CN107987175A - A kind of fragrant newborn mushroom polysaccharide LC-1 and preparation method and application - Google Patents
A kind of fragrant newborn mushroom polysaccharide LC-1 and preparation method and application Download PDFInfo
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- CN107987175A CN107987175A CN201711205932.XA CN201711205932A CN107987175A CN 107987175 A CN107987175 A CN 107987175A CN 201711205932 A CN201711205932 A CN 201711205932A CN 107987175 A CN107987175 A CN 107987175A
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/0003—General processes for their isolation or fractionation, e.g. purification or extraction from biomass
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/006—Heteroglycans, i.e. polysaccharides having more than one sugar residue in the main chain in either alternating or less regular sequence; Gellans; Succinoglycans; Arabinogalactans; Tragacanth or gum tragacanth or traganth from Astragalus; Gum Karaya from Sterculia urens; Gum Ghatti from Anogeissus latifolia; Derivatives thereof
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Abstract
The invention discloses a kind of fragrant newborn mushroom polysaccharide LC 1 and preparation method and application, the heteroglycan that fragrant breast mushroom polysaccharide is made of two monose, i.e. β D glucose and α D-galactolipin is with 2:1 ratio composition, average molecular weight is about 9279D, has the skeleton of (1 4) β D glucose, one → 3 are connected on 6 O) side chain of α D-galactolipin, the newborn mushroom polysaccharide of perfume of doses has significant oxidation resistance, to the Scavenging activity of DPPH free radicals, its IC50It is worth for 0.365mg/mL；To the Scavenging activity of ABTS free radicals, its IC50Value is about 1.599mg/mL, and dosage is bigger, and effect is better.
Description
Technical field
The invention belongs to fungi polysaccharide applied technical field, specifically, is related to one kind perfume breast mushroom polysaccharide LC-1 and its system
Preparation Method and application.
Background technology
Polysaccharide (polysaccharides) is also known as polysaccharide, is a kind of large biological molecule of organism generally existing, to thin
The composition of born of the same parents' skeleton composition and a variety of endogenous bioactive molecules plays the role of important.Polysaccharide plays huge in vivo
Effect, if cellulose and chitin are that cyto-architectural important substance is maintained in organism, as starch and glycogen are to provide energy
Measure the material in source.At present, the research level of polysaccharide will lag behind the research of protein and nucleic acid significantly, mainly due to polysaccharide
Development it is later, plus the complexity and diversity of polysaccharide in itself, huge choose is proposed to polysaccharide structures measure, characterization identification
War.
In the past 20 years, the research to polysaccharide is mainly concentrated in bioactivity, especially immunological regulation, antitumor, anti-
Viral, anti-oxidant and hypoglycemic etc., its effect is multipath, too many levels, Mutiple Targets.
Fragrant breast mushroom, Classification system is Lactarius camphoratum (Bull.) Fr., its fructification is small, bacteria cover diameter
2-5cm, initial stage oblate spheroid, after it is gradually recessed, often there is kick at middle part, does not glue, deep cinnamon to birchleaf pear color.Bacterial context color is shallower than bacterium
Lid.Milk white is constant.Lamella white to faint yellow, it is old after color it is similar to cap, close, growing straight to slightly downward, the nearly long 2- of handle
5cm, thick 0.4-0.8cm, nearly cylindricality, color is similar to cap, internal soft, rear hollow.Spore print milky.Spore is colourless, closely
It is spherical, there are wart and reticulate pattern, 6.4-8 μm of 7.3-9 μ ms.Pleat side utricule fusiformis, tool point, 7.3-10.9 μm of 60-90 μ ms.Summer and autumn
In woods on the ground, scattered or all living creatures.
At present, the fine structure of fragrant newborn mushroom polysaccharide LC-1 is studied with antioxidation activity and its is applied there is not yet any report
Road.
The content of the invention
In view of this, the present invention provides a kind of fragrant newborn mushroom polysaccharide LC-1 and preparation method and application.
