CN103990105B - For composition and method of making the same and the application of memory reinforcing - Google Patents
For composition and method of making the same and the application of memory reinforcing Download PDFInfo
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- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Abstract
The invention discloses a kind of composition and method of making the same for memory reinforcing and application, belong to drugs and health care products development technique field.Said composition is prepared from by the raw material of following percentage by weight: the Cerebrolysin Vial of 10.0%-30.0%, the perilla oil of 90.0%-70.0%.Compositions for memory reinforcing of the present invention, after Cerebrolysin Vial and perilla oil being coordinated with specific ratio, can play synergism, creates unexpected, significant especially effect to memory reinforcing.
Description
Technical field
The present invention relates to a kind of material with memory reinforcing effect, particularly relate to a kind of composition and method of making the same for memory reinforcing and application.
Background technology
Cause the reason of hypomnesis a lot, as: learning pressure is excessive, and the ability of learning and memory of numerous children and adolescents is weakened greatly; What also have along with computer is day by day universal, and adolescence is subject to the puzzlement of hypomnesis just more and more, and main cause is that this crowd is excessively strong to the dependence of computer, thus the brain function of oneself is weakened on the contrary; For another example, in middle-aged and elderly people, the particularly Disease such as hypertension, cerebral arteriosclerosis, its memory ability has obvious fluctuation.Also has some disease, if neurasthenia, depression, cerebral arteriosclerosis, chronic sinusitis etc. are all the common causes causing hypomnesis.The quality of sleep quality also has close ties with memory, and sleeping badly can affect memory function equally.Well-known, the brain of smoking to people has infringement, may cause its hypomnesis along with smoker's age increases; The hobby person that drinks is had also to there will be the phenomenon of hypomnesis.
Domestic and international control memory disorder or the medicine gone down have a lot, and as current clinical conventional oxiracetam and piracetam etc., they are all by improving brain metabolism, reach hypermnesis, improve learning capacity but object.But, the untoward reaction that these chemicalses more or less have people can not accept, and at present only for the treatment of brain injury and the neurological deficit caused, memory and disturbance of intelligence, cannot to general hypomnesis crowd medication.
Current domestic and international market also has the product of some brain boosting and supplementings, but there is the defect of poor effect or complicated in great majority.
Summary of the invention
Based on this, the object of the invention is to the defect overcoming prior art, provide a kind of compositions for memory reinforcing, it is simple that said composition has composition, the extraordinary advantage of effect.
For achieving the above object, the present invention takes following technical scheme:
For a compositions for memory reinforcing, be prepared from by the raw material of following percentage by weight:
Cerebrolysin Vial 10.0%-30.0%
Perilla oil 90.0%-70.0%.
Compositions for memory reinforcing of the present invention, is made up of Cerebrolysin Vial, perilla oil.
Wherein, Cerebrolysin Vial is a kind of bioactive peptide hydrolysate extracted from the fresh cerebral tissue of health pig; containing materials such as several amino acids and cephalin, lecithin, peptide class nerve growth factor; brain is entered by blood brain barrier; and promote the synthesis of brain internal protein, affect respiratory chain, there is the protective capability of anti-hypoxia; improve the metabolism of brain self-energy, activated adenyl cyclase and other Hormone system of catalysis.The effects such as neurotransmitter, peptide hormone and coenzyme precursors are provided.
Perilla oil is from Labiatae medicinal plants Folium Perillae (Lamiales, Perillafrutescens) seed obtains, it is a kind of high undersaturated natural oil, contained Main Ingredients and Appearance is alpha-linolenic acid, content, up to 50%-70%, is the natural oil that in all crude vegetals found at present, alpha-linolenic acid content is the highest.
Although, current people have known that Cerebrolysin Vial or perilla oil can promote the function of learning and memory of little mouse, but, the present invention finds after a large amount of experiments, after Cerebrolysin Vial and perilla oil are coordinated with specific ratio, can synergism be played, unexpected, particularly preferred effect is created to memory reinforcing.And, active ingredient in said composition is only Cerebrolysin Vial and perilla oil, the safety of the two is very high, therefore, said composition is relative to the drug regimen of some active ingredient complexity, have the advantage that side effect is less, be almost applicable to all groups and eat, especially anemia of pregnant woman, child, student and mental worker, old people and myopia, hyperlipidemia, hepatopath often eat.
Wherein in some embodiments, the described compositions for memory reinforcing, is prepared from by the raw material of following percentage by weight:
Cerebrolysin Vial 15.0%-20.0%
Perilla oil 85.0%-80.0%.
Wherein in some embodiments, the described compositions for memory reinforcing, is prepared from by the raw material of following percentage by weight:
Cerebrolysin Vial 17.24%
Perilla oil 82.76%.
The present invention also provides a kind of preparation method of the above-mentioned compositions for memory reinforcing, and wherein, the preparation method of described Cerebrolysin Vial is: get Medulla sus domestica, homogenate after cleaning, after adding pepsin and pancreatin hydrolysis, obtains the mixture of molecular weight less than 10000 after ultrafiltration, drying, to obtain final product.
