CN101215338B - Iron isomaltum oligosaccharide and preparing method thereof - Google Patents
Iron isomaltum oligosaccharide and preparing method thereof Download PDFInfo
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- CN101215338B CN101215338B CN2007100001470A CN200710000147A CN101215338B CN 101215338 B CN101215338 B CN 101215338B CN 2007100001470 A CN2007100001470 A CN 2007100001470A CN 200710000147 A CN200710000147 A CN 200710000147A CN 101215338 B CN101215338 B CN 101215338B
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- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 title claims abstract description 111
- 229910052742 iron Inorganic materials 0.000 title claims abstract description 56
- 238000000034 method Methods 0.000 title claims abstract description 37
- 229920001542 oligosaccharide Polymers 0.000 title claims abstract description 30
- 150000002482 oligosaccharides Chemical class 0.000 title claims description 28
- 238000002360 preparation method Methods 0.000 claims abstract description 32
- 238000006243 chemical reaction Methods 0.000 claims abstract description 19
- 150000003839 salts Chemical class 0.000 claims abstract description 13
- 238000000502 dialysis Methods 0.000 claims abstract description 12
- NCNCGGDMXMBVIA-UHFFFAOYSA-L iron(ii) hydroxide Chemical compound [OH-].[OH-].[Fe+2] NCNCGGDMXMBVIA-UHFFFAOYSA-L 0.000 claims abstract description 10
- 239000003960 organic solvent Substances 0.000 claims abstract description 10
- 235000014413 iron hydroxide Nutrition 0.000 claims abstract description 9
- 238000005342 ion exchange Methods 0.000 claims abstract description 6
- 150000001875 compounds Chemical class 0.000 claims abstract description 5
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 59
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 24
- 238000001556 precipitation Methods 0.000 claims description 23
- 238000000108 ultra-filtration Methods 0.000 claims description 21
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 19
- 238000003756 stirring Methods 0.000 claims description 16
- 239000000047 product Substances 0.000 claims description 15
- 238000005406 washing Methods 0.000 claims description 15
- CDBYLPFSWZWCQE-UHFFFAOYSA-L Sodium Carbonate Chemical compound [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 claims description 14
- 230000004913 activation Effects 0.000 claims description 13
- 230000000536 complexating effect Effects 0.000 claims description 12
- 239000007791 liquid phase Substances 0.000 claims description 12
- 239000006228 supernatant Substances 0.000 claims description 12
- 239000012528 membrane Substances 0.000 claims description 10
- 238000000926 separation method Methods 0.000 claims description 8
- 102000011759 adducin Human genes 0.000 claims description 7
- 108010076723 adducin Proteins 0.000 claims description 7
- 239000012535 impurity Substances 0.000 claims description 7
- RBTARNINKXHZNM-UHFFFAOYSA-K iron trichloride Chemical compound Cl[Fe](Cl)Cl RBTARNINKXHZNM-UHFFFAOYSA-K 0.000 claims description 7
- 235000017550 sodium carbonate Nutrition 0.000 claims description 7
- 229910000029 sodium carbonate Inorganic materials 0.000 claims description 7
- 229920005654 Sephadex Polymers 0.000 claims description 6
- 238000005227 gel permeation chromatography Methods 0.000 claims description 6
- 238000006386 neutralization reaction Methods 0.000 claims description 6
- 239000002244 precipitate Substances 0.000 claims description 5
- 239000000126 substance Substances 0.000 claims description 5
- 238000010792 warming Methods 0.000 claims description 5
- 238000007796 conventional method Methods 0.000 claims description 4
- 150000002500 ions Chemical class 0.000 claims description 4
- 238000000605 extraction Methods 0.000 claims description 3
- 238000005374 membrane filtration Methods 0.000 claims description 3
- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 claims description 2
- 239000007795 chemical reaction product Substances 0.000 claims description 2
- 238000010612 desalination reaction Methods 0.000 claims description 2
