Selenium-rich brassica campestris extract and application thereof in preparation of medicine for relieving psoriasis symptom

Abstract

The invention belongs to the technical field of medicines, and discloses a selenium-enriched brassica campestris extract and application thereof in preparation of a medicine for relieving psoriasis symptoms. Effective substances of the selenium-rich brassica campestris are extracted by using a specific extraction mode, the extract is further found to be capable of effectively relieving psoriasis symptoms, and when the effective selenium content in the extract is far lower than that of MSC, the extract still has a good curative effect and is safe and reliable. Other substances in the extract can be combined with the selenium element to achieve a synergistic treatment effect. The selenium-enriched brassica campestris is used as the unique raw material, so that the skin atrophy caused by hormone treatment and the infection risk caused by monoclonal antibody drugs in the traditional psoriasis therapy are thoroughly avoided.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to selenium-enriched rapeseed extract and its application in the preparation of drugs to relieve psoriasis symptoms. Background Technology

[0002] Psoriasis (commonly known as "cowhide rash") is a chronic inflammatory skin disease characterized by scaly erythematous plaques. It commonly affects young adults and shows no significant gender difference. Epidemiological statistics show that there are over 60 million patients worldwide. Its course is characterized by being "difficult to cure, prone to relapse, and often accompanied by comorbidities." More than one-third of patients develop systemic complications such as depression, joint problems, and metabolic abnormalities. This high prevalence and complex pathological process not only cause long-term physical and psychological suffering for patients but also place a continuous burden on family finances and public health systems, thus earning it a place in the category of major intractable diseases by the World Health Organization (WHO).

[0003] Kadry and Rashed used plasma and tissue samples from 20 psoriasis patients and 10 healthy controls, finding that plasma selenium levels in psoriasis patients were lower than in the control group, and concluded that low plasma selenium levels are a predictive factor for the development of psoriasis. Studies by Kharaeva et al. have shown that routine treatment combined with supplementation with vitamin E, coenzyme Q10, and selenium can improve clinical symptoms in patients with severe psoriasis. Supplementation with inorganic forms of selenium (sodium selenite and selenate) has also been reported to improve clinical symptoms in psoriasis patients.

[0004] The above studies indicate that plasma selenium levels are closely associated with psoriasis, and that selenium supplementation can partially alleviate symptoms, suggesting that selenium may play a potential role in regulating the inflammatory pathways of psoriasis.

[0005] Selenium-enriched rapeseed is a high-selenium vegetable hybrid created by Academician Wang Hanzhong's team through multiple generations of breeding. When planted in non-selenium-rich soils across the country, the selenium content in the rapeseed shoots can reach 0.01-0.07 mg·kg⁻¹. -1 It is also rich in nutrients such as calcium, vitamin C, amino acids, and zinc, and is characterized by its tender texture and comprehensive nutrition. Its core advantage lies in the specific accumulation of methylselenocysteine—compared to inorganic selenium, this organic form of selenium has significantly lower toxicity and enters cells through amino acid transport mechanisms (rather than passive diffusion), resulting in a longer retention time in the body and reduced selenium loss. Since methionine-tRNA cannot distinguish between methionine and L-selenomethionine, L-selenomethionine can largely substitute for methionine in protein synthesis when methionine intake is insufficient. Therefore, methylselenocysteine ​​has higher biocompatibility and great potential in the development of functional foods.

[0006] However, current research has not yet evaluated the intervention effects of selenium-enriched rape, a novel organic selenium source with high bioavailability, on psoriasis. Although selenium-enriched rape has the advantages of high bioactivity and low toxicity, whether it can improve psoriasis symptoms still needs further verification. Summary of the Invention

[0007] The purpose of this invention is to provide a method for preparing selenium-enriched rapeseed extract, which is simple, easy to implement, and suitable for industrial production.

[0008] A final objective of this invention is to provide the use of the above-mentioned extract in the preparation of a medicament for relieving psoriasis symptoms. To achieve the above objectives, this invention employs the following technical measures:

[0009] A method for preparing selenium-enriched rapeseed extract includes impregnating selenium-enriched rapeseed powder with DMEM, followed by ultrasonic disruption, high-speed centrifugation, and then filtering and sterilizing the supernatant.

[0010] The method described above, preferably, refers to Zhongyou Gaoxi No. 1, Zhongyou Gaoxi No. 2, Xiziyuan No. 1, or / and Xiziyuan No. 2.

