A composition containing lemon peel extract, its preparation method and application

Abstract

The composition containing lemon peel extract disclosed in this invention comprises lemon peel extract, 1,2-hexanediol, 1,2-pentanediol, propylene glycol, and polyoxyethylene hydrogenated castor oil. The lemon peel extract is composed of lemon essential oil, lemon extract, and lemon absolute. The composition of this invention possesses a naturally soft, full-bodied, and long-lasting fragrance. Lemon essential oil provides a fresh and soft top note, lemon extract is full-bodied and rounded, and lemon absolute is long-lasting. The combination of these three elements satisfies the unified characteristics of the top, middle, and base notes, thus contributing to the fragrance of cosmetics. Lemon essential oil, lemon extract, and lemon absolute all contain abundant active ingredients, endowing the composition of this invention with varying degrees of antibacterial, antioxidant, anti-inflammatory, and whitening properties. 1,2-hexanediol, 1,2-pentanediol, and propylene glycol enhance the antibacterial and preservative functions of the lemon peel extract, providing antibacterial and preservative effects for cosmetics.

Description

Technical Field

[0001] This invention relates to the preparation and application of a composition containing Rutaceae fruit peel extract, specifically to a composition containing lemon fruit peel extract, its preparation method and application. This composition containing lemon fruit peel extract not only has fragrance, antibacterial and preservative effects, but also has multiple functions such as anti-oxidation, anti-allergy, whitening and moisturizing, and can be used as a multi-functional raw material for cosmetics. Background Technology

[0002] Currently, most cosmetics use synthetic fragrances. Compared to natural fragrances, synthetic fragrances lack the natural, soft, and mellow aroma, and most synthetic fragrances lack functional benefits. Most cosmetics require preservatives to maintain their shelf life. Developing plant extracts that possess natural fragrances, preservative properties, and certain functional benefits for use in cosmetics aligns with the current trend towards natural cosmetics and is more likely to be favored by consumers.

[0003] Lemon is the third largest variety of plant in the genus Citrus of the Rutaceae family, and a well-known fruit used for both medicinal and culinary purposes. Its consumption and processing generate a large amount of byproducts such as lemon peel. Modern pharmacological experiments have confirmed that lemon peel has pharmacological effects such as antibacterial and anti-inflammatory properties, as well as anti-asthmatic, expectorant, antitussive, analgesic, and promotion of gastrointestinal motility and digestive juice secretion. Lemon peel is rich in volatile terpenes, alkaloids, phenolic acids, and other active ingredients. We have found that these components not only have a captivating fragrance but also antibacterial properties and unique biological activities, which can be applied to the field of beauty and skincare products. While replacing synthetic fragrances and providing antibacterial and preservative effects, they also endow cosmetics with special efficacy such as anti-oxidation (preventing skin aging), anti-inflammation (soothing, less allergenic), anti-tyrosinase (whitening), and moisturizing. Therefore, this invention proposes a composition containing lemon peel extract, its preparation method, and its application. Summary of the Invention

[0004] The technical problem to be solved by the present invention is to provide a composition containing lemon peel extract with fragrance, antibacterial and antiseptic, antioxidant, anti-allergic, whitening and moisturizing effects, as well as its preparation method and application.

[0005] The technical solution adopted by the present invention to solve the above-mentioned technical problem is as follows: a composition containing lemon peel extract, the composition being composed of lemon peel extract, 1,2-hexanediol, 1,2-pentanediol, propylene glycol, and polyoxyethylene hydrogenated castor oil in a mass ratio of (20-60):(10-20):(10-20):(10-20):(10-20), wherein the lemon peel extract is composed of lemon essential oil, lemon extract, and lemon absolute in a mass ratio of (20-60):(20-50):(20-30).

[0006] The composition of this invention containing lemon peel extract has a naturally soft, full-bodied, and long-lasting fragrance. Lemon essential oil has a fresh and soft top note, lemon extract has a full and rounded aroma, and lemon absolute has a long-lasting fragrance. The combination of these three elements satisfies the characteristics of a unified fragrance profile across the top, middle, and base notes. This lemon peel extract not only has the function of adding fragrance to cosmetics but also possesses antibacterial, antioxidant, anti-allergic, whitening, and moisturizing effects.

[0007] The lemon essential oil, lemon extract, and lemon absolute contained in the composition of this invention all have antibacterial and preservative effects. When combined with 1,2-hexanediol, 1,2-pentanediol, and propylene glycol in a certain proportion, the antibacterial and preservative effects are enhanced, thus providing a preservative effect for cosmetics.

[0008] The addition of polyoxyethylene hydrogenated castor oil to the composition of this invention helps to mix the components evenly, resulting in a product with a uniform appearance without layering, and making it easier to be compatible with other cosmetic ingredients.