In order to solve the above-mentioned technical problem, the invention discloses a kind of fragrant newborn mushroom polysaccharide LC-1, fragrant breast mushroom polysaccharide is by two
The heteroglycan of a monose composition, i.e. β-D-Glucose and α-D- galactolipins is with 2:1 ratio composition, average molecular weight are about
9279D, has the skeleton of (1-4)-β-D-Glucose, and one → 3 are connected on 6-O) side chains of-α-D- galactolipins, fragrant breast mushroom is more
Sugar structural formula be：
The invention also discloses a kind of preparation method of fragrant newborn mushroom polysaccharide LC-1, comprise the following steps：
Step 1, mix fragrant the breast massee fruiting bodies powder and distilled water after crushing and boil in a water bath, collects supernatant dense
Contracting, is prepared concentrate；
Step 2, add absolute ethyl alcohol formation flocky precipitate in concentrate, collects and precipitates simultaneously drying, obtains fragrant breast mushroom
Polysaccharide LC-1 crude extracts；
Step 3, isolated and purified with chromatographic column, obtains fragrant newborn mushroom polysaccharide LC-1.
Further, the volume ratio of the newborn massee fruiting bodies powder of perfume and distilled water after the crushing in step 1 is 1：2-5.
Further, the temperature of the water-bath in step 1 is 100 DEG C；When boiling time is 2-5 small, number is boiled for 6 times.
Further, the volume ratio of the concentrate in step 2 and absolute ethyl alcohol is 1：2-1：6.
Further, the chromatographic column isolated and purified in step 3 is DEAE-cellulose column and Sephadex
G-100column。
Compared with prior art, the present invention can be obtained including following technique effect：
The newborn mushroom polysaccharide of perfume of doses has significant oxidation resistance, to the Scavenging activity of DPPH free radicals, its IC50
It is worth for 0.365mg/mL；To the Scavenging activity of ABTS free radicals, its IC50Value is about 1.599mg/mL, and dosage is bigger, and effect is got over
It is good.
Certainly, implement any of the products of the present invention and it is not absolutely required to reach all the above technique effect at the same time.
Brief description of the drawings
Attached drawing described herein is used for providing a further understanding of the present invention, forms the part of the present invention, this hair
Bright schematic description and description is used to explain the present invention, does not form inappropriate limitation of the present invention.In the accompanying drawings：
The fragrant newborn mushroom holosaccharide LC-1 weight average molecular weight of Fig. 1；
The INFRARED SPECTRUM of the fragrant newborn mushroom polysaccharide LC-1 of Fig. 2；
The 1H NMR spectras of the fragrant newborn mushroom polysaccharide LC-1 of Fig. 3；
The 13C NMR spectras of the fragrant newborn mushroom polysaccharide LC-1 of Fig. 4；
The structure of the fragrant newborn mushroom polysaccharide LC-1 of Fig. 5；
Scavenging actions of Fig. 6 perfume breast mushroom polysaccharide LC-1 to DPPH free radicals；
Scavenging actions of Fig. 7 perfume breast mushroom polysaccharide LC-1 to ABTS free radicals.
Embodiment
Carry out the embodiment that the present invention will be described in detail below in conjunction with embodiment, thereby to the present invention how application technology hand
Section solves technical problem and reaches the process of realizing of technical effect to fully understand and according to this implement.
The preparation method of the fragrant newborn mushroom polysaccharide LC-1 of embodiment 1
The newborn massee fruiting bodies powder of perfume and distilled water after crushing is with 1：3.6 volume ratio boils 3.5 in 100 DEG C of water-bath
Hour, boil 6 times, collect supernatant and be concentrated into 150ml.The absolute ethyl alcohol for adding 4 times of volumes forms flocky precipitate, and it is heavy to collect
Form sediment and dry, obtain fragrant newborn mushroom polysaccharide LC-1 crude extracts.With DEAE-cellulose column and Sephadex G-
100column is isolated and purified, and obtains fragrant newborn mushroom polysaccharide LC-1.
