CN105884925A - Pumpkin peel polysaccharide and preparation method and application thereof - Google Patents

Pumpkin peel polysaccharide and preparation method and application thereof Download PDF

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Publication number
CN105884925A
CN105884925A CN201610259947.3A CN201610259947A CN105884925A CN 105884925 A CN105884925 A CN 105884925A CN 201610259947 A CN201610259947 A CN 201610259947A CN 105884925 A CN105884925 A CN 105884925A
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pumpkin peel
polysaccharide
pumpkin
peel polysaccharide
water
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李伟光
冯昇
钱景怡
王诗豪
田欣
魏怡超
王森怡
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China Pharmaceutical University
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China Pharmaceutical University
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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
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/715—Polysaccharides, i.e. having more than five saccharide radicals attached to each other by glycosidic linkages; Derivatives thereof, e.g. ethers, esters
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/42—Cucurbitaceae (Cucumber family)
    • 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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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Natural Medicines & Medicinal Plants (AREA)
  • Molecular Biology (AREA)
  • Engineering & Computer Science (AREA)
  • Biochemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Polymers & Plastics (AREA)
  • Organic Chemistry (AREA)
  • Materials Engineering (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Veterinary Medicine (AREA)
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  • Animal Behavior & Ethology (AREA)
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  • Alternative & Traditional Medicine (AREA)
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  • Medical Informatics (AREA)
  • Botany (AREA)
  • Biotechnology (AREA)
  • Sustainable Development (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Medicines Containing Plant Substances (AREA)

Abstract

The invention relates to botanical pumpkin peels, in particular to pumpkin peel polysaccharide and a preparation method and application thereof. The pumpkin peel polysaccharide is characterized in that polysaccharide content is higher than 90%, and number-average molecular weight is 2000-3000KDa; the polysaccharide is composed of glucose, galactose, mannose and xylose. In a pumpkin peel polysaccharide preparation process, proteins are removed more thoroughly through purification by chloroform-butanol solution and ion exchange resin; by further refining through molecular sieve gel columns, obtained pumpkin peel polysaccharide is low in impurity content and high in purity; according to hypoglycemic activity of the pumpkin peel polysaccharide and pumpkin peel extracts, a new idea is provided for application of the pumpkin peel polysaccharide. By recycling of pumpkin peels which are wastes, the pumpkin peel polysaccharide and the preparation method and application thereof have promising application prospect and high social benefits and economic benefits.