[0009] Our tests revealed that:

[0010] 1) Lemon essential oil contains seven main compounds (relative percentage >1%), accounting for 94.66% of the total peak area, including D-limonene, β-pinene, and γ-terpinene. The most abundant component is D-limonene (58.05%), which has good antibacterial, antitussive, and expectorant effects. It can be used as a raw material for formulating artificial orange blossom, sweet blossom, lemon, and bergamot oils, and can also be used as a fresh top note in cosmetic fragrances. β-pinene (14.28%) is an important raw material for the fragrance industry and the synthesis of other fine chemicals. It has broad-spectrum antifungal activity, inhibiting the growth of many fungi, and can be used for astringent skin treatment, antidepressant, anti-cold, relieving swelling and pain, and has certain anti-cancer effects. γ-terpinene (11.98%) is a monoterpene, an orally effective antioxidant that can directly scavenge free radicals. It can be used to relieve pain and also has antifungal and antitrypanosome properties.

[0011] 2) There are 16 main compounds (relative percentage >1%) in lemon extract, accounting for 92.96% of the total peak area, including D-limonene, β-bisabolene, and α-bergamotene. D-limonene has the highest content (22.95%). β-bisabolene (19.39%) is a component with both fragrance and health benefits, possessing woody, citrus, floral, fruity, green, and sweet balsamic aromas, and is widely used in fragrance formulations for daily cosmetics; it is also a good antioxidant with excellent anti-itch and anti-inflammatory properties. α-bergamotene (11.82%), also known as bergamot lactone, is found in high concentrations in bergamot, star anise, and toon. Studies have shown that α-bergamotene is a natural and effective anti-tumor drug, and it is frequently consumed and used as both a food flavoring ingredient and a natural medicinal component.

[0012] 3) Seven main compounds (relative percentage >1%) were found in lemon absolute, accounting for 98.47% of the total peak area, including D-limonene, γ-terpinene, and β-pinene. Among them, D-limonene accounted for 63.79%, γ-terpinene for 18.82%, and β-pinene for 5.13%.

[0013] 4) Lemon essential oil, lemon extract, and lemon absolute contain seven common components: β-pinene, myrcene, D-limonene, γ-terpinene, citral, linalool, and α-terpineol. Citral, linalool, and α-terpineol can all be used as fragrance agents in the formulation of lemon flavorings, and they also possess antibacterial, antioxidant, and anti-inflammatory activities.

[0014] 5) Terpenes were the most abundant component in all three lemon peel extracts (81.96%–96.64%). Alcohols, aldehydes, and esters were also major sources of aroma components in lemon peel extracts. The relative contents of oxygenated compounds in the three lemon peel extracts were as follows: lemon extract (17.8%) > lemon absolute (6.99%) > lemon essential oil (3.36%).

[0015] The preparation method of the above-mentioned composition containing lemon peel extract involves preparing lemon essential oil by enzymatic hydrolysis and steam distillation, preparing lemon extract by ethanol extraction, and preparing lemon absolute oil by petroleum ether extraction and alcohol precipitation.

[0016] Specifically, the preparation process of lemon essential oil is as follows: Weigh 100g of lemon peel into a round-bottom flask, add 1000-3000mL of distilled water, and then add 1-3% sodium chloride and 1-2% cellulase mixture (by weight of lemon peel). This cellulase mixture consists of cellulase with an activity ≥10,000 U / mg and hemicellulase with an activity ≥10,000 U / mg in a 2:1 mass ratio. Adjust the pH to 5-5.5 with citric acid, heat to 40-55℃, and enzymatically hydrolyze for 60-120 minutes. Then, increase the temperature to a gentle reflux extraction until no oil droplets remain in the distillate. After separation with an oil-water separator, collect the oily, transparent liquid, dry with anhydrous sodium sulfate, store in a sealed brown bottle, and refrigerate at 4℃ for later use. The prepared lemon essential oil is a colorless, transparent liquid that retains a natural, soft, and fresh lemon aroma and is highly volatile.

[0017] Specifically, the preparation process of lemon extract is as follows: Weigh 100g of lemon peel, add 2-6% (by weight of lemon peel) of propylene glycol and 300-600mL of 90% ethanol, and extract 2-3 times at 40-60℃ for 1-2 hours. Combine the extracts, let stand, concentrate the supernatant under reduced pressure to remove ethanol, and store the paste in a sealed brown bottle at 4℃ for later use. The prepared lemon extract is a yellowish-brown semi-paste, retaining the unique aroma of lemon, but with a fuller and more rounded flavor.

[0018] Specifically, the preparation process of lemon absolute is as follows: Weigh 100g of lemon peel, add 300-600mL of petroleum ether, and reflux extract 2-3 times at 40-60℃ for 1-2 hours. Combine the extracts, let stand, and concentrate the supernatant under reduced pressure to a paste. Then mix it with anhydrous ethanol at a volume ratio of 1:(5-10), and let it stand overnight at -20℃. Take the supernatant, concentrate it under reduced pressure to remove the solvent, and store it in a sealed brown bottle at 4℃ for later use. The prepared lemon absolute is an orange-yellow liquid with a longer-lasting and more persistent aroma than lemon essential oil, and its fragrance and color are superior to lemon extract.