Wherein in some embodiments, the preparation method of described perilla oil is: get dry Fructus Perillae, obtains crude oil in the mode of mechanical expression, and then this crude oil is standing, precipitation, solids removed by filtration impurity, to obtain final product.
The present invention also provides above-mentioned composition for the preparation of the application in the medicine of memory reinforcing.
Wherein in some embodiments, the dosage form of described medicine is oral liquid, hard capsule, soft capsule, tablet, pill, drop pill, granule, powder, sweet unguentum or distillate medicinal water.Above-mentioned dosage form, all by ordinary skill in the art means, adds the applicable excipient meeting dosage form requirement and prepares.
The present invention also provides above-mentioned compositions for the preparation of the application in the health food of memory reinforcing.
Wherein in some embodiments, described health food is teabag, medicated wine, fresh juice agent.Said composition, all by routine techniques means, joins in applicable food, prepares using this food as substrate by above-mentioned health food.
Compared with prior art, the present invention has following beneficial effect:
A kind of compositions for memory reinforcing of the present invention, after Cerebrolysin Vial and perilla oil are coordinated with specific ratio, synergism can be played, detect through Behavior Test, biochemical indicator and confirm after pathology detection, said composition is to memory reinforcing, particularly in Behavior Test, create unexpected, significant especially effect.
Particularly in Behavior Test, after the proportioning of wherein Cerebrolysin Vial and perilla oil is changed, or select other vegetable oil being rich in alpha-linolenic acid (as Semen Lini oil) to coordinate with Cerebrolysin Vial, or the undersaturated natural oil selecting perilla oil and other to have beneficial brain effect coordinates, or alone brain Proteolytic enzyme or perilla oil, all do not reach the particularly preferred effect that the present composition has.
Further, the remember consolidation disorders mice that said composition causes normal mouse and scopolamine is all effective, and in normal mouse, show particularly preferred effect.
Accompanying drawing explanation
Fig. 1-5 is respectively M group, L group, D group, A group, E group Hippocampus of Mice aspect graph (× 100);
Fig. 6-10 is respectively M group, L group, D group, A group, E group Hippocampus of Mice aspect graph (× 400);
Figure 11-15 is respectively M ' group, L ' group, D ' group, A ' group, E ' organize Hippocampus of Mice aspect graph (× 100);
Figure 16-20 is respectively M ' group, L ' group, D ' group, A ' group, E ' organize Hippocampus of Mice aspect graph (× 400).
Detailed description of the invention
The present invention is described in detail below in conjunction with the drawings and specific embodiments.
Embodiment 1
For a compositions for memory reinforcing, be prepared from by the raw material of following percentage by weight:
Cerebrolysin Vial 17.24%
Perilla oil 82.76%.
The preparation method of the above-mentioned compositions for memory reinforcing, comprises the following steps:
1) preparation of Cerebrolysin Vial.
Get Medulla sus domestica, homogenate after cleaning, after adding pepsin and pancreatin hydrolysis in the usual way, obtains the mixture of molecular weight less than 10000 after ultrafiltration, dry, obtains this Cerebrolysin Vial.
2) preparation of perilla oil.
Get Fructus Perillae, after cleaning, drying, adopt the mode of mechanical expression to obtain crude oil in the usual way, then this crude oil is left standstill, precipitate, filter and obtain.
3) for the preparation of the compositions of memory reinforcing.
By above-mentioned weight ratio, accurately take Cerebrolysin Vial and perilla oil, the two is used in combination.
Embodiment 2
A kind of compositions for memory reinforcing of the present embodiment, substantially identical with the compositions of embodiment 1, difference is, the compositions of the present embodiment is prepared from by the raw material of following percentage by weight:
Cerebrolysin Vial 10.0%
Perilla oil 90.0%.
Embodiment 3
A kind of compositions for memory reinforcing of the present embodiment, substantially identical with the compositions of embodiment 1, difference is, the compositions of the present embodiment is prepared from by the raw material of following percentage by weight:
Cerebrolysin Vial 30.0%
Perilla oil 70.0%.
Comparative example 1
This comparative example prepares a kind of compositions, substantially identical with the compositions of embodiment 1, and difference is, the compositions of this comparative example is prepared from by the raw material of following percentage by weight:
Cerebrolysin Vial 5.0%
Perilla oil 95.0%.
Comparative example 2
This comparative example prepares a kind of compositions, substantially identical with the compositions of embodiment 1, and difference is, the compositions of this comparative example is prepared from by the raw material of following percentage by weight:
Cerebrolysin Vial 40.0%
Perilla oil 60.0%.
Comparative example 3
This comparative example prepares a kind of compositions, is prepared from by the raw material of following percentage by weight:
Cerebrolysin Vial 17.0%
Semen Lini oil 83.0%.
Comparative example 4
This comparative example prepares a kind of compositions, with reference to the method in CN1166386C document, is prepared from by the raw material of following percentage by weight:
Perilla oil 50.0%
Seed oil of Herba Apii graveolentis 50.0%.
Effect experimental examples one
The compositions that the investigation embodiment of the present invention obtains is to the effect of normal mouse memory reinforcing.
Laboratory animal and grouping: ICR mice, by body weight random packet, often organize 14.
