CN108403734A - The application of glasswort extract and active component in preparing antidepressant - Google Patents
The application of glasswort extract and active component in preparing antidepressant Download PDFInfo
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Abstract
The invention discloses a kind of application of glasswort extract and active component (dichloromethane layer, ethyl acetate layer and n-butanol layer extract) on the drug or health products for preparing antidepressant effect, internal pharmacodynamic experiment shows that glasswort extract and its active component have certain antidepressant effect to chronic stress depression mouse, its mechanism of action may with reduce cortisone and promotion neurotransmitter regulator, anti-oxidant related, and Small side effects, it can be used for the research and development application of antidepressants.
Description
Technical field
The present invention relates to a kind of hairs to belong to medicine, field of health care products, is related to a kind of glasswort extract and active component
Application in terms of preparing antidepressant.
Background technology
Depression belongs to a kind of common central nervous system disease, main clinical manifestation be it is pessimistic and worldweary, depressed,
Feel oneself inferior oppressive, illusion, have introgression etc., commonly referred to as " STOP Emotion Cold ".With people's the accelerating rhythm of life, work
The increasing of pressure, the probability that people suffer from depression increase increasingly.The Western medicine of the common depression of clinic includes mainly that monoamine aoxidizes
Enzyme inhibitor (MAOI), tricyclic drugs, selective serotonin reuptake inhibithors (SSRI) etc..Although Western medicine can be effective
Depression is treated on ground, but side effect is apparent, has certain damage to organs such as stomach, livers, and be unable to answering for prevention of depression
Hair.Therefore the development of Western medicine is restricted, and people are very urgent to the Research Requirements of related natural drug.
Glasswort (Salicornia europaea L.), also known as extra large asparagus, belongs to annual dicotyledonous for Chenopodiaceae salicornia europaeal
Herbaceous plant.Glasswort is grown in lick more, seabeach is taken up in salt lake, has saline taste, is answered as food and natural plants salt
With.Recent study, which finds it not only, soothing the liver, diuresis, decompression, also has hypoglycemic, lipid-loweringing, antitumor, immune adjusts
The multiple biological activities such as section, therefore the medicinal and health value research of glasswort, are increasingly becoming the popular neck studied both at home and abroad
Domain, but have no it in the new application for preparing anti-depression aspect drug.
Invention content
The object of the present invention is to provide a kind of purposes of glasswort extract and active component in preparing antidepressant,
It is shown through test result：Glasswort ethanol extract, water extract and active component (dichloromethane layer, ethyl acetate layer with just
Butanol layer extract) significant antidepressant effect is all had to chronic stress depression model, especially to insomnia
Depression has apparent therapeutic effect.Its mechanism of action may be with reduction cortisone and promotion DA neurotransmitter regulators and antioxygen
Change related, and Small side effects, can be applied to the auxiliary treatment of depressed patient, overcomes insufficient in the prior art.
A kind of purposes of glasswort extract and active component of the present invention in the drug for preparing treatment depression.
The present invention is using the isolated glasswort extract of the alcohol extracting method, water extraction and polarity gradient solvent of routine and has
Imitate drug made of position.
Water extraction, ethanol extraction method or the separation of polarity gradient solvent is respectively adopted in Herba Achilleae extract of the present invention,
Glasswort ethanol extract, water extract and glasswort active component are obtained, concrete operations follow these steps to carry out：
Glasswort ethanol extraction method：
Using glasswort as bulk pharmaceutical chemicals, extracted 3 times with the ethyl alcohol of a concentration of 50-95%, solid-liquid ratio is 10 times, 8 times, 5 times, is carried
It is 2,2,1 hours to take 60~100 DEG C of temperature, each time, merges gained extracting solution three times, and filtrate is concentrated into medicinal extract, and vacuum is dry
It is dry, obtain glasswort extract.
Glasswort water extraction method：
It takes the water that glasswort adds 5~10 times of amounts to impregnate 12~24 hours, adds water refluxing extraction 3 times, solid-liquid ratio is respectively 10
Again, 8 times, 5 times, merge 3 gained extracting solutions, filtrate is concentrated into medicinal extract, is dried in vacuo, obtains glasswort water extract.