[0019] Specifically, the preparation process of the composition containing lemon peel extract is as follows: weigh lemon essential oil, lemon extract, and lemon absolute oil and mix them in a mass ratio of (20-60):(20-50):(20-30) to obtain lemon peel extract; take 20-60 parts of lemon peel extract and 10-20 parts of polyoxyethylene hydrogenated castor oil, stir them thoroughly, and then add 10-20 parts of 1,2-hexanediol, 10-20 parts of 1,2-pentanediol and 10-20 parts of propylene glycol and stir them thoroughly.

[0020] The above-mentioned composition containing lemon peel extract is used in cosmetics as a fragrance agent, preservative, antibacterial agent, antioxidant, anti-allergy agent (skin soothing agent), anti-tyrosinase agent (whitening agent), and moisturizer.

[0021] Compared with the prior art, the present invention has the following advantages:

[0022] ① This invention contains lemon peel extract, which is formulated with lemon essential oil, lemon extract, and lemon absolute in a specific ratio. The resulting fragrance is naturally soft, smooth, and long-lasting. Lemon essential oil is fresh and gentle, lemon extract is full-bodied and smooth, and lemon absolute is long-lasting. The combination of these three ingredients satisfies the unified characteristics of the top, middle, and base notes of a fragrance, thus providing a scenting effect for cosmetics. These three lemon peel extracts—lemon essential oil, lemon extract, and lemon absolute—all contain abundant active ingredients, giving the composition varying degrees of antibacterial, antioxidant, anti-inflammatory, and whitening properties.

[0023] ② The addition of 1,2-hexanediol, 1,2-pentanediol and propylene glycol to the composition containing lemon peel extract in this invention enhances the antibacterial and preservative function of lemon peel extract, thus providing antibacterial and preservative effects for cosmetics.

[0024] ③ The addition of polyoxyethylene hydrogenated castor oil to the composition of the present invention helps to improve the compatibility and uniform dispersion of the composition with other ingredients in cosmetics.

[0025] ④ The composition containing lemon peel extract of the present invention can be used as a fragrance agent, preservative, antibacterial agent, antioxidant, anti-allergic agent, anti-tyrosinase agent and moisturizer in cosmetics such as skin care water, skin care lotion and skin care cream. Attached Figure Description

[0026] Figure 1 The above are the total ion current chromatograms of the three lemon peel extracts in the examples;

[0027] Figure 2 The electronic nose response values ​​of the three lemon peel extracts in the examples are shown.

[0028] Figure 3 The response curves show the maximum odor response values ​​of the three lemon peel extracts in the examples. Detailed Implementation

[0029] The present invention will be further described in detail below with reference to the accompanying drawings and embodiments.

[0030] I. The lemon peel extract in the composition containing lemon peel extract of the present invention is composed of lemon essential oil, lemon extract and lemon absolute in a mass ratio of (20-60):(20-50):(20-30).

[0031] The following are the preparation and evaluation test results of three lemon peel extracts: lemon essential oil, lemon extract, and lemon absolute:

[0032] (1) Preparation and sensory evaluation of three lemon peel extracts

[0033] Wash fresh lemon peels, remove the white inner layer, dry at 40℃ for 1-3 hours, then pulverize and pass through an 8-16 mesh sieve. Seal in a plastic bag and store for later use.

[0034] Preparation of lemon essential oil: Weigh 100g of lemon peel into a round-bottom flask, add 1000-3000mL of distilled water, and then add 1-3% sodium chloride and 1-2% cellulase mixture (by weight of lemon peel). This cellulase mixture consists of cellulase with an activity ≥10,000 U / mg and hemicellulase with an activity ≥10,000 U / mg in a 2:1 mass ratio. Then, adjust the pH to 5-5.5 with citric acid, heat to 40-55℃, and enzymatically hydrolyze for 60-120min. Then, raise the temperature to maintain a gentle boil and reflux extraction until no oil droplets are found in the distillate. After separation with an oil-water separator, collect the oily transparent liquid, dry it with anhydrous sodium sulfate, store it in a sealed brown bottle, and refrigerate at 4℃ for later use.

[0035] Preparation of lemon extract: Weigh 100g of lemon peel, add 2-6% of the lemon peel weight of propylene glycol and 300-600mL of 90% ethanol, and extract 2-3 times at 40-60℃ for 1-2h. Combine the extracts, let stand, concentrate the supernatant under reduced pressure to remove ethanol, and store the yellow-brown paste in a sealed brown bottle at 4℃ for later use.

[0036] Preparation of lemon absolute: Weigh 100g of lemon peel, add 300-600mL of petroleum ether, and reflux extract 2-3 times at 40-60℃ for 1-2h. Combine the extracts, let stand, and concentrate the supernatant under reduced pressure to a paste. Then mix it with anhydrous ethanol at a volume ratio of 1:(5-10) and let it stand overnight at -20℃. Take the supernatant, concentrate it under reduced pressure to remove the solvent, and store the orange-yellow liquid in a brown bottle in a sealed container at 4℃ for later use.

