CN102302479A - Application of lycopene in preparation of medicament for treating cerebral ischemia injury - Google Patents

Abstract

The invention discloses the application of lycopene in the preparation of medicine for treating cerebral ischemia injury. The invention provides the application of lycopene in the preparation of a drug for treating cerebral ischemic injury; the drug for treating cerebral ischemic injury is at least one of the following 1)-3): 1) a drug for treating cerebral edema; 2) treating Drugs for cerebral infarction; 3) drugs for treating nerve damage. The present invention provides a possible clinically urgently needed drug for acute treatment, that is, a drug with therapeutic and protective effects in the acute cerebral ischemia-reperfusion injury period, and has strong clinical pertinence. The research of the present invention shows that the administration of lycopene during the reperfusion period of acute cerebral ischemia can still exert a good anti-cerebral infarction and neuroprotective effect, so the present invention has real clinical therapeutic significance and great clinical advantages .

Description

Application of Lycopene in Preparation of Drugs for Treating Cerebral Ischemia Injury

technical field

The invention relates to the new application of lycopene in the field of medicine preparation, in particular to the application of lycopene in the preparation of medicine for treating cerebral ischemia injury.

Background technique

Lycopene is a fat-soluble component in edible tomato, its molecular formula is C ₄₀ H ₅₆ , and its molecular weight is 536.85. Its chemical structure contains 11 conjugated double bonds and 2 non-conjugated double bonds. This polyunsaturated aliphatic hydrocarbon structure makes the compound have excellent antioxidant properties and become the most effective capture agent in natural carotene , and its rate constant for quenching singlet oxygen is 2 times higher than that of β-carotene. In 1990, Paolo et al. reported that more than 30 kinds of biological antioxidants, such as carotenoids and tocopherols, quenched singlet oxygen, and lycopene was the strongest for quenching singlet oxygen. In addition, lycopene can eliminate other forms of active oxygen free radicals through chemical reactions, including hydrogen peroxide, nitrite, etc. These oxygen free radicals can cause lipids, proteins, biological enzymes and DNA and other macromolecules in organisms Peroxidation, and this peroxidation has a significant relationship with human aging, cancer, autoimmune diseases and neurodegenerative diseases.

Studies have shown that, in addition to antioxidant properties, lycopene can also inhibit the growth, proliferation and differentiation of a variety of tumor cells by affecting growth factor receptors and intercellular gap junction signal transduction.

Contents of the invention

One object of the present invention is to provide the application of lycopene in the preparation of medicine for treating cerebral ischemic injury.

The invention provides the application of lycopene in the preparation of a drug for treating cerebral ischemic injury; the drug for treating cerebral ischemic injury is at least one of the following 1)-3):

1) Drugs for the treatment of cerebral edema;

2) Drugs for the treatment of cerebral infarction;

3) Drugs for the treatment of nerve damage.

The medicine is as shown in a) or b) below:

a) the drug consists of lycopene, unsaturated phospholipids and dimethyl sulfoxide;

b) The drug consists of lycopene, water and dimethylsulfoxide.

The mass ratio of lycopene, unsaturated phospholipid and dimethyl sulfoxide in a) is 1:1:99～1:1:495;

The mass ratio of lycopene, water and dimethyl sulfoxide in b) is 1:1:49.5˜1:1:198.

The unsaturated phospholipid described in a) is olive oil.

Another object of the present invention is to provide a medicine for treating cerebral ischemic injury.

The active ingredient of the medicine for treating cerebral ischemic injury provided by the invention is lycopene.

The medicine for the treatment of cerebral ischemic injury is at least one of the following 1)-3):

1) Drugs for the treatment of cerebral edema;

2) Drugs for the treatment of cerebral infarction;

3) Drugs for the treatment of nerve damage.

The medicine for the treatment of cerebral ischemic injury is as shown in a) or b) below:

a) the drug consists of lycopene, unsaturated phospholipids and dimethyl sulfoxide;

b) The drug consists of lycopene, water and dimethylsulfoxide.

The mass ratio of lycopene, unsaturated phospholipid and dimethyl sulfoxide in a) is 1:1:99～1:1:495;

The mass ratio of lycopene, water and dimethyl sulfoxide in b) is 1:1:49.5˜1:1:198.

The unsaturated phospholipid described in a) is olive oil.