The extraction of glasswort active component：
Glasswort medicinal material coarse powder is taken, is 50%-95% ethyl alcohol or water with volumetric concentration, heating and refluxing extraction 0.5-3 hours,
It extracts 3 times altogether, merges extracting solution, filtrate is recycled to concentrate, and it is molten that petroleum ether, dichloromethane, ethyl acetate, n-butanol is added
Agent is extracted, each solvent adding amount according to concentration liquid measure 1: 1 ratio, respectively repeatedly extract 3 times, extraction until
Until upper solution color is thin out, petroleum ether layer, dichloromethane layer, ethyl acetate layer, n-butanol layer and water layer are obtained successively.
It is concentrated to dryness to get each position sample of glasswort under the conditions of 60 DEG C with Rotary Evaporators.
Compared with prior art, the present invention haing the following advantages and advantageous effect：Have present invention is disclosed glasswort and resists
Depression effect provides new approach for its application, and glasswort small toxicity, compliance are good, and no conventional antidepressant agents are not
Good reaction.
Description of the drawings
Fig. 1 is the protein expression of Mice brain tissues Nrf2, Keap1, NQO1, HO-1.
Specific implementation mode
It is further illustrated the present invention below by way of specific embodiment, not limitation of the invention, according to known in this field
The prior art, embodiments of the present invention are not limited to specific embodiment.In the premise without prejudice to spirit of that invention and principle
Under, it is fallen in the scope of the present invention to inventing any modifications or changes that individual technical steps carry out.
Embodiment 1
Pharmacodynamic experiment of the drug to CUMS blahs aypnia mouse models
1 experiment material
1.1 experimental animal
Male ICR mouse (20 ± 2g of weight) is purchased from Beijing HFK Bio-Technology Co., Ltd..Raising 3 before experiment
It adapts to environment.22 ± 2 DEG C of room temperature, relative humidity 65%~70%, day illumination 12h, free diet take the photograph water.
1.2 reagents and drug
Glasswort extract (laboratory self-control)；(Chengdu Kang Hong medicine companies group share has venlafaxine hydrochloride slow-release capsule
Limit company)；Resolving stagnation for tranquilization particle (Tonghua Limin Pharmaceutical Industry Co., Ltd.)；Amitriptyline hydrochloride tablet (Hunan Dongting Lake medicine company stock
Part Co., Ltd) rat cortical ketone (CORT) enzyme-linked immunoassay kit (the Tianjin bio tech ltd Zi Han)
2 experimental methods
2.1 animal packets and administration
Mouse is randomly divided into 7 groups, respectively blank group, model group, Venlafaxine group, resolving stagnation for tranquilization group, Hai Peng by weight
The high, medium and low dosage group of son, every morning 8:30-9:30 gastric infusions continue 21 days, and dosage regimen is as shown in table 1.
1 animal packet of table and dosage regimen
The method for building up of 2.2 CUMS blahs aypnia mouse models
10 kinds of stress factors of random arrangement are pressed in 21d, arbitrary two kinds of stimulations are given once daily, to avoid mouse from generating adaptation
Mild-natured each stimulation is given 4 times.
(1) moist raising：Suitable quantity of water is added in mouse bedding and padding makes its humidity, and survive 12h in a humid environment.
(2) fasting：12h is jejunitas.
(3) prohibit water：12h cuts off the water supply.
(4) raising is tilted：Mouse cage is tilted into 45 ° of raising mouse 12h.
(5) tail is pressed from both sides：Mouse is put into fixed cage, is clamped away from lasting 1min at root of the tail 1cm with haemostatic clamp.
(6) it is raised without bedding and padding：Mouse, each 12h are raised in the mouse cage of no bedding and padding.
(7) sleep deprivation：Sleep deprivation is for 24 hours.Using chain-wales water environment method, in container bottom setting diameter 3cm, height
The platform of 2cm, periphery fill water (depth of water 0.5cm), can fall into the water if mice sleep.