[0011] The preferred embodiment of the above method is as follows: the mixed system formed by the dry powder impregnated with DMEM is placed under ice bath conditions, and ultrasonic extraction is performed for 30-50 minutes. The ultrasonic mode is set to 3-7 seconds working / 3-7 seconds intermittent, and the output power is 15-35%.

[0012] The preferred method described above involves high-speed centrifugation at 8000-12000 r / min for 5-15 min at low temperature.

[0013] The method described above preferably employs aseptic filtration using a 0.22μm PES membrane for filtration and sterilization.

[0014] In the method described above, preferably, after ultrasonic disruption, the extract is left to stand at 4°C in the dark for 1-3 hours.

[0015] Preferably, the extract obtained after filtration and sterilization is aliquoted and stored at -80°C.

[0016] The scope of protection of this invention also includes:

[0017] Selenium-enriched rapeseed extract prepared using the above method;

[0018] Application of selenium-enriched rapeseed extract in the preparation of drugs to relieve psoriasis symptoms.

[0019] In the above-described applications, preferably, the drug includes selenium-enriched rapeseed extract and petrolatum.

[0020] Compared with the prior art, the present invention has the following advantages:

[0021] 1) This invention uses a specific extraction method to extract the effective substances from selenium-enriched rapeseed. During the extraction process, the DMEM medium provides a physiological buffer pH of 7.2-7.4 to maintain the ionization balance of methylselenocysteine. Low temperature conditions are maintained throughout the process to ensure that seleno-methylselenocysteine ​​is not oxidized or volatilized during the extraction process. Ultrasonic treatment is used to efficiently break down the cell walls and fully release seleno-methylselenocysteine ​​from the tissue. A 0.22μm PES filter membrane is used for sterile filtration to thoroughly remove bacteria and fungi from the extract.

[0022] 2) This invention further discovered that the extract can effectively alleviate the symptoms of psoriasis. Even when the effective selenium content in the extract is much lower than that of MSC, it still has a good therapeutic effect, indicating that other substances in the extract can combine with selenium to achieve a synergistic therapeutic effect.

[0023] 3) This invention uses selenium-rich rapeseed as the sole raw material, completely avoiding the skin atrophy associated with hormone therapy and the infection risks of monoclonal antibody drugs in traditional psoriasis treatments. Attached Figure Description

[0024] Figure 1 Flowchart of the preparation process for selenium-enriched rapeseed extract.

[0025] Figure 2 A schematic diagram showing the drug administration time for mouse modeling.

[0026] Figure 3 The images show skin lesions and PASI scores for each group of mice.

[0027] Figure 4 Images of HE and Ki67 staining of mouse skin tissue from each group. Detailed Implementation

[0028] To make the technical problems, technical solutions, and advantages of this invention clearer, a detailed description will be provided below in conjunction with the accompanying drawings and specific embodiments. Unless otherwise specified, the technical solutions described in this invention are conventional solutions in the art, and the reagents or materials described, unless otherwise specified, are all from commercial sources.

[0029] Example 1:

[0030] Preparation of selenium-enriched rapeseed extract

[0031] (1) Accurately weigh 1.00 g of freeze-dried and ground sample powder of selenium-enriched rapeseed (Selenium No. 2) into a 50 mL sterile centrifuge tube; (2) Add 20 mL of serum-free DMEM cell culture medium and vortex mix until the solid and liquid are completely infiltrated;

[0032] (3) The mixture was placed in an ice bath and extracted by ultrasonic disruption for 40 min. The ultrasonic mode was set to 5 s working / 5 s intermittent and the output power was 25%.

[0033] (4) The extract was allowed to stand at 4°C in the dark for 2 hours;

[0034] (5) Centrifuge at 10000r / min for 10min at low temperature, take the supernatant and filter it aseptically through a 0.22μm PES filter membrane to obtain the extract. After the above extract is dispensed, it is stored at -80℃ to obtain the selenium-enriched rapeseed extract (SeRFSE), which is used in the following examples.

[0035] (6) The total selenium content and selenium speciation in the extract were accurately determined by inductively coupled plasma mass spectrometry (ICP-MS). Figure 1 ).

[0036] Analysis showed that 1g of lyophilized powder could yield 18-19ml of extract, which contained 1931.36±218.47ug / L of methylselenocysteine, 117.72±7.12ug / L of selenocysteine, 17.63±7.60ug / L of selenomethionine, and SeO3. 2- 41.41±11.69ug / L, SeO4 2- 6.57±2.14ug / L.