Embodiment 2
The newborn massee fruiting bodies powder of perfume and distilled water after crushing is with 1：It is small that 2 volume ratio boils 2 in 100 DEG C of water-bath
When, boil 6 times, collect supernatant and be concentrated into 150ml.The absolute ethyl alcohol for adding 2 times of volumes forms flocky precipitate, collects precipitation
And it is dry, obtain fragrant newborn mushroom polysaccharide LC-1 crude extracts.With DEAE-cellulose column and Sephadex G-
100column is isolated and purified, and obtains fragrant newborn mushroom polysaccharide LC-1.
Embodiment 3
The newborn massee fruiting bodies powder of perfume and distilled water after crushing is with 1：It is small that 5 volume ratio boils 5 in 100 DEG C of water-bath
When, boil 6 times, collect supernatant and be concentrated into 150ml.The absolute ethyl alcohol for adding 2 times of volumes forms flocky precipitate, collects precipitation
And it is dry, obtain fragrant newborn mushroom polysaccharide LC-1 crude extracts.With DEAE-cellulose column and Sephadex G-
100column is isolated and purified, and obtains fragrant newborn mushroom polysaccharide LC-1.
The Structural Identification of the fragrant newborn mushroom polysaccharide LC-1 of embodiment 4
With hydrolysis, methylation analysis, gas-chromatography and mass spectrometric hyphenated technique (GC-MS), scanning electron-microscopy (SEM),
Infrared spectral technology (IR), nuclear magnetic resonance technique (NMR), structure elucidation is carried out to fragrant newborn mushroom polysaccharide LC-1 sterlings polysaccharide.
1st, the measure of molecular weight
The fragrant newborn mushroom polysaccharide LC-1 samples of 3mg are dissolved in 1mL ddH2In O, ultrasonic 5min, carries out HPGPC analyses, data
With after GPC software analysis molecular weight 1.1 × 10 is measured with standard curve control4D (standard items T-10,20,50,80,130).
Fragrant breast mushroom holosaccharide weight average molecular weight is 9279D (see Fig. 1).
2nd, the infrared spectrum analysis of fragrant newborn mushroom polysaccharide LC-1
As fragrant newborn mushroom polysaccharide LC-1 samples 2mg or so, KBr tablettings, infrared spectrophotometer 4000cm-1-400cm-1。
As shown in Fig. 2, fragrant breast mushroom polysaccharide shows 3428.78cm in INFRARED SPECTRUM-1Wide absworption peak be appointed as O-H and N-H
Stretching vibration peak, there is intramolecular and intermolecular hydrogen bond.2926.89cm-1It is appointed as-CH2, the stretching vibration peak of-CH3.
2127.55cm-1It is appointed as-CH2,-CH3Stretching vibration peak.1651.82cm-1It is appointed as C=O, C=C vibration peaks.
1124.92cm-1, 1082.90cm-1It is for ehter bond C-O-C stretching vibrations in C-O-H stretching vibrations and pyranoid ring respectively.
791.10cm-1The fragrant newborn mushroom polysaccharide of instruction has α type pyranoid rings.
3rd, the nuclear magnetic resonance spectroscopy of fragrant newborn mushroom polysaccharide LC-1
Fragrant newborn mushroom polysaccharide LC-1 samples 10mg or so is taken to be dissolved in 0.5mL heavy water (D2O in), room temperature is surveyed in Nuclear Magnetic Resonance
Fixed, 400MHz is surveyed1HNMR is composed, and 400MHz is surveyed13CNMR is composed.
Anomer hydrogen signal δ 4.865, δ 4.854 indicate that α types anomer hydrogen exists (see Fig. 3) in fragrant newborn mushroom polysaccharide, and both
Shown with INFRARED SPECTRUM consistent.In carbon spectrum13The resonance signal peak in 95-105ppm regions of C NMR (400MHz) belong to β-
The carbonaceous subsignal of D-Glucose and α-D- galactolipins (see Fig. 4, table 1).
