a ratio of 5:5 (v / v).

[0082] Comparative Example 1

[0083] The present comparative example provides a preparation method of a fermented compound vegetable oil, which is only different from the example 2 in that in step (1), no cellulase is added for pretreatment.

[0084] Comparative Example 2

[0085] The present comparative example provides a preparation method of a fermented compound vegetable oil, which is only different from the example 2 in that in step (2), the fermentation strain is only Lactobacillus plantarum, i.e., only Lactobacillus plantarum is used to inoculate the fermentation co-culture system for fermentation culture in an inoculation amount of 8%, and the culture temperature is constant at 30°C.

[0086] Comparative Example 3

[0087] The present comparative example provides a preparation method of a fermented compound vegetable oil, which is only different from the example 2 in that in step (2), the fermentation strain is only Rhodotorula mucilaginosa, i.e., only Rhodotorula mucilaginosa is used to inoculate the fermentation co-culture system for fermentation culture in an inoculation amount of 8%, and the culture temperature is constant at 30°C.

[0088] Comparative Example 4

[0089] The present comparative example provides a preparation method of fermented compound vegetable oil, which is only different from Example 2 in that in step (2), the constant culture temperature of co-culture is 30℃, i.e. without using two-stage fermentation culture of pre-fermentation and main fermentation.

[0090] Comparative Example 5

[0091] The present comparative example provides a preparation method of fermented compound vegetable oil, which is only different from Example 2 in that in step (1), the vegetable oil only uses olive oil.

[0092] Comparative Example 6

[0093] The present comparative example provides a preparation method of fermented compound vegetable oil, which is only different from Example 2 in that in step (1), the vegetable oil only uses camellia seed oil.

[0094] Comparative Example 7

[0095] The present comparative example provides a preparation method of fermented compound vegetable oil, which is only different from Example 2 in that in step (1), the vegetable oil only uses sea-buckthorn oil.

[0096] Comparative Example 8

[0097] The present comparative example provides a preparation method of fermented compound vegetable oil, which is only different from Example 2 in that in step (4), the operation step of rotary evaporation is removed.

[0098] Comparative Example 9

[0099] The present comparative example provides a preparation method of fermented compound vegetable oil, which is only different from Example 2 in that in step (4), rosemary extract is not added.

[0100] Comparative Example 10

[0101] The present comparative example provides a preparation method of fermented compound vegetable oil, which is only different from Example 2 in that in step (2), Lactobacillus plantarum: Rhodotorula mucilaginosa = 3:1 (v / v) is mixed.

[0102] Effect Example

[0103] 1. Polyphenol content detection:

[0104] According to the polyphenol content detection standard LS / T 6119-2017, the pretreated vegetable oil raw materials and fermented compound vegetable oils after fermentation treatment prepared in Examples 1-4 and Comparative Examples 1, 5-7 are detected for polyphenol content, and the test results are shown in Table 1.

[0105] Table 1

[0106]

[0107] Polyphenols in plant oils have multiple effects such as antioxidant, anti-inflammatory, antibacterial, and penetration promotion, and play a core role in cosmetics, especially in anti-aging, soothing and natural preservation systems. Cellulase can precisely degrade cellulose in plant cell walls, significantly improving the release efficiency of polyphenols in complex plant oils, and the process is mild and environmentally friendly.

[0108] According to the data comparison of each example and comparative example in Table 1, cellulase has a significant effect on releasing polyphenols from enzymatic treatment of plant oils, especially in Example 4, the polyphenol content is increased by 133% compared with the blank control Comparative Example 1. At the same time, the treatment effect of cellulase on different components of plant oil also has a significant difference. In Comparative Examples 5-7, the polyphenol release amount of seabuckthorn oil is the highest after single plant oil is treated by cellulase, and the polyphenol release amount of camellia seed oil is the lowest. As shown by the polyphenol release amount data in Examples 1-4, the polyphenol content of the three plant oils in different proportions has certain differences, and the higher the proportion of seabuckthorn oil and camellia seed oil in the complex plant oil, the higher the polyphenol release amount. Compared with the polyphenol content in plant oil before fermentation, the polyphenol content in plant oil after fermentation in each example is increased to different degrees, and the use of double-strain mixing and two-stage temperature control fermentation can increase the release of polyphenol content in plant oil by 46-115%.

[0109] 2. Acid value and hexanal content detection:

[0110] The acid value of the fermented complex plant oil obtained in Examples 1-2 and 5 and Comparative Examples 2-4 and 8-10 was detected according to the detection standard LS / T 6107-2012, and the test results are shown in Table 2.