(8) cold-wate swimming：Animal is placed on to (depth of water 5cm) 5min in the container for filling 10 DEG C of water.
(9) hot water is swum：Animal is placed on to (depth of water 5cm) 5min in the container for filling 45 DEG C of water.
(10) it overturns round the clock：In early 7:The cage of mouse is placed on dark space when 00 not turn on light, until evening 19:It will when 00
Mouse is placed in the room turned on light until next day early 7:When 00.
2.3 observation indexs and method
(1) tail-suspention test (TST)
Mouse tail end is attached on a horizontal waddy, waddy is liftoff, and 30cm makes animal in vacantly projecting shape, and suspension both sides are used
Partition board blocks mouse sight.It tests and carries out 6min, the mouse dead time in 4min after record.The motionless row of mouse in tail-suspention test
For judgment criteria be that mouse head is downward, four limbs naturally droop, mouse forelimb have a little it is mobile be still considered as it is motionless.
(2) forced swim test (FST)
Mouse is placed individually into the transparent glass cylinder for filling 25 DEG C of deep 10cm or more, temperature water, experiment carries out 6min, note
The mouse dead time in 4min after record.Motionless standard in forced swim test is mouse only by head above water, limbs floating
Maintain motionless state.
(3) syrup consumption experiment
Prohibit water and carry out syrup consumption experiment afterwards for 24 hours, gives every cage mouse syrup, each 200mL of pure water, take two away afterwards for 24 hours
Bottle simultaneously weighs, and records syrup and pure water consumption, and calculate syrup preference [syrup preference=syrup consumption/
Total liquid-consumed amount × 100%].Syrup preference is usually used in evaluating anhedonia degree, and anhedonia refers to syrup preference journey
Degree averagely has decline compared with normal control, reference water.
(4) in Mice brain tissues monoamine neurotransmitters measurement
Each group mouse is after last dose 30min, sacrificed by decapitation, and hippocampus and cortex is taken to be respectively put into two E.P pipes.It adopts
Hippocampus and the content of cortex DA, NE, 5-HT are measured with sepectrophotofluorometer method.Content is calculated according to formula, and is united
Meter analysis.Content calculation formula is as follows：
(5) influence of the glasswort to CORT in CUMS mice serums
For each group mouse after last dose 30min, eyeball centrifuges 10min after taking blood, blood sample to place 30min, takes supernatant, standby
With.The content that CORT in mice serum is measured using ELISA method is operated in strict accordance with kit specification and is executed.
2.4 statistical analysis
Experimental data is with means standard deviationIt indicates, is counted using 17.0 softwares of SPSS, with single factor test
Variance Method is analyzed, using P ＜ 0.05 and P ＜ 0.01 as significant difference standard.
3 results
Influence of the 3.1 glasswort extracts to CUMS blahs aypnia mouse TST
The TST of each group mouse is the results show that compared with model group, when the TST of blank group and other administration group mouse is motionless
Between reduce in various degree, wherein blank group, Venlafaxine group, resolving stagnation for tranquilization group and the TST of glasswort high dose group mouse be not
The dynamic time is substantially less than model group (P ＜ 0.01), and glasswort middle dose group has significant difference (P ＜ compared with model group
0.05).Analysis finds that glasswort is presented a certain amount to the TST dead times of chronic stress depression mouse and imitates relationship.As a result
It is shown in Table 2.
2 glasswort extract of table to CUMS blahs aypnia mouse TST influence (N=10)
Note：Compared with model group,*P ＜ 0.05,**P ＜ 0.01
Influence of the 3.2 glasswort extracts to CUMS blahs aypnia mouse FST
Compared with model group, the FST dead times of blank group and each administration group substantially reduce (P ＜ 0.01), wherein sea
Fluffy son height, middle dose group best results, difference have conspicuousness statistical significance (P ＜ 0.001), and table 3 is the results show that Hai Peng
Seed extract has a certain amount to imitate relationship the FST effects of CUMS mouse.