Dosage regimen: often organizing mice by setting the continuous gastric infusion of dosage 35 days, freely drinking water and ingest during administration, each group administration fashion and dosage arrangement see the following form 1.
Table 1 is group mice administrations table respectively
| Group is numbered | Administration kind | Dosage |
| A | Embodiment 1 compositions | 540mg/kg·d |
| D | Embodiment 2 compositions | 540mg/kg·d |
| E | Embodiment 3 compositions | 540mg/kg·d |
| F | Comparative example 1 compositions | 540mg/kg·d |
| G | Comparative example 2 compositions | 540mg/kg·d |
| H | Comparative example 3 compositions | 540mg/kg·d |
| I | Comparative example 4 compositions | 540mg/kg·d |
| J | The Cerebrolysin Vial that embodiment 1 prepares | 540mg/kg·d |
| K | The perilla oil that embodiment 1 prepares | 540mg/kg·d |
| L | Piracetam | 540mg/kg·d |
| M | Normal saline | 0.1ml/10g·d |
One, Behaviors survey
(1) Morris water maze (Morriswatermazetest, MWMT)
1. experimental technique: successive administration 35 days, last administration to start training next day, and last 7 days, period continues gastric infusion, once a day.Platform is placed in the middle of first quartile, within first 6 days, be location navigation experiment, choose two respectively, three, the centre at four-quadrant edge is place of entry, measure mice and arrive plateau time (incubation period).The mouse experiment time is defined as 120s, if mice does not arrive platform in 120s, is then placed on by mice on platform and stops 10 seconds, and mice is arrived plateau time be designated as 120s, if arrive platform in mice 120s, then taken away, namely terminate training.Within 7th day, carry out space search experiment, remove platform, measure mice spanning platform number of times in 120s, pass through effective district number of times, effectively district's time of staying, the platform time of staying, platform stop distance/distance covered by a swimming contest, platform stop distance, effectively district stop distance/distance covered by a swimming contest, effective district's stop distance.
2. statistical method: adopt SPSS16.0 statistical software to carry out statistical analysis to experimental data, final result is represented by equal numerical expression standard deviation (X ± S) form, employing t inspection is compared between organizing.
3. experimental result: as shown in following table 2-3.
Table 2. normal mouse water maze incubation period (s) (X ± S, n=14)
* P ﹤ 0.05 is compared with Normal group (M)
Table 3. normal mouse the 7th day water maze situation (X ± S, n=14)
* compare P ﹤ 0.05, * * with Normal group (M) and compare P ﹤ 0.01 with Normal group (M)
By the above results, we can find out, with Normal group (M) ratio, positive controls (L) the 3rd, 6 day incubation period obviously shorten (P ﹤ 0.05), though other index is improved, and there was no significant difference.
D group, A group, E group mice all obviously shorten (P ﹤ 0.05) the 2 to 6 day incubation period, and the 7th balance table time of staying also all obviously extended (P ﹤ 0.05), further, A group mice the 7th day spanning platform number of times, platform stops distance/distance covered by a swimming contest, effectively district's stop distance/distance covered by a swimming contest and is also all significantly increased (P ﹤ 0.05).
With Normal group (M) ratio, F, G, H, I group mice incubation period of 1-6 days or have the trend of shortening, but not significant difference.Though the trend that each index in the 7th day water maze test is improved, but and there was no significant difference.
With Normal group (M) ratio, J, K group mice is similar to positive controls, 3rd, obviously shorten (P ﹤ 0.05) 6 day incubation period, but though the trend that is improved of each index in its 7th day water maze test, but and there was no significant difference.
(2) Jumping test (Step-downtest)
1. experimental technique: first day is the training stage, second day is test phase, and period continues gastric infusion.Mice is put into reaction chamber by first day, conform 3 minutes, then mice is put in reaction chamber, be placed on copper grid, the alternating current of logical 36V, record errors number in every mice 5 minutes, in this, as training achievement, 24h recast is test, and is that memory keeps test, records the mistake sum in every mice 5 minutes and incubation period (jumping off the time of platform for the first time).
2. statistical method: adopt SPSS16.0 statistical software to carry out statistical analysis to experimental data, final result is represented by mean scholar standard deviation (X ± S) form, employing t inspection is compared between organizing.
3. experimental result: as shown in table 4 below.
Table 4. normal mouse Jumping test situation (X ± S, n=14)
* P ﹤ 0.05 is compared with Normal group.
Pass through the above results, we can find out, with Normal group (M) ratio, D, A, E group mice the 2nd day errors number all obviously reduces (P ﹤ 0.05), further, A group mice also obviously increases (P ﹤ 0.05) the 2nd day incubation period.
And positive controls (L) though, the indices of F, G, H, I, J, K group mice is improved, but and there was no significant difference.
(3) step-through test (Step-throughtest)
1. experimental technique: first day is the training stage, second day is test phase, and period continues gastric infusion.First day, puts into bright room by mice, and records errors number (entering darkroom number of times) in mice 5 minutes, in this, as training achievement.Recast test after 24h, records errors number in 5 minutes and animal and enters darkroom incubation period from bright room.