This invention is a project supported by the Beijing Natural Science Foundation, and the project number is 7092009. The invention provides a new drug with therapeutic and protective effects in the acute cerebral ischemia-reperfusion injury period. The present invention has the following obvious advantages:

(1) The present invention avoids the problems of blindness and long-term medication faced by prophylactic administration. The purpose of preventive medicine is to prevent diseases that may occur in the future. The first problem we face is who needs medicine, that is, which groups of people are likely to have a high incidence of diseases; for these people, diseases may or may not occur in the future. It will inevitably lead to the blindness of drug use; secondly, even for high-risk groups, drug use to prevent possible diseases that do not know when in the future means that long-term drug use is required, and the problem of poor drug compliance will inevitably occur, and the cost of prevention cannot be ignored;

(2) The present invention provides a possible clinically urgently needed drug for the treatment of the acute phase, that is, a drug with therapeutic and protective effects in the acute cerebral ischemia-reperfusion injury phase, which has strong clinical pertinence. In acute cerebral ischemic injury, acute treatment drugs are the most lacking, especially in the case of routine clinical emergency use of vasodilator drugs, reperfusion injury is inevitable. The research of the present invention shows that the administration of lycopene during the reperfusion period of acute cerebral ischemia can still exert a good anti-cerebral infarction and neuroprotective effect, so the present invention has real clinical therapeutic significance and great clinical advantages .

(3) The present invention provides a possible reference dosage range. In the fight against cerebral ischemia-reperfusion injury in rats, lycopene has obvious protective effects against cerebral infarction, cerebral edema and neurobehavior in the dose range of 50-200 mg/kg (twice a day), and has certain clinical application. guiding significance.

Description of drawings

Figure 1 is a photograph of TTC-stained brain slices in the sham operation (blank) control group.

Figure 2 is the photo of TTC-stained brain slices in the model control group.

Fig. 3 is a photograph of TTC-stained brain slices of the low-dose lycopene group (20 mg/kg).

Fig. 4 is a photograph of TTC-stained brain slices of the lycopene middle-dose group (50 mg/kg).

Fig. 5 is a photograph of TTC-stained brain slices of the lycopene high-dose group (100 mg/kg).

Figure 6 is a photograph of TTC-stained brain slices in the Edaravone group.

Detailed ways

The experimental methods used in the following examples are conventional methods unless otherwise specified.

The materials and reagents used in the following examples can be obtained from commercial sources unless otherwise specified.

Embodiment 1, the application of lycopene in the preparation of the medicine for the treatment of cerebral ischemia injury

1. Preparation of drugs for the treatment of cerebral ischemic injury

Lycopene is used as the main drug, and it is emulsified with unsaturated phospholipids to make a suspension, which is used for the acute and rehabilitation treatment of cerebral ischemia-reperfusion injury.

In this example, 0.1 ml of dimethyl sulfoxide (DMSO) was added to 100 mg of lycopene (content > 99%) (purchased from Xi'an Bosheng Biological Co., Ltd., catalog number 13739-02-1) for rapid grinding Emulsify, then gradually add 9.9ml of olive oil, mix well, and ultrasonically suspend for 20 minutes to obtain a 10mg/ml lycopene preparation. In the 10 mg/ml lycopene preparation, the mass ratio of lycopene, unsaturated phospholipid and dimethyl sulfoxide is 1:1:99.

On this basis, it was diluted with olive oil to obtain 5 mg/ml lycopene preparation and 2 mg/ml lycopene preparation, so as to ensure equal volume of olive oil solvent fed to rats under different lycopene doses. In the 5mg/ml lycopene preparation, the mass ratio of lycopene, unsaturated phospholipid and dimethyl sulfoxide is 1:1:198. In the 2mg/ml lycopene preparation, the mass ratio of lycopene, unsaturated phospholipid and di The mass ratio of methyl sulfoxide is 1:1:495.

2. Evaluation of the effect of drugs in the treatment of cerebral ischemic injury

The present invention adopts the middle cerebral artery occlusion model of thread-tethering method to cause acute cerebral ischemia-reperfusion injury model in rats, and adopts acute treatment administration mode (administration during reperfusion), from cerebral infarction area, degree of cerebral edema and animal The three aspects of behavioral score prove the acute therapeutic effect and neurorehabilitation effect of lycopene on acute cerebral ischemia-reperfusion injury.