3 glasswort extract of table to CUMS blahs aypnia mouse FST influence (N=10)
Note：Compared with model group,**P ＜ 0.01,***P ＜ 0.001
Influence of the 3.3 glasswort extracts to mouse syrup intake
Table 4 is the results show that the raising all different degrees of compared with model group to the preference of syrup of each administration group.Hai Peng
Seed extract is high, middle dose group is higher than positive drug control group to the intake of syrup.
Influence of the 4 glasswort extract of table to CUMS mouse syrup intakes
Influence of the 3.4 glasswort extracts to Hippocampus of Mice monoamine neurotransmitters
Under showing that DA, NE, 5-HT content and blank group are more in model group Hippocampus of Mice by 5 test result of table
Drop.Compared with model group, DA is increased significantly (P ＜ 0.05 in Venlafaxine and glasswort extract low dose group hippocampus of mice
Or P ＜ 0.001), 5-HT contents significantly increase in resolving stagnation for tranquilization group Hippocampus of Mice, significant difference (P ＜ 0.001),
NE, 5-HT have different degrees of raising trend in each administration group Hippocampus of Mice of glasswort, but there was no significant difference (P ＞
0.05)。
5 glasswort extract of table to Hippocampus of Mice monoamine neurotransmitters influence (N=10)
Note：Compared with model group,*P ＜ 0.05,***P ＜ 0.001
Influence of the 3.5 glasswort extracts to mouse cortex tissue monoamine neurotransmitters
6 experimental result of table is shown, compared with blank group, DA, NE, 5-HT content in model group Cerebral Cortex under
Drop, and NE, 5-HT content decline apparent (P ＜ 0.05, P ＜ 0.01).Compared with model group, each administration group mouse of glasswort is big
NE, 5-HT content are on the rise in cortex, and 5-HT contents significantly increase in Venlafaxine group Cerebral Cortex
(P ＜ 0.05), in addition, NE contents have significant difference (P ＜ with model group in glasswort high dose group mouse cortex
0.05)。
6 glasswort extract of table to mouse cortex tissue monoamine neurotransmitters influence (N=10)
Note：Compared with model group,*P ＜ 0.05,**P ＜ 0.01
Influence of 3.6 glassworts to CORT in CUMS mice serums
As seen from the results in Table 7, CORT contents are significantly lower than mould in blank group, Venlafaxine group and glasswort group mice serum
Type group, and each group of data all has significant significant difference (P ＜ 0.05, P ＜ 0.01) compared with model group.
7 glasswort of table to CORT in CUMS mice serums influence (N=10)
Note：Compared with model group,*P ＜ 0.05,**P ＜ 0.01
Embodiment 2
1 experiment material
1.1 experimental animal
With content under " 1.1 " item in embodiment 1.
1.2 reagents and drug
It makes by oneself in glasswort (n-butanol group, ethyl acetate group, dichloromethane group) laboratory
Amitriptyline hydrochloride tablet Hunan Dongting Pharmaceutical Co., Ltd.
Yellow Jackets Shandong West Asia chemical industry Co., Ltd
2 experimental methods
2.1 animal packets and administration
Mouse is randomly divided into 6 groups, and every group 10, it is different that blank group, model group, positive drug group and glasswort is respectively set
Regional administration group.In addition to blank group, remaining each group mouse presses method modeling under " 2.2 " item, every from the daystart of modeling
Its morning 8:00-9:00 gastric infusion continues 21 days once a day, and grouping and dosage regimen are shown in Table 8.
8 animal packet of table and dosage regimen
Note：Glasswort n-butanol layer group, glasswort ethyl acetate group, glasswort dichloromethane group are respectively equivalent to glasswort
Extract 1.8g/kg, it is equivalent to fresh glasswort 60g/kg.
The method for building up of 2.2 CUMS blahs aypnia mouse models
With content under " 2.2 " item in embodiment 1.