Description

Pumpkin peel polysaccharide and preparation method thereof, application
Technical field
The present invention relates to plant amedica pumpkin peel, be exactly in pumpkin peel polysaccharide and pumpkin peel extract for treating diabetic medicine thereof Application.
Background technology
Diabetes are the most universal a kind of endocrine and metabolic disorders diseases, be insulin secretion and Use barriers causes with high blood The chronic disease that sugar is characterized.Onset diabetes rate increases year by year, it has also become global outstanding problem.Type i diabetes, former Name insulin dependent diabetes mellitus (IDDM), mostly occurs at Children and teenager, it is possible to betide the various age.Onset compares drastically, body Interior insulin is the most not enough, is susceptible to ketoacidosis, it is necessary to could obtain satisfactory effect with insulin treatment, otherwise will danger And life.
One kind of Fructus Cucurbitae moschatae (Cucurbita moschata (Duch.ex Lam.) Duch.ex Poiret) Cucurbitaceae Cucurbita, 1 year The raw herbaceous plant that overgrows.Originate in Mexico to carry to Central America one, the most generally cultivate.Incoming China of the Ming Dynasty, existing north and south is each Ground extensively plantation.The fruit of Fructus Cucurbitae moschatae can make sumptuous courses at a meal, also can be for grain.The each portion of Herb hyoscine again, and effect is extensively, middle its people Between also spread the saying of " eating Fructus Cucurbitae moschatae blood sugar lowering ".There are some researches show that in pumpkin pulp, squash polyoses can treat type i diabetes, but Pumpkin peel is not used by people as garbage.The present invention uses the type i diabetes mouse model with STZ abductive approach right for research As, from pumpkin peel, extract squash polyoses and investigate its function of reducing blood sugar to type i diabetes mice, proposing new for pumpkin peel application Thinking, thus turn waste into wealth.
Summary of the invention
The object of the invention provides the application in preparation treatment diabetes medicament of a kind of pumpkin peel polysaccharide and pumpkin peel extract.
Technical scheme
Pumpkin peel polysaccharide, it is characterised in that its polyoses content more than 90%, number-average molecular weight 2000～3000KDa；Monosaccharide group Become to include glucose, galactose, mannose, xylose；Glycosidic bond of bonding has α or β key；
The preparation method of described pumpkin peel polysaccharide, is characterized in that being made up of the following step:
A, acquisition crude extract: the pumpkin peel that will fetch is dried, and pulverizes, adds 1-10 times of volume distilled water, 45 DEG C water Bathe 3-5 hour, filter, concentrate；
B, remove removing protein: described concentrated solution mixes with chloroform-n-butyl alcohol 4:1 by volume, and machine is centrifuged 1min by centrifugation, Rotating speed is 5000rpm/min, removes removing protein, retains lower floor's solution, wherein said chloroform-butanol solution be chloroform with just The solution that butanol is mixed to form for 4:1 by volume；Repeat said process three times, concentrate, lyophilizing, it is thus achieved that pumpkin peel extract Thing；
C, acquisition pumpkin peel polysaccharide crude: take pumpkin peel extract and be dissolved in the water of 20-50 times of weight, add D301-G macropore tree Fat, intermittent stirring 1-3 hour under room temperature, after concentration, 4000r.p.m is centrifugal goes precipitation, adds 1-6 times of volume in supernatant Dehydrated alcohol, place 24 hours, be centrifuged off supernatant for 4 DEG C, precipitate three times with absolute ethanol washing, dried Fructus Cucurbitae moschatae Skin polysaccharide crude；
D, obtain pumpkin peel polysaccharide: described pumpkin peel polysaccharide crude, be dissolved in water, upper DEAE-52 anion-exchange column, With 1mol/L NaCl solution eluting after deionized water eluting, detect with Phenol-sulphate acid method, collect NaCl solution eluting peak, decompression Concentrating, flowing water is dialysed 24 hours, precipitates through 4 times of ethanol, 4 DEG C of placements after concentration, is centrifuged and removes supernatant, with absolute ethanol washing Three times, vacuum drying；Sample after vacuum drying is dissolved in deionized water, on the Sephacryl S-400 molecular sieve that balanced Gel column, is washed with deionized water de-, fraction collection, detects with Phenol-sulphate acid method, collects eluting peak, through lyophilizing after concentration, obtains Obtain pumpkin peel polysaccharide.
Described pumpkin peel polysaccharide purposes in preparation treatment diabetes medicament.
A kind of pumpkin peel extract purposes in preparation treatment diabetes medicament, it is characterised in that described pumpkin peel extract is In claim 1, step B prepares.
Beneficial effect
The present invention, during preparing pumpkin peel polysaccharide, is respectively adopted chloroform-butanol solution method and ion exchange resin carries out pure The method changed, goes removing protein more complete, refines further through molecular sieve gel post, the pumpkin peel polysaccharide of gained, impurity the most again Few, purity is high；By the molecular weight of pumpkin peel polysaccharide, purity, monosaccharide composition, glycosidic bond on-link mode (OLM) etc. are carried out structural table Levy, it is thus achieved that the detailed structural information of pumpkin peel polysaccharide；And to pumpkin peel polysaccharide and the hypoglycemic activity of pumpkin peel extract, for The application of pumpkin peel polysaccharide provides new thinking.Pumpkin peel, as garbage, can be turned waste into wealth, gather around and have broad application prospects, There is good Social benefit and economic benefit.
Detailed description of the invention
Embodiment 1:
The concrete preparation method of pumpkin peel polysaccharide of the present invention is as follows:
1, the pumpkin peel that fetch is dried, dry after its weight be called 117.6 grams, then smashed to pieces, added distilled water, 45 C water bath 3 hours.Filtrate is obtained by filtered through gauze.With Rotary Evaporators to its evaporation and concentration.
2, concentrated solution is carried out severge method Deproteinization；Detailed process is: crude polysaccharides solution (is prepared in advance with chloroform n-butyl alcohol Becoming volume ratio is 4: 1 mixed liquors) ratio of 4:1 by volume, it is placed in tool plug test tube, after shake well 30min, by centrifugation Machine is centrifuged 1min, and rotating speed is 5000r/min, is then separated mutually with chloroform by aqueous phase.Aqueous phase is added and is equivalent to its volume 1/4 Chloroform butanol solution, repeat said process, amount to the most in triplicate, concentrate, lyophilizing, it is thus achieved that pumpkin peel extract；
3, take pumpkin peel water extract 2g in container, be completely dissolved in 100mL deionized water, add D301-G macroporous resin Decolouring and isolating protein, or stirring 3 hours, solution is concentrated into 10mL after filtering, and adds 60mL dehydrated alcohol precipitation, Place 24 hours for 4 DEG C.It is centrifuged off supernatant.With absolute ethanol washing 3 times, vacuum drying obtains pumpkin peel polysaccharide Decolorization product；
4, take pumpkin peel polysaccharide Decolorization product 200mg, be completely dissolved in deionized water, hand over the DEAE-52 ion balanced Changing post to separate, specification is (2.6 × 30cm), and balance liquid is deionized water, and loading volume is 10mL, first goes with 300mL Ionized water eluting, then the NaCl solution eluting with 1mol/L, fraction collection, Phenol-sulphate acid method tracing detection, merge identical group Point.
(4) ion exchange column separating obtained NaCl solution elution fraction carries out Sephacryl S-400 molecular sieve column chromatography, to go Ionized water eluting.Post specification is (1.0 × 100cm), fraction collection, Phenol-sulphate acid method tracing detection, merges same composition. Lyophilization, obtains pumpkin peel polysaccharide.
Purity and the molecular weight of pumpkin peel polysaccharide are measured by efficient molecular exclusion chromatography: with Shodex SUGAR KS-805 (8mmID × 300mm) is performance liquid chromatographic column, uses differential refraction detector；Chromatographic condition is: H2O is flowing phase, Flow velocity 1mL/min, sample concentration is 1mg/mL, each sample introduction 20 μ L, detects time 20min.Obtain pumpkin peel polysaccharide Mean molecule quantity is 2880KDa.
The infrared spectrum of pumpkin peel polysaccharide, wave number 3442cm-1Strong absworption peak be the stretching vibration of O-H, 2933cm-1Absorb Peak is sugar ring C-H stretching vibration, 1000-1200cm-1Between a series of absworption peaks be by the stretching vibration of C-O on sugar ring, 1743 cm-1,1425cm-1And 1257cm-1Absworption peak be the C=O stretching vibration of aldehydic acid group respectively, C-O stretching vibration and O-H Stretching vibration, 855cm-1And 895cm-1Absworption peak show that existing α configuration has again the sugared unit of beta comfiguration.
The monosaccharide composition of pumpkin peel polysaccharide products passes through pre column Derivatization: with 2mol/L trifluoroacetic acid water Solving, be evaporated after methanol washing, with deionized water dissolving, hydrolyzed solution carries out high-efficient liquid after PMP derivatization and detects mutually；Chromatograph Condition: chromatographic column is waters synmetry C18 (150mm × 4.6mm), flowing is ammonium acetate (pH5.5)/acetonitrile (70/30) mutually, Detection wavelength is 250nm.Shown in result, calculate according to retention time and peak area, and with monosaccharide standard substance carry out right According to, it is known that pumpkin peel polysaccharide contains glucose, galactose, mannose, xylose.
Embodiment 2
1, animal: provided experiment kunming mice by Qinglongshan animal cultivation field, Nanjing, adaptability is fed two weeks, packet numbering.
(because STZ easily distils, whole process for preparation should operate on ice chest STZ reagent (Sigma Co., USA), the most existing With now joining, it is finished in preparing rear 30min).Blood sugar test paper (Johnson Co.'s surely person of outstanding talent times easily type reagent paper) is (because STZ easily rises China, whole process for preparation should operate on ice chest, and usual matching while using is finished in preparing rear 30min).
Blood glucose meter (Johnson Co.'s surely person of outstanding talent times benefit type blood glucose meter)
2, method:
(1) randomly drawing 5 mices is Normal group (only injecting isopyknic normal saline), and remaining mice fasting is not Prohibit water and after 12 hours, use STZ method inducing type I diabetes model, inject to mouse peritoneal with 150mg/kg dosage, then Recovering feed, survey its fasting glucose after three days, blood glucose value is more than 11.6mmol/L then modeling success.Rear random taking-up 5 is Negative control group (only injection equal-volume normal saline), then residue mice is often organized 5, packet；It is respectively pumpkin peel to extract Thing 300mg/kg, 600mg/kg, 900mg/kg, pumpkin peel polysaccharide 50mg/kg, 100mg/kg, 150mg/kg and sun Property matched group (0.5U/ml insulin, with 0.054ml/10g dosed administration), result is as follows:
Squash polyoses and the insulin medicament impact on blood glucose in diabetic mice in table 1 pumpkin peel
Note: " * " compares P ＜ 0.05 with diabetes model, " * * " compares P ＜ 0.01 with diabetes model
Illustrate: pumpkin peel polysaccharide of the present invention or pumpkin peel extract, compared with negative control group (i.e. model group), have dosage and depend on The relation of relying, simultaneously by data, high dose and positive controls (insulin is suitable).Find out from above-mentioned data, Fructus Cucurbitae moschatae Skin polysaccharide is the main active of pumpkin peel extract.Meanwhile, for oral, its dosage is the least, is suitable for exploitation and protects Strong product or medicine.