2. statistical method: adopt SPSS16.0 statistical software to carry out statistical analysis to experimental data, final result is represented by mean scholar standard deviation (X ± S) form, employing t inspection is compared between organizing.
3. experimental result: as shown in table 5 below.
Table 5. normal mouse step-through test situation (X ± S, n=14)
* compare P ﹤ 0.05, * * with Normal group and compare P ﹤ 0.01 with Normal group
Pass through the above results, we can find out, with Normal group (M) ratio, D, A, E group mice the 2nd day errors number all obviously reduces (P ﹤ 0.05), further, D, A group mice the 1st day errors number also has remarkable minimizing, the 2nd day incubation period also obviously to increase (P ﹤ 0.01).
And positive controls (L) though, the indices of F, G, H, I, J, K group mice is improved, but and there was no significant difference.
Two, biochemical indicator detects
(1) Serum Indexes detects
Serum sample prepare: after above-mentioned Behaviors survey terminates, water 12-16h is can't help in all mice fasting, and eyeball gets blood, at 4 DEG C preserve, 5000rpm/min, centrifugal 10 minutes, be separated get serum, be placed in-80 DEG C frozen to be measured.
(1) SOD viability examination in serum: (institute of biological products is built up in Nanjing to detecting step reference reagent box, 20140109) description, ultraviolet spectrophotometer measures 550nm place absorbance, and calculates SOD vigor in serum, as shown in table 6 below.
(2) MDA content detection in serum: (institute of biological products is built up in Nanjing to detecting step reference reagent box, 20140109) description, ultraviolet spectrophotometer measures 532nm place absorbance, and calculates MDA content in serum, as shown in table 6 below.
Table 6 Normal Mouse Serum SOD, MDA value (X ± S, n=12)
* compare P ﹤ 0.05, * * with Normal group and compare P ﹤ 0.01 with Normal group
By in the above results, we can find out, with Normal group (M) ratio, positive controls (L), A group, E group mice serum SOD vigor obviously raise (P ﹤ 0.01).Compared with normal group, in each administration group mice serum, MDA content has rising trend, but not statistically significant.
(2) tissue index of correlation detects
Prepared by brain tissue homogenate: the mice marrow that breaks is put to death, and gets its brain, shreds on ice, be placed in test tube, add the normal saline of 9 times of volumes, in ice-water bath with homogenizer grinding fully, be made into 10% homogenate ,-80 DEG C of Refrigerator stores are to be measured.
(1) cerebral tissue determining the protein quantity: homogenate in 4 DEG C, 3500rpm/min, centrifugal 10 minutes.Aspirate supernatant, detecting step reference reagent box (Beijing DingGuo ChangSheng Biology Technology Co., Ltd, 20140210) description, detects its absorbance in 562nm place microplate reader and calculates wherein protein content, as shown in table 7 below.
(2) Brain Tissue SOD Vigor detect: homogenate in 4 DEG C, 3500rpm/min, centrifugal 10 minutes.Aspirate supernatant, detecting step reference reagent box (institute of biological products is built up in Nanjing, 20140109) description, ultraviolet spectrophotometer measures 550nm place absorbance, and calculates SOD vigor in cerebral tissue, as shown in table 7 below.
(3) cerebral tissue MDA content detection: (institute of biological products is built up in Nanjing to detecting step reference reagent box, 20140109) description, ultraviolet spectrophotometer measures 532nm place absorbance, and calculates MDA content in cerebral tissue, as shown in table 7 below.
(4) cerebral tissue Glu assay: get 10% brain tissue homogenate, detecting step reference reagent box (Shang Le institute of biological products, Shanghai, 201403) description, microplate reader measures 550nm place absorbance, according to standard curve, calculate Glu content in cerebral tissue, as shown in table 7 below.
(5) cerebral tissue GABA content measures: get 10% brain tissue homogenate, detecting step reference reagent box (Shang Le institute of biological products, Shanghai, 201403) description, microplate reader measures 550nm place absorbance, according to standard curve, calculate GABA content in cerebral tissue, as shown in table 7 below.
(6) AchE assay in cerebral tissue: get 10% brain tissue homogenate, detecting step reference reagent box (Shang Le institute of biological products, Shanghai, 201403) description, microplate reader measures 550nm place absorbance, according to standard curve, calculate AchE content in cerebral tissue, as shown in table 7 below.
(7) eNOS assay in cerebral tissue: get 10% brain tissue homogenate, detecting step reference reagent box (Shang Le institute of biological products, Shanghai, 201403) description, microplate reader measures 550nm place absorbance, according to standard curve, calculate NOS content in cerebral tissue, as shown in table 7 below.
(8) Na+-K+-ATPase in cerebral tissue: assay: get 10% brain tissue homogenate, detecting step reference reagent box (Shang Le institute of biological products, Shanghai, 201403) description, microplate reader measures 550nm place absorbance, according to standard curve, calculate Na+-K+-ATPase content in cerebral tissue, as shown in table 7 below.
(9) MAO-B assay in cerebral tissue: get 10% brain tissue homogenate, detecting step reference reagent box (Shang Le institute of biological products, Shanghai, 201403) description, microplate reader measures 550nm place absorbance, according to standard curve, calculate MAO-B content in cerebral tissue, as shown in table 7 below.