[0111] Table 2

[0112] Sample origin Acid value (mg KOH / g) Hexanal content (ppm) Example 1 1.26 0.34 Example 2 1.20 0.22 Example 5 1.16 0.25 Comparative Example 2 2.65 0.83 Comparative Example 3 2.79 1.26 Comparative Example 4 3.17 0.74 Comparative Example 8 4.67 - Comparative Example 9 1.89 - Comparative Example 10 3.75 1.46

[0113] During the fermentation of plant oil, lipase secreted by microorganisms will hydrolyze triglycerides into free fatty acids (FFA) and glycerol. Although a moderate amount of FFA may enhance certain functions (such as penetration promotion), excessive FFA content (usually >3%) can cause a series of problems, which need to be reduced to a safe range through process control. Acid value (AV) is a key indicator for measuring the content of free fatty acids in oil, and controlling acid value is crucial for ensuring product quality, safety and stability. Hexanal is one of the key markers of plant oil oxidative degradation, mainly produced by the oxidative decomposition of linoleic acid (C18:2) and linolenic acid (C18:3), and its content directly affects the degree of rancidity and the odor of the product. Through the synergistic metabolism of double strains and two-stage temperature control, the generation of hexanal can be significantly inhibited.

[0114] From the data comparison in Table 2, the acid value of Example 2 using the two-stage temperature fermentation process with double strains was reduced by 121% and 132% compared with Comparative Examples 2-3 single strain fermentation, and the content of hexanal was reduced by 277% and 473% respectively; compared with Comparative Example 4 double strain constant temperature fermentation, the acid value was reduced by 164%, and the content of hexanal was reduced by 236%. The fatty acid content of the fermented compound vegetable oil in Example 2 was detected (peak chart is shown in Figure 3 ), the peak time of linolenic acid was 11.212 min, and the peak area was only about 45.41%, the peak time of linoleic acid was 15.968 min, and the peak area was only about 14.65%, compared with the FFA content of Comparative Example 4, which was reduced from 5.2% to 2.7%.

[0115] The two-stage temperature fermentation process synthesizes lipase at low temperature in pre-fermentation, and synthesizes β-carotene at high temperature in main fermentation stage to promote Rhodotorula to consume FFA, which effectively reduces the content of free fatty acid in fermented vegetable oil to control the acid value range, and the sensory of the product after separation and purification is obviously better than that of Comparative Example 4 (see Figure 2 ); it can be seen from Example 2 compared with Comparative Example 8 that a large amount of short-chain FFA is removed by rotary evaporation operation during post-treatment of fermented vegetable oil, which effectively reduces the overall FFA content in the product, and compared with Comparative Example 9, the appropriate addition of rosemary extract, a natural antioxidant, in the product can also effectively reduce the oxidation of fatty acid dissociation in the product to reduce the acid value level of the product. Example 2 adjusts the inoculation ratio of Lactobacillus plantarum and Rhodotorula, mainly optimizing the metabolic balance in the double strain fermentation process. Lactobacillus plantarum is responsible for hydrolyzing oil to produce FFA in the fermentation process, and Rhodotorula is responsible for consuming FFA to synthesize antioxidant substances. Compared with Comparative Example 10, the inoculation ratio of 2:1 in Example 2 can make the FFA generation and consumption rate basically match, avoid excessive accumulation of FFA and exceed the standard of hexanal, and the initial inoculation amount of 2:1 can make the double strains enter the metabolic vigorous period with a biomass of 1:1.

[0116] 3. DPPH clearance rate detection:

[0117] According to the detection standard GB / T 39100-2020, the antioxidant performance test (DPPH clearance rate detection) of the fermented vegetable oil obtained in Examples 1-7 and Comparative Examples 1-7, 10 was carried out, and the test results are shown in Table 3.

[0118] Table 3

[0119]

[0120]

[0121] DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging rate is a core index for evaluating antioxidant activity, and the higher the value, the stronger the antioxidant capacity of the vegetable oil.

