CN102037962A - Chitosan nano-biological preparation, and preparation method and application thereof - Google Patents
Chitosan nano-biological preparation, and preparation method and application thereof Download PDFInfo
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- CN102037962A CN102037962A CN2010105636049A CN201010563604A CN102037962A CN 102037962 A CN102037962 A CN 102037962A CN 2010105636049 A CN2010105636049 A CN 2010105636049A CN 201010563604 A CN201010563604 A CN 201010563604A CN 102037962 A CN102037962 A CN 102037962A
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Abstract
The invention discloses a chitosan nano-biological preparation, and a preparation method and application thereof. The preparation method comprises the following steps of: dissolving chitosan in 1 volume percent acetic acid solution to keep the concentration of the chitosan at 2mg/ml; adjusting pH value to be between 4.6 and 4.7 by using 10mol/L sodium hydroxide; degrading for 4 to 8 hours to obtain chitosan acetic acid solution; dropwise adding 1mg/mL of sodium tripolyphosphate solution into the chitosan acetic acid solution, wherein the mass ratio of the chitosan acetic acid solution to the sodium tripolyphosphate solution is 5 to 1; magnetically stirring for 30 minutes; performing pulse ultrasonic processing for 15 minutes in an ice bath by using 85 percent of power; and opening the ultrasonic sound for 3 seconds every 2 seconds to prepare the cross-linked and uniformly dispersed chitosan nano-biological preparation. Nano-chitosan is used as a novel plant growth adjusting substance which can induce a plant to secrete resistant enzyme, excite expression of a resistant gene and effectively improve the resistance of the plant against adverse environmental stress.
Description
Technical field
The present invention relates to chitosan nano biologic product and preparation thereof.
Background technology
Shitosan is one of most important derivative behind the de-acetyl chitin.Chitin (Chitin) has another name called chitin, chitin, is a kind of glycosaminoglycan polymer, and chemical name is β-(1,4)-2-acetylaminohydroxyphenylarsonic acid 2-deoxy-D-glucose, and molecular formula is (C
8H
15NO
6)
n, molecular weight is about 1,000,000.Chitin is the positively charged natural biological macromolecular material of the unique existence of occurring in nature, extensively be present in lower animal such as crusts such as crab, shrimp, in the shell of the first skin of insects such as cricket, male whetstone etc. and in the cell wall of some rudimentary plants such as algae, fungi, resource is very abundant, being to be only second to cellulosic second largest class natural polymer in the world, is a kind of inexhaustible renewable resources.
The molecular structural formula of chitin is as follows:
Chitin contains-NH
2Base ,-the OH base; be easy to modify; can generate the different various derivatives of performance; derivative shitosan (chitosan) behind de-acetyl chitin is an alkaline polysaccharide unique in the natural polysaccharide; have the physicochemical characteristics and the biological function of many uniquenesses, can be widely used in fields such as environmental project, weaving, papermaking, food, medical medicine, biotechnology, cosmetics and agrotechnique.In recent years, be subjected to along with its plant growth regulator effect that in agricultural production, manifests and plant pest management immunity protection function paying attention to widely.
At present, all kinds of chitins and the derived product producing and use, its feature mainly is that most products are common size chitin and derivative or its elementary modified product, another one is that common size chitin and derivative are also only as medicinal compatibility material, the chemical property of not fine demonstration chitin and biologically active, limited the range of application and the effect of shitosan to a certain extent, therefore need to its carry out form and
Structural modification, enlarging its range of application and function, and modern nanometer technology is the important channel that addresses the above problem.
Nanometer technology is the structure of research material on 1～100nm yardstick and the cutting edge technology of character, and its final goal is the product that has specific function with the characteristic manufacturing on nanoscale of molecule, atom and material, realizes the revolution of the mode of production.Have unique effect-skin effect, bulk effect and macro quanta tunnel effect as nano material, thereby nano material on the performance with traditional concept on the stock size of same composition or the difference that the micron-scale material has highly significant, show the characteristic of many excellences and brand-new function.In recent years, nanometer technology to the infiltration of high-tech areas such as biological medicine, information, the energy and environment, ocean, national defence and modern agriculture, demonstrates its application and stronger market potential widely.