Table 7. normal mouse tissues biochemical indicator (Mean ± S, n=12)
* P ﹤ 0.05 is compared with Normal group
By the above results, we can find out, with Normal group ratio, each administration group Mice brain tissues protein content, SOD vigor all have rising trend, but not statistically significant.
With Normal group ratio, D group, A group Mice brain tissues MDA content have reduction trend, but not statistically significant, E group Mice brain tissues MDA content obviously reduces (P ﹤ 0.05), positive controls (L) Mice brain tissues MDA content there was no significant difference.
With Normal group ratio, D group, A group, E group Mice brain tissues Glu content obviously raise (P ﹤ 0.05), and other respectively organize there was no significant difference.
With Normal group ratio, D group Mice brain tissues GABA content has rising trend, but not statistically significant, but A group, E group Mice brain tissues GABA content obviously raise (P ﹤ 0.01), positive controls Mice brain tissues GABA content there was no significant difference.
With Normal group ratio, each administration group Mice brain tissues AchE content there are no significant difference.
With Normal group ratio, D group, A group Mice brain tissues eNOS content obviously raise (P ﹤ 0.05), and E group, positive controls (L) are though Mice brain tissues eNOS content has rising trend, not statistically significant.
With Normal group ratio, D group, A group, positive controls (L) Mice brain tissues Na
+-K
+-ATPase content has rising trend, but not statistically significant, and E group Mice brain tissues Na+-K+-ATPase content obviously raises (P ﹤ 0.05).
With Normal group ratio, E group Mice brain tissues MAO-B content obviously reduces (P ﹤ 0.05), and other respectively organizes mouse brain albumen MAO-B content reduction trend, but not statistically significant.
Three, pathology detect
(1) histopathology section is prepared
The mice marrow that breaks is put to death, and gets its brain and to be dipped in 10% formalin fixing, use paraffin embedding according to a conventional method, make wax stone after the dehydration of tissue ethanol, dimethylbenzene.Wax stone is cut into the thin slice of 5 μm through microtome, drags for sheet, is placed in 60 DEG C of baking ovens and dries for 15 minutes.
(2) HE dyeing
(1) paraffin section is dewaxed through dimethylbenzene, ethanol.
(2) use tap water running water 5 minutes, then soak with distilled water.
(3) dye 2-3 minute in hematoxylin, tap water.
(4) hydrochloride alcohol color separation is put into 5 minutes, with tap water running water 10 minutes.
(5) put into Yihong dyeing 2 minutes, use tap water.
(6) section is dewatered through ethanol, dimethylbenzene.
(7), after dehydration terminates, neutral gum mounting is used.Under optical microscope, observe each group of Mice brain tissues Hippocampus morphological change, and photo is got in bat, as shown in Fig. 1-Figure 10, wherein, Fig. 1-Fig. 5 is amplification 100 times of photos, and Fig. 6-Figure 10 is amplification 400 times of photos.
Fig. 1,6 is Normal group (M) Hippocampus of Mice form, Fig. 2,7 is positive controls (L) Hippocampus of Mice form, Fig. 3,8 is D group Hippocampus of Mice form, Fig. 4,9 is A group Hippocampus of Mice form, and Fig. 5,10 is E group Hippocampus of Mice form.
By in above-mentioned figure, we can find out, under the optical microscope of amplification 100 times, with Normal group ratio, and each administration group Hippocampus of Mice form there was no significant difference.Each group of Hippocampus of Mice structure is more complete, even dyeing, clear-cut, is normal hippocampal tissue form.
Under the optical microscope of amplification 400 times, with Normal group ratio, each administration group hippocampus of mice district cellular morphology there was no significant difference.Ge Zu hippocampus of mice district neuronal cell shape is in oval or circular, and marshalling, cell is full, and without edema phenomenon, cell edges is clear, and kernel is visible, acellular cavity and necrosis, is normal hippocampus cellular morphology.
Effect experimental examples two
The compositions that the investigation embodiment of the present invention obtains is to the effect of remember consolidation disorders mice memory reinforcing.
Laboratory animal and grouping: ICR mice, by body weight random packet, often organize 14.
Dosage regimen: often organizing mice by setting the continuous gastric infusion of dosage 35 days, freely drinking water and ingest during administration, each group administration fashion and dosage arrangement see the following form 8.
Remember consolidation disorders mice modeling in the following manner: mice is 10min lumbar injection scopolamine 5mg/kgBW before experiment every day, optionally blocks m receptor to reach the object causing hypomnesis by scopolamine.