2.3 observation indexs and method
2.3.1 weight
Weight of the record mouse before experiment starts and in experimentation respectively is administered the 0th day, the 6th day, administration is administered
12nd day, the weight for being administered the 18th day, being administered the 24th day.
2.3.2 autonomic activities are tested
Each group mouse is individually put into mouse cage after gavage 0.5h by label sequence, in administration the 13rd day independently to live
Dynamic tester voluntarily records its activity condition.Every mouse first adapts to 10min under autonomic activities instrument, records subsequent 5min
The autonomic activities of interior mouse, experiment carry out in room temperature quiet environment.
2.3.3 tail-suspention test (TST)
With content under " 2.3 " (1) item in embodiment 1.
2.3.4 forced swimming (FST)
With content under " 2.3 " (2) item in embodiment 1.
2.3.5 Western Blot methods detect Mice brain tissues Nrf2, Keap1, NQO1, HO-1 protein expression
Brain tissue is ground, the lysate of corresponding proportion is added, 12000r centrifuges 20min, takes supernatant spare.Using BCA
Method carries out the concentration mensuration of protein sample, by protein sample denaturation, SDS-PAGE electrophoresis, gel transferring film, detects small
Nuclear factor E2-related factor 2 (Nrf2), Kelch sample epoxychloropropane related protein-1s (Keap1), quinone oxidation are gone back in murine brain
Protoenzyme -1 (NQO1), DELTA rHO-1 (HO-1) protein expression.
2.4 statistical analysis
With content under " 2.4 " item in embodiment 1.
3 experimental results
Influence of each position of 3.1 glassworts to CUMS blahs aypnia mouse weights
As seen from the results in Table 9, each group mouse weight no significant difference (P ＞ 0.05) shows each position of glasswort to mouse
Weight has no significant effect.
Each position of 9 glasswort of table to CUMS mouse weights influence (N=10)
Influence of each position of 3.2 glassworts to CUMS blahs aypnia mouse autonomic activities
The results are shown in Table 10, and compared with model control group, the autonomic activities number of blank group and each administration group mouse is equal
No significant difference (P ＞ 0.05), each administration group antidepressant effect reach not by enhancing mouse autonomic activities.
Each position of 10 glasswort of table to CUMS mouse autonomic activities influence (N=10)
Influence of each position of 3.3 glassworts to the CUMS blahs aypnia mouse TST dead times
Table 11 the result shows that, amitriptyline group with each position group of glasswort hang the tail dead time subtract compared with model group
Few, wherein dichloromethane layer has statistical significance (P ＜ 0.05, P ＜ 0.01) with n-butanol layer extract group difference, wherein
Glasswort n-butanol group best results (P ＜ 0.01).
Each position of 11 glasswort of table to CUMS mouse TST influence (N=10)
Note：Compared with model group,*P ＜ 0.05,**P ＜ 0.01
Influence of each position of 3.4 glassworts to the CUMS blahs aypnia mouse FST dead times
Difference group other mouse FST dead times the results are shown in Table 12.Compared with model group, blank group and each administration group mouse
The FST dead times are reduced, and wherein dichloromethane layer, ethyl acetate layer and n-butanol layer extract group difference have statistics
Meaning (P ＜ 0.01) shows that this three glasswort positions have antidepressant effect.
Each position of 12 glasswort of table to CUMS mouse FST influence (N=10)
Note：Compared with model group,*P ＜ 0.05,**P ＜ 0.01
Nrf2, Keap1, NQO1, HO-1 protein expression in 3.5 Western Blot detection Mice brain tissues
As shown in Figure 1, compared with model group, glasswort n-butanol group, glasswort ethyl acetate group and glasswort dichloromethane
Alkane group can raise the expression of Nrf2, Keap1, NQO1, HO-1 albumen, show that each position of glasswort may be by adjusting
Nrf2-ARE signal paths enhance antioxidation, to protect nerve cell to a certain extent, reach resolving stagnation for tranquilization work(
Effect.
Claims (1)
1. a kind of application of glasswort extract and active component in preparing medicament for treatment of depression.
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Application publication date: 20180817 |