Nanometer technology combines with biotechnology, and is applied to biological field, has just formed a kind of new multidisciplinary interleaving techniques--nanometer biotechnology.Emerging field that develops just gradually of nanometer biotechnology, and form a new subject gradually.The research range of nanometer biotechnology relates generally to fields such as nano meter biomaterial, Nano medication and gene delivery nano-carrier, nano biological sensor and nano biological preparation.
Because it is big that general shitosan is difficult for dissolving, viscosity height, particle, difficultly directly absorbed by plant, make chitin run into bottleneck in the plant growth regulating application facet.
Summary of the invention
The present invention makes the nano biological preparation of chitin by combined methods such as ionomer, magnetic agitation and impulse ultrasounds.Because it has special natures such as small-size effect, quantum effect, skin effect, therefore, with after the chitosan nanoization because of the characteristic that nano particle had, can make it more effective by plant absorbing, improved availability.
Mentality of designing of the present invention is: chitin is the big molecule natural polysaccharide of a kind of straight chain, after being dissolved in acid, amino protonated back on the shitosan chain is positively charged, become cationic polymer, therefore, adopt ionic cross-linking, can with sodium phosphate trimer anion crosslinking agent, become Nano microsphere by electrostatic interaction with chitosan crosslinked.
The preparation method of the micro-nano biologic product of shitosan provided by the present invention is specially:
Chitin is dissolved in the acetum of 1vt%, the concentration that makes chitin is 2mg/ml, with 10mol/L sodium hydroxide adjust pH 4.6～4.7, degraded 4～8h gets the chitin acetum, in above-mentioned chitin acetum, dropwise add the 1mg/mL sodium tripolyphosphate solution, chitin acetum and sodium tripolyphosphate solution mass ratio are 5: 1, magnetic agitation 30min, impulse ultrasound 15min under condition of ice bath, the impulse ultrasound condition is a power 85%, opens 3 seconds every 2 seconds, obtains through crosslinked and finely dispersed chitosan nano biologic product.
Shitosan is regulated material as new plant growth, can secrete the resistance enzyme by inducing plant, excites the expression of resistant gene, effectively improves the resistance of plant to environment stress.The present invention is on the basis that has prepared chitin nano biological preparation, Primary Study the preparation product be used to alleviate the influence of Cd to wheat seeds sprouting and growth of seedling, through clear water, former dose and nanometer formulation comparative test, alleviate Cd to a certain extent and coerced toxic action to wheat seeds sprouting and growth of seedling, and the index of correlation (germination rate, germination vigor, root growth, seedling growth etc.) of wheat seeds sprouting and seedling growth had obvious facilitation, the result shows evident difference.
Description of drawings
Fig. 1 is the TEM photo of the former medicine of shitosan.
Fig. 2 is degraded 4h shitosan and the crosslinked back of STPP TEM photo.
Fig. 3 is shitosan and the crosslinked back of the STPP TEM photo of degraded 6h.
Fig. 4 the degrade shitosan of 8h and the crosslinked back of STPP TEM photo.
Fig. 5 is the infrared spectrum of crosslinked front and rear casing glycan, a be crosslinked before, b be crosslinked after.
Fig. 6 Nano chitosan is handled the influence of Cd being coerced down the wheat seedlings blade chlorophyll content.
By Fig. 1～4 as can be seen, the TEM photo of chitin after STPP is crosslinked of former medicine of shitosan and degraded different time.As seen with the reducing of the lengthening of degradation time, chitin molecule amount, the shitosan balling-up of crosslinked back is obvious more, disperses evenly more, and particle diameter is also more little.
As can be seen from Figure 4, the chitin nanoparticle is spherical, and is better dispersed, no reunion situation, and particle diameter is evenly distributed, about 200nm.