Table 8 is group mice administrations table respectively
| Group is numbered | Administration kind | Dosage |
| A’ | Embodiment 1 compositions | 540mg/kg·d |
| B’ | Embodiment 2 compositions | 540mg/kg·d |
| C’ | Embodiment 3 compositions | 540mg/kg·d |
| D’ | Embodiment 4 compositions | 540mg/kg·d |
| E’ | Embodiment 5 compositions | 540mg/kg·d |
| F’ | Comparative example 1 compositions | 540mg/kg·d |
| G’ | Comparative example 2 compositions | 540mg/kg·d |
| H’ | Comparative example 3 compositions | 540mg/kg·d |
| I’ | Comparative example 4 compositions | 540mg/kg·d |
| J’ | The Cerebrolysin Vial that embodiment 1 prepares | 540mg/kg·d |
| K’ | The perilla oil that embodiment 1 prepares | 540mg/kg·d |
| L’ | Piracetam | 540mg/kg·d |
| M’ | Normal saline | 0.1ml/10g·d |
(1) remember consolidation disorders mice Morris water maze laboratory (Morriswatermazetest, MWMT)
1. experimental technique: with normal mouse water maze laboratory, except Normal group (M), other group is the front 10min lumbar injection scopolamine 5mg/kgBW of training before experiment every day.
2. statistical method: with normal mouse statistical method.
3. experimental result: as shown in following table 9-10.
Table 9. remember consolidation disorders Mice water maze incubation period (s) (X ± S, n=14)
* P ﹤ 0.05 is compared with Normal group
Table 10. remember consolidation disorders mice the 7th day water maze situation (X ± S, n=14)
# compares P ﹤ 0.05 ， ﹡ and compares P ﹤ 0.05 with model group with Normal group,
Pass through the above results, we can find out, with Normal group (M) ratio, model group (M ') mice the 1st, 2,5,6 day incubation period obviously extend (P ﹤ 0.05), 7th day spanning platform number of times, pass through effective district number of times and obviously reduce, effective district's stop distance, the platform time of staying, effectively district's time of staying, platform stop distance, effectively district's stop distance/distance covered by a swimming contest and all obviously shorten (P ﹤ 0.05), and modeling success is described.
With model group (M ') ratio, D ' organizes mice the 2nd, 6 day incubation period obviously shortened (P ﹤ 0.05), 7th day spanning platform number of times, pass through effective district number of times and obviously increase (P ﹤ 0.05), the platform time of staying, platform stops distance, platform stops distance/distance covered by a swimming contest, effectively district's time of staying, effectively district stop distance, effectively district's stop distance/distance covered by a swimming contest and all obviously extend (P ﹤ 0.05).
With model group (M ') ratio, A ' organizes mice the 1st, 2,5,6 day incubation period obviously shortened (P ﹤ 0.05), 7th balance effective district time of staying, the platform time of staying, effectively district's stop distance, platform stop distance, effectively district's stop distance/distance covered by a swimming contest and all obviously extend (P ﹤ 0.05), though platform stops distance/distance covered by a swimming contest prolongation trend, but and there was no significant difference.
With model group (M ') ratio, E ' organizes mice the 4th, 5,6 day incubation period obviously shortened (P ﹤ 0.05), 7th day effective district's stop distance, effect district's time of staying, the platform time of staying, effectively district stops distance/distance covered by a swimming contest and all obviously extends (P ﹤ 0.05), though platform stops Distance geometry platform stop distance/distance covered by a swimming contest prolongation trend, but and there was no significant difference.
With model group ratio, positive controls (L ') mice the 2nd, 3 day incubation period significantly shorten (P ﹤ 0.05), 7th day spanning platform number of times, pass through effective district number of times and obviously increase (P ﹤ 0.05), effective district obviously extends (P ﹤ 0.05) time of staying, other each index there was no significant differences.
H ', I ' are though mice 1-6 is improved a days incubation period, but and there was no significant difference.During the 7th day water maze laboratory, H ' mice passes through effective district number of times obviously to be increased (P ﹤ 0.05), and I ' mice effective district obviously increases (P ﹤ 0.05) time of staying, other index there was no significant difference.
F ', G ', J ', K ' are though the indices organizing mice is improved, but and there was no significant difference.
(2) remember consolidation disorders mice Jumping test (Step-downtest)
1. experimental technique: with normal mouse Jumping test.After first day experiment (training stage), except matched group, other organize lumbar injection sodium nitrite solution 120mg/kg immediately.Number of shocks in record mice first day 5 minutes, the number of shocks in second day 5 minutes and jump off incubation period of platform.
2. statistical method: adopt SPSS16.0 statistical software to carry out statistical analysis to experimental data, final result is represented by mean scholar standard deviation (X ± S) form, employing t inspection is compared between organizing.
3. experimental result: as shown in table 11 below.
Table 11. remember consolidation disorders mice Jumping test situation (X ± S, n=14)
* compare with model group P ﹤ 0.05, * * with model group than P ﹤ 0.01, # with Normal group than P ﹤ 0.05
Pass through the above results, we can find out, with Normal group (M) ratio, model group (M ') mice first day, second day errors number obviously increase (P ﹤ 0.05), second day incubation period obviously shortened (P ﹤ 0.05), and modeling success is described.
With model group ratio, D ' group, A ' organize first day, second day errors number obviously reduces (P ﹤ 0.05, P ﹤ 0.01), and second day incubation period obviously extended (P ﹤ 0.05); E ' organizes mice second day errors number and obviously reduces (P ﹤ 0.05), though other index is improved trend, but and there was no significant difference.
With model group ratio, positive controls mice first day errors number obviously reduces (P ﹤ 0.05); Other each index there was no significant differences.