[0037] The sensory quality comparison of the three lemon peel extracts is shown in Table 1.

[0038] Table 1. Comparison of sensory qualities of three lemon peel extracts

[0039]

[0040] Lemon essential oil is a colorless and transparent liquid that retains a fresh lemon aroma and is highly volatile; lemon extract is a yellowish-brown semi-paste that retains the natural aroma of lemon more completely; lemon absolute is an orange-yellow liquid with a more lasting and richer scent than lemon essential oil, and its aroma and color are superior to lemon extract.

[0041] (2) GC / MS test of the components contained in the three lemon peel extracts

[0042] GC conditions: The capillary column was an HP-5MS flexible quartz capillary column (30m × 0.25mm × 0.25μm); the carrier gas was helium with a purity > 99.999%; manual injection was used with a split ratio of 200:1; the injection port temperature was 220℃; the temperature program was as follows: initial column temperature 50℃ held for 3 min, increased to 110℃ at 2℃ / min and held for 10 min, increased to 140℃ at 2℃ / min and held for 2 min, increased to 250℃ at 10℃ / min and held for 5 min (total run time 84 min).

[0043] Mass spectrometry conditions: ionization mode was electron impact ionization (EI); electron energy was 70 eV; transfer line temperature was 270 °C; ion source temperature was 230 °C; solvent delay was 2 min; scan range was 35–550 u; scan mode was full scan.

[0044] The NIST MS spectral library was used to analyze the spectra, identify the names of each volatile substance, and the area normalization method was used to calculate the relative content of each component.

[0045] The total ion chromatograms of the three lemon peel extracts are shown below. Figure 1 A total of 22 chromatographic peaks were detected in lemon essential oil, 39 chromatographic peaks were detected in lemon extract, and 9 chromatographic peaks were detected in lemon absolute.

[0046] The chemical composition and relative percentage content of the three lemon peel extracts are shown in Table 2.

[0047] Table 2. Chemical composition and relative percentage content of three lemon peel extracts

[0048]

[0049]

[0050] Note: "-" indicates that the content is <0.05 or not detected.

[0051] Twenty-two components were detected in lemon essential oil, with seven compounds having a relative percentage content >1%, accounting for 94.66% of the total peak area. The most abundant component was D-limonene (58.05%), which possesses excellent antibacterial, antitussive, expectorant, nerve-relieving, analgesic, and detoxifying properties. It can be used as a raw material for formulating artificial orange blossom, sweet blossom, lemon, and bergamot oils, and can also be used as a fresh top note in cosmetic fragrances. β-L-pinene (14.28%) has a resinous and pine resinous aroma and is an important raw material for the fragrance industry and the synthesis of other fine chemicals. It has broad-spectrum antifungal activity, inhibiting the growth of many fungi, and can be used for astringent skin treatment, antidepressant, anti-cold, relieving swelling and pain, and also has certain anti-cancer effects. γ-terpinene (11.98%) is a monoterpene with citrus and lemon aromas and can be used to formulate artificial lemon and peppermint essential oils; γ-terpinene is also an orally effective antioxidant that can directly scavenge free radicals and can be used to relieve pain, and has antifungal and antitrypanosomiasis properties.

[0052] Thirty-nine components were detected in lemon extract, with 16 compounds having a relative percentage content >1%, accounting for 92.96% of the total peak area. The most abundant was D-limonene at 22.95%. β-Bisabolene (19.39%) is a component with both fragrance and health benefits, possessing woody, citrus, floral, fruity, green, and sweet balsamic aromas. It is widely used in fragrance formulations for daily cosmetics and is also a good antioxidant with excellent anti-itch and anti-inflammatory properties. α-Berberinene (11.82%), also known as bergamot lactone, is found in high concentrations in bergamot, star anise, and toon. Studies have shown that bergamotene is a natural and effective anti-tumor drug, and it is frequently consumed and used as both a food flavoring ingredient and a natural medicinal component.

[0053] Seven compounds with a relative percentage content >1% were found in lemon absolute oil, accounting for 98.47% of the total peak area. Among them, D-limonene accounted for 63.79%, γ-terpinene accounted for 18.82%, and β-pinene accounted for 5.13%.

[0054] The component categories and proportions of the three lemon peel extracts are shown in Table 3.

[0055] Table 3. Component categories and percentages (%) of the three lemon peel extracts

[0056]

[0057]

[0058] Among the three lemon peel extracts, the component with the highest proportion is terpene compounds (81.96% - 96.64%). Oxygenated compounds such as alcohols, aldehydes, and esters are also the main sources of the aroma components of lemon peel extracts. The relative contents of oxygenated compounds in the three lemon peel extracts are as follows: lemon extract (17.64%) > lemon absolute (6.99%) > lemon essential oil (3.36%).