The fragrant newborn mushroom polysaccharide LC-1's of table 113Chemical shift in C NMR figures
4th, the Silylation and methylation analysis of fragrant newborn mushroom polysaccharide LC-1
It is as follows that hydrolysate after standard monose and above-mentioned drying is subjected to Silylation step：The sample dissolving of 1.0mg
In the anhydrous pyridine of 0.2mL, then add the hexamethyldisilazane of 0.2mL and the trim,ethylchlorosilane of 0.1mg, 50 DEG C of thermostatted waters
20min, supercentrifuge 10000rpm centrifugation 20min are educated, supernatant liquor is drawn and adds sample injection bottle progress methylation analysis.
5th, fragrant newborn mushroom polysaccharide LC-1GC-MS spectrum analysis
GC conditions：Chromatographic column：Rtx-5sil MS (the μ of 5%phenylmethylsiloxane30mm × 0.25mm × 0.25
m)；
Injector temperature, GC-MS interface temperatures, ion source temperature and detector temperature are respectively
Carrier gas：High-purity helium.
The novel heteroglycan being made of two monose of a structure, i.e. β-D-Glucose and α-D- are obtained from fragrant newborn mushroom
Galactolipin is with 2:1 ratio composition, average molecular weight is about 9279D, has the skeleton of (1-4) β-D-Glucose, is connected on 6-O
One → 3) side chain (table 2) of-α-D- galactolipins, to sum up can tentatively infer the structure of fragrant newborn mushroom polysaccharide (see Fig. 5).
The GC-MS data that methylate of the fragrant newborn mushroom polysaccharide of table 2
The antioxidation activity research of the fragrant newborn mushroom polysaccharide LC-1 of embodiment 5
In vitro with the antioxidation activity of the fragrant newborn mushroom polysaccharide LC-1 holosaccharides of two kinds of chemical gaugings, including DPPH-
The removing of free radical, ABTS+Radical cation is removed.
1 reagent
1,1- diphenyl -2- picryls hydrazines (DPPH-)；30%H2O2；Vitamin C (Vc)；2,6- di-t-butyl -4- methylbenzenes
Phenol (BHT)；ABTS
2 instruments
721 type spectrophotometers (the 3rd analytical instrument factory of Shanghai)；
3 methods
3.1DPPH radical scavenging activities measure
DPPH radical scavenging activities measure with reference to Braca methods, sample with distilled water be configured to suitable concentration (1,2,3,
4,5,6mg/mL).1mL solution to be measured is taken, 2.0mL 0.2mM DPPH- solution (being prepared with absolute ethyl alcohol) is added, is put after shaking up
Put 30min.Light absorption value of the spectrophotometric determination A samples at 517nm.A controls are 1.0mL sample solutions and 2.0mL ethanol
Mixed liquor.Positive control is used as using Vc and BHT.DPPH Scavenging activities=(1-A samples/A controls) × 100%.
3.2ABTS+Radical cation Scavenging activity measures
7mmol/L ABTS aqueous solutions and the reduction of 2.45mmol/L potassium peroxydisulfates lucifuge 12-16h, ABTS solution PBS
(pH7.4) dilute, until 630nm measured values are 0.50 ± 0.05.10 μ L samples, 90 μ LABTS solution hybrid reactions are taken, are placed in dark
Locate 3min.By the use of sample P BS solution as color sample blank reference, PBS solution is blank control, in 734nm in microplate reader
Place's detection.Clearance rate is calculated according to following equation：ABTS clearance rates (%)=[A blank-(A samples-A controls)]/A blank ×
100%
4 results
4.1. perfume breast mushroom polysaccharide LC-1 is to DPPH- free radical scavenging activities
Fig. 6 illustrates that the newborn more DPPH free radicals of mushroom polysaccharide sample of the perfume of purifying have good scavenging capacity, it eliminates DPPH
The IC50 values of free radical are about 0.365mg/mL, and dosage is bigger, and effect is better.