With model group ratio, G ' group, I ' organize first day errors number and obviously reduce (P ﹤ 0.05), though other index is improved trend, but and there was no significant difference.
With model group ratio, F ' group, H ' group, J ' group, K ' are improved trend though organize indices, but and there was no significant difference.
(3) remember consolidation disorders mice step-through test (Step-throughtest)
1. experimental technique: with normal mouse step-through test.Except matched group, other groups were in second day experiment (test phase) first 30 minutes gastric infusion 30% ethanol 0.lml/10g.Errors number (entering darkroom number of times) in record mice first day 5 minutes, the errors number in second day 5 minutes and animal enter darkroom incubation period from bright room.
2. statistical method: adopt SPSS16.0 statistical software to carry out statistical analysis to experimental data, final result is represented by mean scholar standard deviation (X ± S) form, employing t inspection is compared between organizing.
3. experimental result: as shown in table 12 below.
Table 12. remember consolidation disorders mice step-through test situation (X ± S, n=14)
* compare with model group P ﹤ 0.05, * * with model group than P ﹤ 0.01, # with Normal group than P ﹤ 0.05
Pass through the above results, we can find out, with Normal group (M) ratio, model group (M ') mice second day errors number showed increased (P ﹤ 0.05), second day incubation period obviously shortened (P ﹤ 0.05), and modeling success is described.
With model group ratio, D ', A ' organize mice second day errors number and obviously reduce (P ﹤ 0.05, P ﹤ 0.01), and second day incubation period obviously extended (P ﹤ 0.05); E ' organizes mice first day, second day errors number all obviously reduces (P ﹤ 0.05), and second day incubation period obviously extended (P ﹤ 0.01).
With model group ratio, positive controls (L ') mice second day errors number obviously reduces (P ﹤ 0.05), and other each indexs are improved trend, but and there was no significant difference.
With model group ratio, F ', G ', I ', K ' organize mice second day errors number and obviously reduce (P ﹤ 0.05), and G ' organizes mice obviously extends (P ﹤ 0.05) second day incubation period, other each index there was no significant differences.
H ', J ' organize each index there are no significant difference.
Two, biochemical indicator detects
(1) Serum Indexes detects: the same normal mouse of method, result is as shown in table 13 below.
Table 13 remember consolidation disorders mice serum SOD, MDA value (X ± S, n=12)
* P ﹤ 0.05, # is compared with model group with Normal group than P ﹤ 0.05
By in the above results, we can find out, with blank group (M) ratio, model group (M ') mice serum SOD vigor obviously reduces (P ﹤ 0.05); With model group ratio, D group, A ' organize mice serum SOD vigor rising trend, but not statistically significant.E ' organizes mice serum SOD vigor and obviously raises (P ﹤ 0.05).
With blank group (M) ratio, model group (M ') mice serum MDA content has rising trend, but not statistically significant; With model group ratio, other respectively organize mice serum MDA content trend, but not statistically significant.
(2) tissue index of correlation detects: the same normal mouse of method, result is as shown in table 14 below.
Table 14. remember consolidation disorders mouse tissue biochemical indicator (Mean ± S, n=12)
* compare with model group P ﹤ 0.05, # with Normal group than P ﹤ 0.05, ## with Normal group than P ﹤ 0.01
By the above results, we can know, with Normal group (M) ratio, model group (M ') Mice brain tissues protein content obviously reduces (P ﹤ 0.05); With model group (M ') ratio, positive controls (L '), D ' group, A ' group, E ' organizes Mice brain tissues protein content rising trend, but not statistically significant.
With Normal group ratio, model group Mice brain tissues SOD vigor obviously reduces (P ﹤ 0.05); With model group ratio, positive controls, D ' group, A ' group, E ' organize Mice brain tissues SOD vigor rising trend, but not statistically significant.
With normal group ratio, model group Mice brain tissues MDA content obviously raises (P ﹤ 0.01); With model group ratio, and positive controls (L '), D ', A ' group, E ' Mice brain tissues MDA content obviously declines (P ﹤ 0.05).
With normal group ratio, model group Mice brain tissues Glu content significantly reduces (P ﹤ 0.05); With model group ratio, D ' group, A ' group, E ' organize Mice brain tissues Glu content and obviously raise, and positive controls (L ') Mice brain tissues Glu content there was no significant difference.
With normal group ratio, model group Mice brain tissues GABA content obviously reduces (P ﹤ 0.01); Compared with model group, it is on the rise that D ' organizes Mice brain tissues GABA content, but not statistically significant, A ' group, E ' organize Mice brain tissues GABA content and obviously raise, positive controls Mice brain tissues GABA content there was no significant difference.
With Normal group ratio, model group Mice brain tissues AchE content obviously raises (P ﹤ 0.05); With model group ratio, D ' group, A ' group, E ' group, positive controls Mice brain tissues AchE content there are no significant difference.
With Normal group ratio, model group cerebral tissue eNOS content obviously reduces (P ﹤ 0.05); With model group ratio, D ' group, positive controls Mice brain tissues eNOS content has rising trend, but not statistically significant.A ' group, E ' organize Mice brain tissues eNOS content and obviously raise (P ﹤ 0.05).