[0037] Example 2:

[0038] Application of selenium-enriched rapeseed extract (SeRFSE) in the preparation of drugs to relieve psoriasis symptoms:

[0039] 1) Pretreatment methods for laboratory animals:

[0040] like Figure 2 As shown: Healthy male BALB / c mice aged 6-8 weeks (weighing 20±2g) were selected and housed in an SPF-grade environment for 7 days of acclimatization (temperature 22±1℃, 12-hour day-night cycle, free access to food and water). The back hair of the mice was removed using a motorized small animal shaver until no residue remained. The mice were restrained during the procedure to prevent skin abrasion. A non-irritating depilatory cream was evenly applied to the shaved area and left for 10 minutes. The cream and loose hair were then gently removed with a sterile cotton swab. The skin of the shaved area was cleaned with saline solution and allowed to air dry before observing the integrity of the epidermis (individuals with broken skin were excluded).

[0041] 2) Grouping of experimental animals:

[0042] Normal control group: 5 animals;

[0043] IMQ only manufactures 5 modules;

[0044] IMQ modeling followed by topical application of MSCs (methylselenocysteine): 5 mice;

[0045] IMQ modeling followed by topical application of SerFSE: 5 animals;

[0046] 3) Methods for handling laboratory animals:

[0047] (1). Normal control group

[0048] Drug substance: Vaseline

[0049] Administration method: Apply petroleum jelly evenly to the shaved area daily;

[0050] Dosage standard: 10g / kg;

[0051] Intervention process:

[0052] Apply 10g / kg of petroleum jelly every afternoon at 16:00;

[0053] Intervene once daily for 5 days.

[0054] (2). IMQ Modeling Group

[0055] Modeling drug: 5% imiquimod (IMQ) cream (chemical purity ≥98%)

[0056] Administration method: Apply a measured amount of IMQ cream to the hairless area of ​​the back. Weigh the IMQ cream onto pre-sterilized weighing paper, accurately dispense it into sterile EP tubes, and spread it evenly.

[0057] Dosage standard: 3.125g (imiquimod cream) / kg (mice);

[0058] Intervention process:

[0059] Apply 3.125g / kg IMQ cream daily at 10:00 AM;

[0060] Apply 10g / kg of petroleum jelly every afternoon at 16:00;

[0061] Intervene twice daily for 5 days.

[0062] 3. MSC group

[0063] Drug delivery substance: Methylselenocysteine ​​(MSC)

[0064] Administration method: MSCs are mixed with petrolatum and applied topically to the IMQ-treated area.

[0065] Dosage standard: 50 μg (MSC) / kg (mice)

[0066] Intervention process:

[0067] Apply 3.125g / kg IMQ cream daily at 10:00 AM;

[0068] Apply 50 μg / kg MSC daily at 16:00.

[0069] Intervene twice daily for 5 days.

[0070] 4. SerFSE group

[0071] Drug agent: Selenium-enriched rapeseed extract (SeRFSE)

[0072] Administration method: Apply SeRFSE mixed with petroleum jelly topically to the IMQ-treated area.

[0073] Dosage standard: 5 ml (SeRFSE) / kg (mice)

[0074] Intervention process:

[0075] Apply 3.125g / kg IMQ cream daily at 10:00 AM;

[0076] Apply 5ml / kg of SeRFSE daily at 16:00.

[0077] Intervene twice daily for 5 days.

[0078] result:

[0079] PASI score:

[0080] Assessment of the severity of back skin inflammation: An objective scoring system was established based on the clinical PASI score. Independent scoring was conducted according to the degree of erythema, scaling, and thickening (0-4 points), with the following scoring rules: 0 = none, 1 = mild, 2 = moderate, 3 = significant, 4 = very significant. The cumulative score (erythema + scaling + thickening, 0-12 points) was used to represent the degree of skin inflammation. All scoring items were completed independently by three researchers in separate sessions.