Claims (4)

1. pumpkin peel polysaccharide, it is characterised in that its polyoses content more than 90%, number-average molecular weight 2000 ~ 3000KDa；Monosaccharide composition includes glucose, galactose, mannose, xylose.
The preparation method of pumpkin peel polysaccharide the most according to claim 1, is characterized in that being made up of the following step:
A, acquisition crude extract: the pumpkin peel that will fetch is dried, and pulverizes, add 1-10 times of volume distilled water, 45 C water bath 3-5 hour, filter, concentrate；
B, remove removing protein: described concentrated solution mixes with chloroform-n-butyl alcohol 4:1 by volume, machine is centrifuged 1min by centrifugation, and rotating speed is 5000rpm/min, removes removing protein, retaining lower floor's solution, wherein said chloroform-butanol solution is the solution that chloroform is mixed to form for 4:1 by volume with n-butyl alcohol；Repeat said process three times, concentrate, lyophilizing, it is thus achieved that pumpkin peel extract；
C, acquisition pumpkin peel polysaccharide crude: take pumpkin peel extract and be dissolved in the water of 20-50 times of weight, add D301-G macroporous resin, intermittent stirring 1-3 hour under room temperature, after concentration, 4000r.p.m is centrifugal goes precipitation, in supernatant, add the dehydrated alcohol of 1-6 times of volume, place 24 hours, be centrifuged off supernatant for 4 DEG C, precipitate three times with absolute ethanol washing, dried pumpkin peel polysaccharide crude；
D, acquisition pumpkin peel polysaccharide: described pumpkin peel polysaccharide crude, be dissolved in water, upper DEAE-52 anion-exchange column, with 1mol/L NaCl solution eluting after deionized water eluting, detect with Phenol-sulphate acid method, collect NaCl solution eluting peak, concentrating under reduced pressure, flowing water is dialysed 24 hours, precipitate through 4 times of ethanol after concentration, 4 DEG C of placements, be centrifuged and remove supernatant, with absolute ethanol washing three times, vacuum drying；Sample after vacuum drying is dissolved in deionized water, on the Sephacryl S-400 molecular sieve gel post that balanced, be washed with deionized water de-, fraction collection, detect with Phenol-sulphate acid method, collect eluting peak, through lyophilizing after concentration, it is thus achieved that pumpkin peel polysaccharide.
The pumpkin peel polysaccharide the most according to claim 1 purposes in preparation treatment diabetes medicament.
4. a pumpkin peel extract is preparing the purposes treated in diabetes medicament, it is characterised in that described pumpkin peel extract is that in claim 1, step B prepares.
CN201610259947.3A 2016-04-25 2016-04-25 Pumpkin peel polysaccharide and preparation method and application thereof Pending CN105884925A (en)

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116606385A (en) * 2023-04-28 2023-08-18 广州工程技术职业学院 Zedoary turmeric polysaccharide and preparation method and application thereof

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102603907A (en) * 2012-01-17 2012-07-25 中国药科大学 Glabrous sarcandra herb polysaccharide and preparation method and application thereof

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102603907A (en) * 2012-01-17 2012-07-25 中国药科大学 Glabrous sarcandra herb polysaccharide and preparation method and application thereof

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
崔亦华: "应用广泛的天然多糖及其提取方法", 《广州化工》 *
白娣斯等: "南瓜皮多糖PP1的提纯、组分分析及原子力显微镜技术观测研究", 《食品科学》 *
赵大庆等: "南瓜水溶性粗多糖提取工艺的优化", 《农产品加工.学刊》 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116606385A (en) * 2023-04-28 2023-08-18 广州工程技术职业学院 Zedoary turmeric polysaccharide and preparation method and application thereof
CN116606385B (en) * 2023-04-28 2024-04-16 广州工程技术职业学院 Zedoary turmeric polysaccharide and preparation method and application thereof

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Application publication date: 20160824