[0059] There are 7 common components in lemon essential oil, lemon extract, and lemon absolute, namely β - (-)-pinene, myrcene, D - limonene, γ - terpinene, citral, linalool, and α - terpineol. Citral, linalool, and α - terpineol can all be used as flavoring agents, to formulate lemon essence and manufacture citrus spices, etc., and also have strong antibacterial, antioxidant, and anti - inflammatory activities. The components of lemon peel extracts obtained by different extraction processes have certain similarities and differences, which result in the coordination and complementarity of the three lemon peel extracts in terms of fragrance, function, etc.

[0060] The content comparison of total flavonoids and total polyphenols in the three lemon peel extracts is shown in Table 4. Flavonoids and polyphenols have multiple effects. They are very strong antioxidants, can effectively scavenge free radicals in the body, prevent cell degradation and aging, can inhibit the exudation of inflammatory bio - enzymes, promote wound healing and relieve pain, and can be used for various types of sensitivity.

[0061] Table 4 Content comparison of total flavonoids and total polyphenols in three lemon peel extracts ( n = 3)

[0062]

[0063] (3) Electronic nose test analysis of the odors of three lemon peel extracts

[0064] Precisely pipette 2 mL of the three lemon peel extracts into a 10 - mL headspace vial and equilibrate for 1 min. Using the headspace aspiration method, at room temperature, detect with a PEN3 - type electronic nose. Detection conditions: the self - cleaning time of the sensor is 150 s, the zeroing time of the sensor is 5 s, the sample preparation time is 5 s, and the signal acquisition time is 300 s. Analyze the data using the WinMuster software supporting the PEN3 - type sensor. The changes in the electronic nose response values of the three lemon peel extracts are shown in Figure 2 .

[0065] From Figure 2It can be seen that the ten sensing elements all showed different response intensities to the sample odor, with the response values ​​mainly concentrated on probes S2, S7, and S9. The S7 response value was highest for essential oils and absolute oils, while the S2 response value was highest for extracts. Tables 2-3 show that terpenoids were the main components in all three lemon peel extracts, accounting for 96.64%, 81.96%, and 93.01% in essential oils, extracts, and absolute oils, respectively. Therefore, the S7 response values ​​for essential oils and absolute oils were relatively high. The extract was the only one containing isothiocyanates, although its relative content was only 0.22%, its S2 response value was the highest. This is because phenyl isothiocyanate has a strong, pungent odor, which can be captured by the S2 probe even at extremely low concentrations.

[0066] Plot its response curve based on the maximum response value. Figure 3 The peak values ​​of the odors of three lemon peel extract samples were identified, and the changes in their response values ​​after reaching the peak value for 100 seconds were measured, as shown in Table 5.

[0067] Table 5. Peak odor response values ​​and their variations for three lemon peel extracts.

[0068]

[0069] comprehensive Figure 3 According to Table 5, based on the maximum response peak value, the aroma intensity was: lemon essential oil (43.4134) > lemon extract (20.3461) > lemon absolute (16.6896). This is because extracts and absolutes contain plant waxes and pigments in addition to terpenes and aromatic components, and their high viscosity makes it difficult for aroma components to overcome the barrier, which is the main reason for the overall low response value. The fragrance retention was determined by the difference between the peak value and the corresponding G / G0 value within the same time period after the peak. Three time periods were selected: 60s, 100s, and 140s after the peak. The fragrance persistence was: lemon absolute > lemon essential oil > lemon extract. This indicates that differences in the types and contents of components in lemon peel extracts obtained by different extraction methods affect the intensity, fullness, and persistence of the aroma.

[0070] Therefore, it can be seen that the aroma of lemon essential oil is mainly concentrated in the initial stage and is slightly less rich; the aroma of lemon extract is full and mellow, with the scent appearing in the middle and later stages; the aroma of lemon absolute is not very strong but has good persistence. Lemon extract can compensate for and connect the middle aromas of essential oil and absolute, and the combination of the three creates a harmonious and layered aroma that can also enhance the persistence of the fragrance.

[0071] (4) Determination of the in vitro antioxidant activity of three lemon peel extracts

[0072] DPPH free radicals: Add 1 mL of sample solution of different concentrations, 3 mL of 1 mmol / L DPPH solution, and 1 mL of deionized water to a test tube to a total volume of 5 mL as the test group. Shake well and react at room temperature in the dark for 30 min. Zero the instrument with the sample solution solvent and measure the absorbance A1 at 517 nm. Use the sample solution solvent instead of the sample solution as the blank group and measure A0. Use anhydrous ethanol instead of DPPH solution as the background group and measure A2. Use vitamin C as a positive control. Calculate according to the formula (1) below. The results are shown in Table 6.

[0073] ABTS free radicals: Add 1 mL of sample solution of different concentrations and 3 mL of ABTS working solution (A) to the test tube in sequence. 734nm 0.7±0.02) and 1 mL of deionized water were added to a total volume of 5 mL as the test group. After thorough mixing, the mixture was reacted at room temperature in the dark for 30 min. The absorbance A1 at 734 nm was measured using the sample solvent as the zeroing point. A0 was measured using the sample solvent instead of the sample solution as the blank group. A2 was measured using anhydrous ethanol instead of the ABTS solution as the background group. Vitamin C was used as the positive control. The results are shown in Table 7, calculated according to formula (1) below.