[0081] The anti-psoriatic effects of MSCs and SerFSE were evaluated using an IMQ-induced psoriatic dermatitis mouse model. Figure 3 As shown, IMQ induced severe psoriatic phenotypes (erythema, scaling, and thickening) on ​​the back skin of mice. Both MSC and SeRFSE treatments improved IMQ-induced psoriasis symptoms, while the skin of mice in the SeRFSE group was smoother and more even. On day 2, mice in all groups had measurable PASI scores. The PASI scores of all groups were highest on day 5 and slightly decreased on day 6. The PASI scores of mice in the MSC and SeRFSE groups were significantly lower than those in the IMQ group, with the decrease in PASI scores being more significant in the SeRFSE group.

[0082] Skin histopathological examination:

[0083] Depend on Figure 4 As shown, compared with the normal control group, the use of IMQ alone significantly thickened the epidermal layer of the mouse dorsal skin to 84.68±3.71 μm. Compared with the IMQ group, the thickness of the mouse epidermal layer was significantly reduced after topical application of MSCs and SerFSE. The thickness of the epidermal layer of the MSC group decreased to 54.55±4.72 μm, while that of the SerFSE group decreased to 41.60±3.15 μm, a more significant reduction compared to the MSC group. Ki67 immunohistochemical staining showed that the Ki67-positive area in the skin of the normal control group was 2.83±0.23%. The use of IMQ alone significantly increased the Ki67-positive area in the mouse dorsal skin to 9.90±1.44%, while the Ki67-positive area in the mouse skin significantly decreased after topical application of MSCs and SerFSE. The area of ​​Ki67-positive skin in MSC group mice decreased to 7.0% ± 1.50%, while the area of ​​Ki67-positive skin in SerFSE group mice decreased to 2.73 ± 0.39%, which was more significant compared to the MSC group.

[0084] Although the MSC and total selenium content in the SeRFSE group (5 ml / kg) were significantly lower than those in the MSC group (50 μg / kg), it still showed significantly better symptom relief in the psoriasis model. This difference may be due to the synergistic effect of other naturally occurring key active ingredients in selenium-enriched rapeseed extract with methyl selenocysteine ​​(MSC) to enhance therapeutic efficacy. Among them, allyl thioglycoside, hydrolyzed by endogenous myrosinase to generate allyl isothiocyanate (AITC), can target cysteine ​​residues of the IKKβ protein in the inflammatory pathway, effectively inhibiting NF-κB activation; the quercetin-3-O-selenocysteine ​​complex, which is present at the same time, binds the benzopyranone skeleton and the active selenium source through stable chemical bonds, achieving the slow-release function of selenium while scavenging hydroxyl free radicals; in addition, zinc ions enriched in plant tissues exist in the form of chelated peptide complexes, which enhance the zinc finger protein-mediated barrier repair process by activating epidermal metallothionein. These three types of substances, from the three dimensions of inflammation regulation, redox homeostasis maintenance, and physical barrier reconstruction, respectively, form a therapeutic network with four synergistic effects with the core components of MSC.

Claims

1. A method for preparing selenium-enriched rapeseed extract, comprising impregnating selenium-enriched rapeseed powder with DMEM, followed by ultrasonic disruption, high-speed centrifugation, and then filtering and sterilizing the supernatant.

2. The preparation method according to claim 1, wherein the selenium-enriched rapeseed is Zhongyou Gaose No. 1, Zhongyou Gaose No. 2, Xiziyuan No. 1, or / and Xiziyuan No.

2.

3. The preparation method according to claim 1, wherein the ultrasonic disruption step is as follows: the mixed system formed by the dry powder impregnated with DMEM is placed under ice bath conditions, and ultrasonic disruption extraction is performed for 30-50 min, with the ultrasonic mode set to working for 3-7 s / intermittent for 3-7 s and the output power being 15-35%.

4. The preparation method according to claim 1, wherein the high-speed centrifugation parameters are 8000-12000 r / min and low-temperature centrifugation for 5-15 min.

5. The preparation method according to claim 1, wherein the filtration and sterilization adopts aseptic filtration using a 0.22 μm PES material filter membrane.

6. According to the preparation method of claim 1, after ultrasonic disruption, the extract is allowed to stand at 4°C in the dark for 1-3 hours.

7. According to the preparation method of claim 1, the extract obtained after filtration and sterilization is aliquoted and stored at -80°C.

8. The selenium-enriched rapeseed extract prepared by the method of claim 1.

9. The use of the selenium-enriched rapeseed extract according to claim 8 in the preparation of a medicament for relieving psoriasis symptoms.

10. The application according to claim 9, wherein the drug comprises selenium-enriched rapeseed extract and petrolatum.

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