1. Experimental materials and methods

Animals and groups:

Twenty-four Wistar male rats, weighing 240-260 g, were purchased from Beijing Huafukang Biotechnology Co., Ltd. The experiment was divided into 6 groups with 8 rats in each group: (1) sham operation (blank) control group; (2) model control group; (3) lycopene low dose group (20mg/kg) (i.e. the 2mg/ml lycopene preparation, the dosage is 1.0ml/100g); (4) lycopene medium dose group (50mg/kg) (that is, the 5mg/ml lycopene preparation prepared in the above experiment 1, the dosage is 1.0ml/ 100g); (5) lycopene high-dose group (100mg/kg) (that is, the above-mentioned experiment 1 prepares 10mg/ml lycopene preparation, and the consumption is 1.0ml/100g); (6) Edaravone group (3mg/ml kg) (purchased from Sigma, catalog number M70800). The lycopene group was intragastrically administered immediately after reperfusion and 2 hours after reperfusion, the Edaravone group was administered intravenously at the same time, and the model control group was given the same dose at the same reperfusion time point after modeling The solvent (olive oil containing 1% DMSO), and the blank control group were sham-operated animals without thread plugs inserted.

Preparation of cerebral ischemia-reperfusion model:

According to the method of Zea Longa et al., male Wistar rats were anesthetized with 3% pentobarbital sodium 1ml·kg ^-1 , fixed in the supine position, cut along the median of the neck, and separated and exposed the left common carotid artery (CCA) and internal carotid ( ICA), external carotid artery (ECA), clamp the CCA, ligate and make an incision at the outer end of the ECA at 0.7 cm, and tie the polylysine-treated nylon suture (length 2 cm, Φ=0.26 mm, smooth at the top) The diameter of the round head is 0.36mm) and gently inserted into the cranium through the ECA and ICA to the bifurcation of the middle cerebral artery (MCA) (the nylon thread enters the bifurcation of the CCA about 18mm) and then ligated to block the cerebral blood flow of the MCA. Ischemia was performed by unclamping the CCA. After 2 hours of ischemia, the nylon suture was gently withdrawn to achieve reperfusion. Afterwards, the wounds of the animals were sutured and kept in individual cages. The body temperature of the animals was maintained at 34°C and the room temperature at 28°C throughout the experiment.

Neurobehavioral Score:

The animals in each group were taken out 24 hours after reperfusion, and the behavioral defects of the animals were graded and scored according to the Bederson method. The standards were: 0 points: no neurological symptoms were observed, and the behavior was normal; The contralateral forelimb showed wrist and elbow flexion, shoulder internal rotation, elbow abduction, and was close to the chest wall; 2 points: the animal was placed on a smooth surface, and the resistance decreased when the operated shoulder was pushed to the opposite side; 3 points: the animal was free Turn around or turn in circles to the opposite side of the operation when walking; 4 points: flaccid paralysis, no spontaneous movement of limbs.

The detection methods of cerebral infarct size and cerebral edema are as follows:

After the behavioral test, the animals were decapitated, and the complete brain was quickly taken out. The blood was washed away with 4°C normal saline, frozen at -20°C for 10 minutes, and then the brain was transected into 5 mm, 7 mm, 9 mm, 11 mm and 13 mm from the olfactory bulb. A 2mm thick slice was placed in a 1% TTC solution at 37°C for 10 minutes in a water bath, protected from light, and photographed after taking it out. The size of the cerebral infarction and the degree of edema were analyzed by computer. There were 8 mice in each group, and the results were averaged.

Cerebral infarction volume (%)=(non-injured brain area-injured non-white area)/(2×non-injured brain area)×100%.

Cerebral edema (%)=(brain area of injured side-brain area of uninjured side)/(2×brain area of uninjured side)×100%.

The results are shown in Table 1 and Fig. 1-Fig. 6; Fig. 1 is the photo of TTC stained brain slice of sham operation (blank) control group; Fig. 2 is the photo of TTC stained brain slice of model control group; Fig. 3 is the photo of lycopene small dose group (20mg /kg) TTC stained brain slice photo; Fig. 4 is the TTC stained brain slice photograph of lycopene medium dose group (50mg/kg); Fig. 5 is the TTC stained brain slice photograph of lycopene large dose group (100mg/kg); Fig. 6 is the photograph of TTC-stained brain slices in the Edaravone group.

Table 1 Effect of lycopene olive oil suspension on cerebral infarction volume, cerebral edema and neurobehavioral score caused by acute cerebral ischemia-reperfusion in rats (Mean ± SD, n=8)

^a : p<0.01 compared with the sham group. ^{b, c} : p<0.05, p<0.01 compared with the model group

The results showed that lycopene had a significant therapeutic effect on cerebral ischemia-reperfusion injury, reduced the volume of cerebral infarction, and significantly reduced the degree of cerebral edema; at the same time, the neurobehavioral scores of animals were significantly improved, indicating that lycopene had a strong effect on neurological function. protective effect.