As can be seen from Figure 5, crosslinked front and back, the not too big variation of the absworption peak of chitin, this shows chitin behind ionomer, its structure does not change substantially.
Embodiment
Chitin is dissolved in the acetum of 1vt%, the concentration that makes chitin is 2mg/ml, with 10mol/L sodium hydroxide adjust pH 4.6～4.7, degraded 4～8h gets the chitin acetum, in above-mentioned chitin acetum, dropwise add the 1mg/mL sodium tripolyphosphate solution, chitin acetum and sodium tripolyphosphate solution mass ratio are 5: 1, magnetic agitation 30min, impulse ultrasound 15min under condition of ice bath, the impulse ultrasound condition is a power 85%,, opened 3 seconds every 2 seconds, obtain through crosslinked and finely dispersed chitosan nano biologic product.
Ultraviolet specrophotometer is surveyed the chitosan nano polymeric microspheres stabilize:
Adopt turbidimetry, measure the stability of Nano chitosan.The crosslinked suspension that obtains is left standstill room temperature 20 ℃ of condition 3d, 7d, 14d, 20d, 30d respectively, measure the variation of its absorbance with ultraviolet specrophotometer, the result shows: can deposit though chitosan nanoparticles was crossed about a week, but that still can disperse after the vibration is comparatively even, suspension dispersed fine, the change of size amplitude of chitosan nano is little.The chitosan nano system that the ionomer preparation is described is comparatively stable.
With the seed soaking of the Nano chitosan that utilizes method provided by the present invention to make to Cd coerce down wheat seeds sprouting influence and and utilize this Nano chitosan processing Cd to be coerced down the influence of wheat seedling growth, choosing, disinfecting and mensuration of the index of germinateing, the calculating of seed vitality etc. from seed, all according to national seed quality test stone, 4d surveys germination vigor, and 8d surveys germination rate.It is long that each handles root length and the bud of measuring 10 strain seedling simultaneously.
Germination index and vitality index are calculated as follows: germination index (GI)=∑ Gt/Dt; Vitality index (VI)=germination index (GI) * S; Gt is the germinative number in the t time, and Dt is corresponding germination fate; S is a bud length.
1, the influence of wheat seeds sprouting is coerced down in the Nano chitosan seed soaking to Cd
The influence of wheat seeds sprouting is coerced down in table 1 variable concentrations shitosan and Nano chitosan seed soaking to Cd
Former dose of shitosan and Nano chitosan all can be alleviated Cd and coerce the influence that wheat seed is sprouted as can be seen from Table 1.The Nano chitosan of 100mg/L is the most obvious to the effect of wheat germination gesture and germination rate, has improved 11.5% and 16.6% than CK respectively, significant difference.The control group optium concentration is 100mg/L, and its germination vigor and germination rate have improved 5.8% and 9.8% than CK respectively.The Nano chitosan of 100mg/L is also remarkable to the influence that Cd coerces down the wheat seed vigor, has improved 21.86% and 78.15% than CK respectively, and is significantly higher than the optimization process effect of former dose of shitosan.As seen Nano chitosan has tangible mitigation to the sprouting that Cd coerces down wheat seed, and action effect is better than former dose of shitosan.
2, Nano chitosan is handled the influence of Cd being coerced down wheat seedling growth
Table 2 Nano chitosan is handled the influence of Cd being coerced down wheat seedling growth
Table 2 shows, compares with the clear water contrast, and the wheat seedling plant height that 100mg/LCd coerces, root length all have obvious decline, have reduced by 28.5% and 50.6% respectively.The wheat seedling of handling through former dose of shitosan and Nano chitosan, when it was subjected to Cd and coerces, cadmium that its growth all obviously is better than not dealing with was coerced contrast, and alleviated and coerce effect and present to rise earlier with the increase of chitosan concentration and afterwards reduce.Influence to plant height is the most obvious with the treatment effect of former dose of the shitosan of 100mg/L and 200mg/L Nano chitosan, and coercing contrast than cadmium has respectively increased by 22.1% and 25.6%, and the treatment effect of Nano chitosan is better than former dose of shitosan.