[0122] As can be seen from the data in Table 3, the addition of cellulase during the pretreatment of the mixed vegetable oil effectively degrades the plant cell wall and releases the wrapped polyphenols, thereby effectively improving the antioxidant performance of the fermented vegetable oil. As can be seen from the comparison of the data in Examples 1-4 and Comparative Examples 5-7, the proportion of sea buckthorn oil and olive oil in the compound vegetable oil significantly affects the antioxidant performance of the fermented vegetable oil, and the antioxidant performance of the compound vegetable oil is significantly better than that of the single vegetable oil fermentation. The DPPH scavenging rate of the fermented oil in Example 2 is increased by 62% compared to that in Comparative Example 6. At the same time, compared with the unfermented mixed vegetable oil and single vegetable oil, the DPPH scavenging rate of the fermented vegetable oil is increased by different degrees (10.6%-34%). Compared with single-bacterium fermentation (Comparative Examples 2-3), the advantage of double-bacterium fermentation is more obvious, and the DPPH scavenging rate of the fermented oil is increased by 27%-30.9%. During the low-temperature pre-fermentation stage of Lactobacillus plantarum, lipase is secreted to hydrolyze triglycerides to generate free fatty acids, and the carboxyl group thereof is chelated with trace metal ions in the culture medium to inhibit the oxidative chain reaction. During the main fermentation stage, Rhodotorula glutinis synthesizes β-carotene, superoxide dismutase (SOD) and other antioxidant substances to further improve the DPPH scavenging rate of the fermented compound vegetable oil. As can be seen from the comparison of the data in Example 2 and Examples 6-7, the volume ratio of the fermentation medium to the compound vegetable oil directly and significantly affects the DPPH scavenging rate of the fermented vegetable oil. The increase in the proportion of vegetable oil cannot further improve the DPPH scavenging rate of the fermented vegetable oil. Rhodotorula glutinis needs trace oxygen to synthesize carotenoids, but too much oxygen will accelerate the oxidation of oil. When Lactobacillus plantarum and Rhodotorula glutinis are inoculated at a ratio of 2:1 (v / v), the micro-anaerobic environment created by Lactobacillus plantarum is most conducive to the synthesis of β-carotene and other antioxidant substances by Rhodotorula glutinis to improve the DPPH scavenging rate of the fermented vegetable oil. As shown in the experimental data in Table 3, the DPPH scavenging rate of Comparative Example 10 (3:1) is increased by 33%.

[0123] Finally, it should be noted that the above examples are only used to illustrate the technical solutions of the present application and do not limit the protection scope of the present application. Although the present application has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solutions of the present application can be modified or replaced by equivalents without departing from the essence and scope of the technical solutions of the present application.

Claims

1. A method for preparing a fermented compound vegetable oil, characterized by, The method comprises the following steps: S1, adding cellulase to the composite vegetable oil for enzymolysis, and then performing ultrasonic treatment to obtain pretreated composite vegetable oil; S2, mixing the pretreated composite vegetable oil with a fermentation medium, inoculating Lactobacillus plantarum and Rhodotorula mucilaginosa for co-fermentation culture to obtain a fermentation liquor; S3, centrifuging the fermentation liquor, collecting the upper oil phase, performing demulsification treatment on the upper oil phase, and then performing ultrafiltration to remove impurities to obtain an ultrafiltrate; S4, evaporating and concentrating the ultrafiltrate to obtain a concentrated oil phase product, and then adding an antioxidant to the concentrated oil phase product to obtain the fermented composite vegetable oil; The composite vegetable oil comprises olive oil, camellia seed oil and sea buckthorn oil; In step S2, the specific method of fermentation culture is pre-fermentation at 10-15℃ for 22-26h, and then main fermentation at 25-30℃.

2. The production method according to claim 1, wherein The volume ratio of the olive oil, the camellia seed oil and the sea buckthorn oil is (6-7):(4-5):(2-3).

3. The production method according to claim 1, wherein In step S1, the volume percentage content of the cellulase in the composite vegetable oil is 0.8-1%; and the specific conditions of enzymolysis are enzymolysis at 45-55℃ for 1.5-2.5h. The specific conditions of ultrasonic treatment are ultrasonic treatment at 40-60kHz for 15-25min.

4. The production method according to claim 1, wherein The composite vegetable oil further comprises peony seed oil, and the volume ratio of the peony seed oil to the olive oil is (1-2):(6-7).

5. The production method according to claim 1, wherein In step S2, the volume ratio of the pretreated composite vegetable oil to the fermentation medium is (3-5):(5-7).

6. The production method according to claim 1, wherein In step S2, the inoculation amount ratio of the Lactobacillus plantarum to the Rhodotorula mucilaginosa is (1-2):

1.

7. The production method according to claim 1, wherein In step S4, the antioxidant comprises rosemary extract.

8. The fermented composite vegetable oil prepared by the preparation method according to any one of claims 1-7.

9. The fermented composite vegetable oil according to claim 8 for use in cosmetics.

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