Example 2, the application of lycopene in the preparation of medicines for the treatment of cerebral ischemic injury

1. Preparation of drugs for the treatment of cerebral ischemic injury

With lycopene as the main drug, the olive oil in the above-mentioned Example 1 was replaced with pure water to make a suspension, which was used for the treatment of acute and convalescent cerebral ischemia-reperfusion injury.

In this example, 0.2 ml of dimethyl sulfoxide (DMSO) was added to 200 mg of lycopene (content > 99%) (purchased from Xi'an Bosheng Biological Co., Ltd., catalog number 13739-02-1) for rapid grinding Emulsify, then gradually add 9.9ml of pure water, mix well, and ultrasonically suspend for 20 minutes to obtain a 20mg/ml lycopene preparation. In the 20 mg/ml lycopene preparation, the mass ratio of lycopene, water and dimethyl sulfoxide is 1:1:49.5.

On this basis, 10 mg/ml lycopene preparation and 5 mg/ml lycopene preparation were obtained by pure dilution, so as to ensure that the volume of water solvent fed to rats under different lycopene doses was equal. Among them, in the 10mg/ml lycopene preparation, the mass ratio of lycopene, water and dimethyl sulfoxide is 1:1:99, and in the 5mg/ml lycopene preparation, the mass ratio of lycopene, water and dimethyl sulfoxide The mass ratio of sulfone is 1:1:198.

2. Evaluation of the effect of drugs in the treatment of cerebral ischemic injury

The method is the same as in Example 1. The results are shown in Table 2.

Table 2 Effect of lycopene water preparation on cerebral infarction volume, cerebral edema and neurobehavioral score caused by acute cerebral ischemia-reperfusion in rats (Mean±SD, n=4)

^{b, c} : p<0.05, p<0.01 compared with the model group.

The results showed that lycopene aqueous suspension also had obvious therapeutic effect on cerebral ischemia-reperfusion injury, reduced the volume of cerebral infarction, and significantly reduced the degree of cerebral edema; at the same time, the neurobehavioral scores of animals improved significantly, indicating that lycopene water The suspension also has a strong protective effect on nerve function.

Claims (9)

1. The application of lycopene in the preparation of the medicine for the treatment of cerebral ischemic injury; the medicine for the treatment of cerebral ischemic injury is at least one of the following 1)-3): 1) Drugs for the treatment of cerebral edema; 2) Drugs for the treatment of cerebral infarction; 3) Drugs for the treatment of nerve damage. 2. The application according to claim 1, characterized in that: the drug is as shown in a) or b) below: a) the drug consists of lycopene, unsaturated phospholipids and dimethyl sulfoxide; b) The drug consists of lycopene, water and dimethylsulfoxide. 3. The application according to claim 2, characterized in that: The mass ratio of lycopene, unsaturated phospholipid and dimethyl sulfoxide in a) is 1:1:99～1:1:495; The mass ratio of lycopene, water and dimethyl sulfoxide in b) is 1:1:49.5˜1:1:198. 4. The application according to claim 2 or 3, characterized in that: The unsaturated phospholipid described in a) is olive oil. 5. A medicine for treating cerebral ischemic injury, the active ingredient of which is lycopene. 6. The medicine according to claim 5, characterized in that: the medicine for treating cerebral ischemic injury is at least one of the following 1)-3): 1) Drugs for the treatment of cerebral edema; 2) Drugs for the treatment of cerebral infarction; 3) Drugs for the treatment of nerve damage. 7. The medicine according to claim 5 or 6, characterized in that: the medicine for treating cerebral ischemic injury is as shown in a) or b) below: a) the drug consists of lycopene, unsaturated phospholipids and dimethyl sulfoxide; b) The drug consists of lycopene, water and dimethylsulfoxide. 8. The medicine according to claim 7, characterized in that: The mass ratio of lycopene, unsaturated phospholipid and dimethyl sulfoxide in a) is 1:1:99～1:1:495; The mass ratio of lycopene, water and dimethyl sulfoxide in b) is 1:1:49.5˜1:1:198. 9. The medicine according to claim 7 or 8, characterized in that: the unsaturated phospholipid in said a) is olive oil.

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