4.2. the activity that perfume breast mushroom polysaccharide LC-1 removes ABTS+ radical cations
The activity removed using the fragrant newborn mushroom polysaccharide of measure of the spectrophotometer at 734nm to ABTS+ radical cations.
The result shows that (Fig. 7), the IC50 values that fragrant breast mushroom polysaccharide eliminates ABTS+ radical cations are about 1.599mg/mL, and dosage is bigger,
Effect is better.Some preferred embodiments of invention have shown and described in described above, but as previously described, it should be understood that invention is not
Form disclosed herein is confined to, is not to be taken as the exclusion to other embodiment, and available for various other combinations, modification
And environment, and can be carried out in the scope of the invention is set forth herein by the technology or knowledge of above-mentioned teaching or association area
Change., then all should be in power appended by invention and changes and modifications made by those skilled in the art do not depart from the spirit and scope of invention
In the protection domain that profit requires.
Claims (7)
1. a kind of fragrant newborn mushroom polysaccharide LC-1, it is characterised in that the heteroglycan that fragrant breast mushroom polysaccharide is made of two monose, i.e. β-D-
Glucose and α-D- galactolipins are with 2:1 ratio composition, average molecular weight is about 9279D, has (1-4)-β-D-Glucose
Skeleton, one → 3 are connected on 6-O) side chains of-α-D- galactolipins, the structural formula of fragrant breast mushroom polysaccharide is：
2. a kind of preparation method of fragrant newborn mushroom polysaccharide LC-1, it is characterised in that comprise the following steps：
Step 1, mix fragrant the breast massee fruiting bodies powder and distilled water after crushing and boil in a water bath, collects supernatant concentration,
Concentrate is prepared；
Step 2, add absolute ethyl alcohol formation flocky precipitate in concentrate, collects and precipitates simultaneously drying, obtains fragrant breast mushroom polysaccharide
LC-1 crude extracts；
Step 3, isolated and purified with chromatographic column, obtains fragrant newborn mushroom polysaccharide LC-1.
3. preparation method according to claim 2, it is characterised in that newborn mushroom of perfume after the crushing in the step 1 is real
The volume ratio of body powder and distilled water is 1：2-5.
4. preparation method according to claim 2, it is characterised in that the temperature of the water-bath in the step 1 is 100 DEG C；
When boiling time is 2-5 small, number is boiled for 6 times.
5. preparation method according to claim 2, it is characterised in that concentrate and absolute ethyl alcohol in the step 2
Volume ratio is 1：2-1：6.
6. preparation method according to claim 2, it is characterised in that the chromatographic column isolated and purified in the step 3 is
DEAE-cellulose column and Sephadex G-100 column.
A kind of 7. applications of perfume breast mushroom polysaccharide LC-1 in anti-oxidation medicine is prepared.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110229244A (en) * | 2019-06-26 | 2019-09-13 | 西华师范大学 | A kind of Ramaria stricta polysaccharide and its preparation method and application |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104686654A (en) * | 2014-08-14 | 2015-06-10 | 深圳市大百汇技术有限公司 | Lactarius volemus polysaccharide yoghourt and preparation method thereof |
-
2017
- 2017-11-27 CN CN201711205932.XA patent/CN107987175A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104686654A (en) * | 2014-08-14 | 2015-06-10 | 深圳市大百汇技术有限公司 | Lactarius volemus polysaccharide yoghourt and preparation method thereof |
Non-Patent Citations (1)
| Title |
|---|
| 王芳: ""香乳菇多糖（LC-1）的分离纯化、结构鉴定以及生物活性的研究"", 《中国优秀硕士学位论文全文数据库 医药卫生科技辑》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110229244A (en) * | 2019-06-26 | 2019-09-13 | 西华师范大学 | A kind of Ramaria stricta polysaccharide and its preparation method and application |
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