[0074]

[0075] Table 6. Scavenging rates and IC50 values ​​of three lemon peel extracts against DPPH free radicals. 50 ( n=3)

[0076]

[0077] As shown in Table 6, the DPPH free radical scavenging ability of the three lemon peel extracts increased with increasing concentration. The DPPH free radical scavenging rate was: lemon extract > lemon absolute > lemon essential oil. The lemon extract showed the best effect, with a DPPH scavenging rate as high as 92.44±1.99% at 8 mg / mL.

[0078] Table 7. Scavenging rates and IC50 values ​​of three lemon peel extracts against ABTS free radicals. 50 ( n=3)

[0079]

[0080] As shown in Table 7, within the tested concentration range, the scavenging ability of the three lemon peel extracts against ABTS free radicals increased with increasing concentration. The scavenging rate against ABTS free radicals was: lemon absolute oil > lemon extract > lemon essential oil. Lemon absolute oil at 0.7 mg / mL achieved a scavenging rate of 91.80 ± 2.46% against ABTS free radicals.

[0081] The three lemon peel extracts showed varying effects on free radical scavenging, which was related to the content of total flavonoids and total polyphenols in the extracts. Table 5 shows that lemon extract and absolute oil contained more total flavonoids and total polyphenols than lemon essential oil. Flavonoids and polyphenols have good anti-free radical effects, and the scavenging patterns of these two free radicals generally corresponded to the variation patterns of total flavonoid and total polyphenol content in the three extracts.

[0082] (5) Determination of the in vitro anti-inflammatory activity of three lemon peel extracts

[0083] Albumin inhibition assay was used for evaluation. 0.4 mL of fresh egg white diluent and 3.1 mL of phosphate buffer (pH 6.8) were added to the test tubes, and after thorough mixing, 1.5 mL of sample solutions of different concentrations were added to a total volume of 5 mL as the test group. The tubes were incubated in a 37℃ water bath for 15 min, then immediately heated in a 70℃ water bath for 10 min, cooled, and the absorbance A1 at 660 nm was measured using deionized water as the zeroing tube. PBS buffer was used instead of the sample solution as the blank group, and A0 was measured. PBS buffer was used instead of the egg white diluent as the background group, and A2 was measured. Ibuprofen (standard drug) was used as a positive control. The results were calculated according to formula (2) and are shown in Table 8.

[0084]

[0085] Table 8. Inhibition rate (%) of three lemon peel extracts on albumin n=3)

[0086]

[0087] Table 8 shows that, within the experimental concentration range of 5 to 200 μg / mL, all three lemon peel extracts and the positive control ibuprofen could protect albumin from heat-induced damage and denaturation to varying degrees in a concentration-dependent manner. Their inhibition rates were: lemon absolute oil > lemon extract > lemon essential oil. Lemon absolute oil (200 μg / mL) showed the best effect, exhibiting an inhibition rate of 72.69 ± 1.13%, comparable to ibuprofen. Previous studies have demonstrated the anti-inflammatory effects of limonene, linalool, and α-pinene, and some studies have also shown that β-bisabolene has anti-inflammatory functions. Among the three, lemon absolute oil exhibited the strongest anti-inflammatory effect.

[0088] (6) Determination of in vitro antityrosinase activity of three lemon peel extracts

[0089] Add the reagents in the order listed in Table 9 from top to bottom, shake thoroughly, and incubate in a 37°C water bath for 10 minutes. Immediately add the tyrosine solution and incubate at 37°C in the dark for 30 minutes. Cool, zero the chamber with deionized water, and determine the A content of each group of samples. 475nm Arbutin was used as a positive control. The inhibition rate of tyrosinase was calculated according to formula (3), and the results are shown in Table 10.

[0090]

[0091] Table 9. Composition of the reaction solution for determining the tyrosinase inhibition rate.

[0092]

[0093] Table 10. Inhibition rates and IC50 values ​​of three lemon peel extracts against tyrosinase. 50 ( n=3)

[0094]

[0095] Table 10 shows that the inhibitory effect of the three lemon peel extracts on tyrosinase activity increased with increasing concentration, and the inhibition rate of tyrosinase was in the order of lemon essential oil > lemon extract > lemon absolute. At 20 mg / mL, lemon essential oil achieved an inhibition rate of 93.82 ± 1.06%, far exceeding the inhibition rate of the positive control arbutin. Furthermore, the IC50 values ​​of the three lemon peel extracts... 50 The values ​​were all lower than the IC50 value of arbutin. 50 The concentration of 29.40 mg / mL indicates that its anti-tyrosinase effect is superior to that of arbutin. Lemon peel is rich in flavonoids, polyphenols, and other bioactive substances. Due to their structural similarity to the substrate of tyrosinase, they can act as substrate analogs to bind to tyrosinase, thereby inhibiting melanin production. Citral, nerol acetate, and pinene are also major contributors to the inhibition of tyrosinase activity. The synergistic effect of different compounds results in the extract having different degrees of inhibitory effect on tyrosinase.