With normal control ratio, model group Mice brain tissues Na
+-K
+-ATPase Content obviously reduces (P ﹤ 0.05); With model group ratio, D ' group, A ' group, E ' organize Mice brain tissues Na
+-K
+-ATPase content obviously raises (P ﹤ 0.05), and positive controls (L ') Mice brain tissues Na
+-K
+-ATPase content has rising trend, but not statistically significant.
With Normal group ratio, model group Mice brain tissues MAO-B content significantly raises (P ﹤ 0.05); With model group ratio, E ' group, positive controls Mice brain tissues MAO-B content has reduction trend, but not statistically significant.And A ' group, D ' organize Mice brain tissues MAO-B content obviously reduces (P ﹤ 0.05).
Three, pathology detect
(1) histopathology section is prepared
The same normal mouse of method, clap and get photo as shown in Figure 11-20, wherein, Figure 11-Figure 15 is amplification 100 times of photos, and Figure 16-Figure 20 is amplification 400 times of photos.
Figure 11,16 is model group (M ') Hippocampus of Mice form, Figure 12,17 is positive controls (L ') Hippocampus of Mice form, Figure 13,18 organizes Hippocampus of Mice form for D ', Figure 14,19 organizes Hippocampus of Mice form for A ', and Figure 15,20 organizes Hippocampus of Mice form for E '.
By in above-mentioned figure, we can find out, under the optical microscope of amplification 100 times, compare with Normal group (M), model group (M ') Hippocampus of Mice structure is more complete, uneven dyeing, Hippocampus soft edge, hippocampal tissue generation pathomorphism changes; With model group ratio, D ' group, A ' group, E ' organize Hippocampus of Mice pathomorphism and obviously improve, each group Hippocampus of Mice structural integrity, even dyeing, and Hippocampus profile is more clear, and configuration is tending towards normal; Positive controls Hippocampus of Mice pathomorphism and model group are without significant difference.
Under the optical microscope of amplification 400 times, Normal group (M) hippocampus of mice district cellular morphology is normal; There is obvious pathomorphism and change in model group (M ') hippocampus of mice district cell, cell arrangement loosens, out-of-shape, and most cells is downright bad, pyknosis, vacuolar degeneration, and kernel is invisible, addicted to the dense dye in Yihong; With model group (M ') ratio, D ' group, A ' group, E ' organize hippocampus of mice district cell pathology form and obviously improve, most cells arrangement closely, regular shape, kernel is visible, respective cells pyknosis, necrosis, and configuration is tending towards normal; Positive controls hippocampus of mice district cell pathology form and model group are without significant difference.
The above embodiment only have expressed several embodiment of the present invention, and it describes comparatively concrete and detailed, but therefore can not be interpreted as the restriction to the scope of the claims of the present invention.It should be pointed out that for the person of ordinary skill of the art, without departing from the inventive concept of the premise, can also make some distortion and improvement, these all belong to protection scope of the present invention.Therefore, the protection domain of patent of the present invention should be as the criterion with claims.
Claims (8)
1. for a compositions for memory reinforcing, it is characterized in that, be made up of the raw material of following percentage by weight:
Cerebrolysin Vial 15.0%-20.0%
Perilla oil 85.0%-80.0%
The preparation method of described Cerebrolysin Vial is: get Medulla sus domestica, and homogenate after cleaning, after adding pepsin and pancreatin hydrolysis, obtains the mixture of molecular weight less than 10000 through ultrafiltration, dry, to obtain final product;
The preparation method of described perilla oil is: get dry Fructus Perillae, obtains crude oil in the mode of mechanical expression, and then this crude oil is standing, precipitation, solids removed by filtration impurity, to obtain final product.
2. the compositions for memory reinforcing according to claim 1, is characterized in that, is made up of the raw material of following percentage by weight:
Cerebrolysin Vial 17.24%
Perilla oil 82.76%.
3. the preparation method of the compositions for memory reinforcing described in any one of claim 1-2, it is characterized in that, the preparation method of described Cerebrolysin Vial is: get Medulla sus domestica, homogenate after cleaning, after adding pepsin and pancreatin hydrolysis, the mixture of molecular weight less than 10000 is obtained through ultrafiltration, dry, to obtain final product.
4. the preparation method of the compositions for memory reinforcing according to claim 3, it is characterized in that, the preparation method of described perilla oil is: get dry Fructus Perillae, obtains crude oil in the mode of mechanical expression, then this crude oil is standing, precipitation, solids removed by filtration impurity, to obtain final product.
5. the compositions described in any one of claim 1-2 is for the preparation of the application in the medicine of memory reinforcing.
6. compositions according to claim 5 is for the preparation of the application in the medicine of memory reinforcing, it is characterized in that, the dosage form of described medicine is oral liquid, hard capsule, soft capsule, tablet, pill, drop pill, granule, powder, sweet unguentum or distillate medicinal water.
7. the compositions described in any one of claim 1-2 is for the preparation of the application in the health food of memory reinforcing.
8. compositions according to claim 7 is for the preparation of the application in the health food of memory reinforcing, it is characterized in that, described health food is teabag, medicated wine, fresh juice agent.
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