The most obvious with former medicine of the shitosan of 150mg/L and Nano chitosan effect to the influence that root is long, coercing contrast than cadmium has increased by 16.3% and 23.7% respectively.From the depression effect of cadmium to wheat growth, under ground portion＞acrial part; And the relieving effect of Cd being coerced from shitosan, acrial part＞under ground portion, and the treatment effect of Nano chitosan is better than former dose of shitosan.
3, Nano chitosan is handled the influence of Cd being coerced down the wheat seedlings blade chlorophyll content
Green cellulose content is measured according to the method for Armon.Take by weighing the fresh blade of 0.5g, shred grinding, add 10mL 80% acetone soln and extract chlorophyll.The centrifugal 5min of 3000 * g gets supernatant, measures the OD value at 663nm, 645nm place respectively, and the following formula of gained OD value substitution calculates chlorophyllous content (mg/gFw)
C
a＝12.7×OD
663-2.69×OD
645；C
b＝22.9×OD
645-4.68×OD
663；
C
a+b＝C
a+C
b＝8.02×OD
663+20.2×OD
645
As seen from Figure 6, compare with the clear water contrast, chlorophyll a and chlorophyll a+b content that cadmium is coerced the contrast wheat seedlings blade significantly descend, and are respectively 61.3% and 67.4% of CK.Shitosan is handled, and can effectively alleviate the reduction that Cd coerces the leaf chlorophyll that causes.Through the increase of former dose of shitosan and Nano chitosan processing and concentration thereof, the content of chlorophyll a and chlorophyll a+b generally improves, and all coerces contrast greater than Cd.The Nano chitosan processed group, the treatment effect of 150mg/L and 200mg/L all is better than former dose of processed group of shitosan of 150mg/L, wherein the chlorophyll content of 200mg/L processing is the highest, chlorophyll a and a+b are respectively 90.4% and 92.4% of clear water contrasts, coerce contrast high 47.4% and 37.2% than Cd.
The chitin nano biological preparation that the present invention developed directly uses and gets final product.But act on foliage-spray on the plant, soil-perfusion; Administer water environment and agriculture soil heavy metal Cd etc., can directly put into the water Adsorption of Heavy Metal Ions.The utilization nanometer technology prepares the nano biological preparation, can improve the service efficiency of chitin, improves effect greatly.Its preparation method is simple, normal temperature and pressure, and behaviour is convenient with control, suitability for industrialized production.
Claims (3)
1. the preparation method of chitosan nano biologic product, chitin is dissolved in the acetum of 1vt%, the concentration that makes chitin is 2mg/ml, with 10mol/L sodium hydroxide adjust pH 4.6～4.7, degraded 4～8h gets the chitin acetum, in above-mentioned chitin acetum, dropwise add the 1mg/mL sodium tripolyphosphate solution, chitin acetum and sodium tripolyphosphate solution mass ratio are 5: 1, magnetic agitation 30min, impulse ultrasound 15min under condition of ice bath, the impulse ultrasound condition is a power 85%, opened 3 seconds every 2 seconds, obtain through crosslinked and finely dispersed chitosan nano biologic product.
2. the chitosan nano biologic product that makes of method according to claim 1.
3. the purposes of chitosan nano biologic product as claimed in claim 2 in wheat plant growth regulating process.