[0096] (7) Antibacterial properties of three lemon peel extracts

[0097] Preparation of test strains: The strains were activated using MH medium. *Escherichia coli* and *Staphylococcus aureus* were cultured at 37°C, and *Candida albicans* at 28°C to the logarithmic growth phase. The bacterial suspension was centrifuged at 5000 rpm for 10 min, and the precipitate was collected. The precipitate was washed twice with 0.9% sterile physiological saline, and the bacteria were resuspended in MH medium and the bacterial concentration was adjusted to 1 × 10⁻⁶. 8 CFU / mL represents the bacterial solution to be tested.

[0098] Sample solution preparation: Accurately weigh 0.100g of each of the three lemon peel extracts and dissolve each in 1mL of DMSO to obtain a sample stock solution with a concentration of 100mg / mL.

[0099] Antibacterial rate determination: The experiment was divided into a blank group, a solvent blank group, and an experimental group. The experimental group included lemon essential oil, extract, and pure oil groups. A sterile 96-well plate was used. For the blank group, 170 μL of MH medium and 30 μL of the bacterial solution to be tested were added; for the solvent blank group, 200 μL of MH medium containing different concentrations of the sample was added; for the experimental group, 170 μL of MH medium containing the sample and 30 μL of the bacterial solution to be tested were added. After incubation for 18 hours, the absorbance was measured at 600 nm using a microplate reader. Each sample was tested in triplicate.

[0100] The antibacterial rate was calculated according to formula (4), and the results are shown in Table 11.

[0101]

[0102] Table 11. Inhibition rates (%) of three lemon peel extracts against Escherichia coli, Staphylococcus aureus, and Candida albicans. n=3)

[0103]

[0104] Comparing the antibacterial properties of the three lemon peel extracts in Table 11, it was found that lemon essential oil had stronger antibacterial activity than lemon extract and lemon absolute at the same concentration. It was more sensitive to *Escherichia coli* than to *Staphylococcus aureus* and *Candida albicans*, with inhibition rates exceeding 90% for all three bacteria at concentrations >12.5 mg / mL. At a concentration of 50 mg / mL, the inhibition rates of the three lemon peel extracts against the three bacteria reached over 93% (except for *Staphylococcus aureus* against lemon extract).

[0105] II. The lemon peel extract contained in the composition of this invention is composed of lemon essential oil, lemon extract, and lemon absolute in a mass ratio of (20-60):(20-50):(20-30). Based on the fact that the three lemon peel extracts each have advantages and differences in aroma, anti-free radical, anti-inflammatory, anti-tyrosinase, and antibacterial properties, specifically:

[0106] (1) Aroma: The aromas of the three have their own characteristics in terms of intensity, fullness and persistence, and they can complement each other;

[0107] (2) DPPH resistance: Lemon extract > Lemon absolute > Lemon essential oil;

[0108] (3) Anti-ABTS ability: Lemon absolute oil > Lemon extract > Lemon essential oil;

[0109] (4) Anti-inflammatory effect: Lemon absolute oil > Lemon extract > Lemon essential oil;

[0110] (5) Anti-tyrosinase properties: lemon essential oil > lemon extract > lemon absolute;

[0111] (6) Antibacterial properties: Lemon essential oil is superior to extract and absolute oil.

[0112] Therefore, this invention prepares a mixture of three lemon peel extracts in a certain proportion to achieve a complementary and enhancing effect.

[0113] Example 1: Weigh 50 parts lemon essential oil, 20 parts lemon extract, and 30 parts lemon absolute oil, mix them evenly to obtain a lemon peel extract. The extract is a yellow liquid with a pure, fresh, full-bodied, and long-lasting lemon aroma. Anti-free radical, anti-tyrosinase, antibacterial, and anti-inflammatory experiments were conducted according to the aforementioned method, and the results are shown in Table 12.

[0114] Table 12 Inhibition rate (%) of lemon peel extract on relevant indicators ( n=3)

[0115]

[0116] It is evident that mixing the three lemon peel extracts in a certain proportion results in a complementary and enhanced effect on their aroma and efficacy, making them more suitable as fragrance and efficacy agents in cosmetics.

[0117] III. The composition containing lemon peel extract of this invention is composed of lemon peel extract, 1,2-hexanediol, 1,2-pentanediol, propylene glycol, and polyoxyethylene hydrogenated castor oil (such as PEG-40 hydrogenated castor oil) in a mass ratio of (20-60):(10-20):(10-20):(10-20):(10-20). The lemon peel extract is composed of lemon essential oil, lemon extract, and lemon absolute in a mass ratio of (20-60):(20-50):(20-30). The composition containing lemon peel extract of this invention can be used as an additive in cosmetics such as skin care water, skin care lotion, and skin care cream, serving as a fragrance agent, preservative, antibacterial agent, antioxidant, anti-allergic agent, whitening agent, moisturizing agent, anti-tyrosinase agent, and skin soothing agent.