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Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103027035A (en) * | 2012-12-28 | 2013-04-10 | 辽宁师范大学 | Compound sodium nitrophenolate nano preparation and preparation method |
| CN103719150A (en) * | 2013-12-31 | 2014-04-16 | 辽宁师范大学 | Preparation method of selenium-rich biogenic nanometer antibacterial growth regulator |
| CN104745198A (en) * | 2015-02-12 | 2015-07-01 | 上海贯发海洋生物科技有限公司 | Preparation method of soil heavy metal curing detoxicant |
| CN105638663A (en) * | 2016-02-03 | 2016-06-08 | 河南农业大学 | Application of nanometer chitin to prevention and control of wheat aphids |
| CN105994269A (en) * | 2016-04-25 | 2016-10-12 | 王合中 | Application of chitin to promoting production of wheat |
| CN108371281A (en) * | 2018-02-01 | 2018-08-07 | 江南大学 | A method of cadmium in small Dried fish is removed using chitosan nano microballoon |
| CN109769823A (en) * | 2019-02-13 | 2019-05-21 | 河海大学 | A kind of load medicine chitosan sustained-release particle algae-inhibiting agent and preparation method |
| CN110419543A (en) * | 2019-06-05 | 2019-11-08 | 广西天浩农业发展有限公司 | A kind of slow-release insecticide and preparation method thereof |
| CN114304171A (en) * | 2022-01-11 | 2022-04-12 | 甘肃省科学院生物研究所 | Seed coating agent of ligularia virgaurea extract and preparation method and application thereof |
| CN114793528A (en) * | 2022-03-25 | 2022-07-29 | 浙江大学 | Application of nano enzyme in promoting corn growth under heavy metal stress |
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2010
- 2010-11-29 CN CN2010105636049A patent/CN102037962A/en active Pending
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| 孙庆申等: "壳聚糖微纳米粒制备及其特性的研究", 《黑龙江大学自然科学学报》 * |
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Cited By (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103027035A (en) * | 2012-12-28 | 2013-04-10 | 辽宁师范大学 | Compound sodium nitrophenolate nano preparation and preparation method |
| CN103719150A (en) * | 2013-12-31 | 2014-04-16 | 辽宁师范大学 | Preparation method of selenium-rich biogenic nanometer antibacterial growth regulator |
| CN103719150B (en) * | 2013-12-31 | 2015-08-12 | 辽宁师范大学 | A kind of preparation method of selenium-rich biogenic nanometer antibacterial growth regulator |
| CN104745198A (en) * | 2015-02-12 | 2015-07-01 | 上海贯发海洋生物科技有限公司 | Preparation method of soil heavy metal curing detoxicant |
| CN104745198B (en) * | 2015-02-12 | 2018-03-30 | 上海贯发海洋生物科技有限公司 | Heavy metal-polluted soil solidifies the preparation method of antidote |
| CN105638663B (en) * | 2016-02-03 | 2017-10-31 | 河南农业大学 | Application of the nanometer chitin in terms of wheat aphid is prevented and treated |
| CN105638663A (en) * | 2016-02-03 | 2016-06-08 | 河南农业大学 | Application of nanometer chitin to prevention and control of wheat aphids |
| CN105994269A (en) * | 2016-04-25 | 2016-10-12 | 王合中 | Application of chitin to promoting production of wheat |
| CN108371281A (en) * | 2018-02-01 | 2018-08-07 | 江南大学 | A method of cadmium in small Dried fish is removed using chitosan nano microballoon |
| CN109769823A (en) * | 2019-02-13 | 2019-05-21 | 河海大学 | A kind of load medicine chitosan sustained-release particle algae-inhibiting agent and preparation method |
| CN110419543A (en) * | 2019-06-05 | 2019-11-08 | 广西天浩农业发展有限公司 | A kind of slow-release insecticide and preparation method thereof |
| CN114304171A (en) * | 2022-01-11 | 2022-04-12 | 甘肃省科学院生物研究所 | Seed coating agent of ligularia virgaurea extract and preparation method and application thereof |
| CN114304171B (en) * | 2022-01-11 | 2023-11-28 | 甘肃省科学院生物研究所 | Ligularia virgaurea extract seed coating agent and preparation method and application thereof |
| CN114793528A (en) * | 2022-03-25 | 2022-07-29 | 浙江大学 | Application of nano enzyme in promoting corn growth under heavy metal stress |
| CN114793528B (en) * | 2022-03-25 | 2023-02-14 | 浙江大学 | Application of nano enzyme in promoting corn growth under heavy metal stress |
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Application publication date: 20110504 |