[0118] (1) Example 2 and Comparative Example: Formulation and Preparation of Softening Cream

[0119] Table 12 Formulas of Cream Products

[0120]

[0121] Preparation of Example 2: Component A was added sequentially to an emulsifying pot and heated to 80°C with stirring. Component B was mixed and heated to 80°C, then poured into component A. The mixture was kept warm and homogenized with stirring for 15 minutes. Stirring was continued until the temperature dropped to 45-50°C, then component C was added. Stirring was continued for another 15 minutes before discharging.

[0122] Preparation of the comparative example: Component A was added sequentially to an emulsifying pot and heated to 80°C with stirring. Component B and Component D were mixed and heated to 80°C. Component B+D was then poured into Component A, and the mixture was kept warm and homogenized with stirring for 15 minutes. Stirring was continued until the temperature dropped to 45-50°C. Component E was added, mixed thoroughly, and then discharged.

[0123] (2) Example 2 and Comparative Example: The sensory, spreadability, stability and microbiological tests of the cream were compared according to the "Cosmetic Safety Technical Specifications" (2015 edition, Announcement No. 40 of the National Medical Products Administration in 2019). The test results are shown in Table 13 and Table 14.

[0124] Table 13 Sensory indicators and stability of cream products

[0125]

[0126]

[0127] 50g of the cream products from Example 2 and the comparative example were placed in bottles, sealed tightly, and placed in a 40℃ oven for 10 days. Samples were then taken for microbial testing. The results are as follows:

[0128] Table 14 Microbiological test results of cream products

[0129]

Claims

1. A composition containing lemon peel extract, characterized in that, The composition consists of lemon peel extract, 1,2-hexanediol, 1,2-pentanediol, propylene glycol, and polyoxyethylene hydrogenated castor oil in a mass ratio of (20~60):(10~20):(10~20):(10~20):(10~20). The lemon peel extract is composed of lemon essential oil, lemon extract, and lemon absolute in a mass ratio of (20~60):(20~50):(20~30). The lemon absolute is prepared by petroleum ether extraction and alcohol precipitation.

2. The method for preparing the composition containing lemon peel extract according to claim 1, characterized in that, Lemon essential oil was prepared by enzymatic hydrolysis and steam distillation, and lemon extract was prepared by ethanol extraction.

3. The method for preparing the composition containing lemon peel extract according to claim 2, characterized in that, The preparation process of lemon essential oil is as follows: Weigh 100g of lemon peel into a round-bottom flask, add 1000-3000mL of distilled water, and then add 1-3% sodium chloride and 1-2% cellulase mixture by weight of lemon peel. The cellulase mixture consists of cellulase with an enzyme activity ≥10,000 U / mg and hemicellulase with an enzyme activity ≥10,000 U / mg in a 2:1 mass ratio. Then, adjust the pH value to 5-5.5 with citric acid, heat to 40-55℃, and enzymatically hydrolyze for 60-120min. Then, raise the temperature to maintain a gentle reflux extraction until there are no oil droplets in the distillate. After separation with an oil-water separator, collect the oily transparent liquid, dry it with anhydrous sodium sulfate, and store it in a brown bottle with a seal.

4. The method for preparing the composition containing lemon peel extract according to claim 2, characterized in that, The preparation process of lemon extract is as follows: Weigh 100g of lemon peel, add 2-6% of the lemon peel weight of propylene glycol and 300-600mL of 90% ethanol, and extract 2-3 times at 40-60℃ for 1-2 hours. Combine the extracts, let stand, concentrate the supernatant under reduced pressure to remove ethanol, and store the paste in a brown bottle and seal it.

5. The method for preparing the composition containing lemon peel extract according to claim 2, characterized in that, The preparation process of lemon absolute is as follows: Weigh 100g of lemon peel, add 300~600mL of petroleum ether, and reflux extract 2~3 times at 40~60℃ for 1~2h. Combine the extracts, let stand, concentrate the supernatant under reduced pressure to a paste, and then mix it with anhydrous ethanol at a volume ratio of 1:(5~10). Place it at -20℃ overnight, take the supernatant, concentrate under reduced pressure to remove the solvent, and store it in a brown bottle and seal it.

6. The method for preparing the composition containing lemon peel extract according to claim 2, characterized in that, The preparation process of the composition containing lemon peel extract is as follows: weigh lemon essential oil, lemon extract, and lemon absolute oil and mix them in a mass ratio of (20~60):(20~50):(20~30) to obtain lemon peel extract; take 20~60 parts of lemon peel extract and 10~20 parts of polyoxyethylene hydrogenated castor oil, stir them thoroughly, and then add 10~20 parts of 1,2-hexanediol, 10~20 parts of 1,2-pentanediol and 10~20 parts of propylene glycol and stir them thoroughly.

7. The use of the composition containing lemon peel extract as described in claim 1 in cosmetics as a fragrance agent, preservative, antibacterial agent, antioxidant, anti-inflammatory agent, anti-tyrosinase agent, and moisturizer.

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