- 238000001514 detection method Methods 0.000 claims description 2
- 239000000706 filtrate Substances 0.000 claims description 2
- 229910052500 inorganic mineral Inorganic materials 0.000 claims description 2
- 239000011707 mineral Substances 0.000 claims description 2
- 235000010755 mineral Nutrition 0.000 claims description 2
- 238000001728 nano-filtration Methods 0.000 claims description 2
- 238000000746 purification Methods 0.000 claims description 2
- 238000005516 engineering process Methods 0.000 abstract description 6
- MVZXTUSAYBWAAM-UHFFFAOYSA-N iron;sulfuric acid Chemical compound [Fe].OS(O)(=O)=O MVZXTUSAYBWAAM-UHFFFAOYSA-N 0.000 abstract description 5
- 238000010521 absorption reaction Methods 0.000 abstract description 3
- 230000000694 effects Effects 0.000 abstract description 3
- 239000003513 alkali Substances 0.000 abstract description 2
- 150000001719 carbohydrate derivatives Chemical class 0.000 abstract description 2
- 238000001914 filtration Methods 0.000 abstract description 2
- AYRXSINWFIIFAE-SCLMCMATSA-N Isomaltose Natural products OC[C@H]1O[C@H](OC[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C=O)[C@@H](O)[C@@H](O)[C@@H]1O AYRXSINWFIIFAE-SCLMCMATSA-N 0.000 abstract 4
- -1 isomaltose oligosaccharide Chemical class 0.000 abstract 4
- 230000003213 activating effect Effects 0.000 abstract 2
- 208000015710 Iron-Deficiency Anemia Diseases 0.000 abstract 1
- 230000008021 deposition Effects 0.000 abstract 1
- 238000000151 deposition Methods 0.000 abstract 1
- 244000144972 livestock Species 0.000 abstract 1
- 239000000243 solution Substances 0.000 description 38
- 229920002307 Dextran Polymers 0.000 description 8
- 241001465754 Metazoa Species 0.000 description 8
- 208000007502 anemia Diseases 0.000 description 4
- 238000010926 purge Methods 0.000 description 4
- 238000004062 sedimentation Methods 0.000 description 4
- 241001071864 Lethrinus laticaudis Species 0.000 description 3
- 239000003814 drug Substances 0.000 description 3
- 230000007062 hydrolysis Effects 0.000 description 3
- 238000006460 hydrolysis reaction Methods 0.000 description 3
- 238000002347 injection Methods 0.000 description 3
- 239000007924 injection Substances 0.000 description 3
- 210000003205 muscle Anatomy 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 231100000215 acute (single dose) toxicity testing Toxicity 0.000 description 2
- 238000011047 acute toxicity test Methods 0.000 description 2
- 230000037396 body weight Effects 0.000 description 2
- 238000001816 cooling Methods 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 238000009826 distribution Methods 0.000 description 2
- 239000013049 sediment Substances 0.000 description 2
- 238000013112 stability test Methods 0.000 description 2
- 239000005715 Fructose Substances 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 206010053759 Growth retardation Diseases 0.000 description 1
- 101000993059 Homo sapiens Hereditary hemochromatosis protein Proteins 0.000 description 1
- 241000581650 Ivesia Species 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 238000013019 agitation Methods 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 150000008064 anhydrides Chemical class 0.000 description 1
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 229940041984 dextran 1 Drugs 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 230000003203 everyday effect Effects 0.000 description 1
- 230000002349 favourable effect Effects 0.000 description 1
- 238000000855 fermentation Methods 0.000 description 1
- 230000004151 fermentation Effects 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 150000004676 glycans Chemical class 0.000 description 1
- 231100000001 growth retardation Toxicity 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 235000020256 human milk Nutrition 0.000 description 1
- 210000004251 human milk Anatomy 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-M hydroxide Chemical compound [OH-] XLYOFNOQVPJJNP-UHFFFAOYSA-M 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 238000010253 intravenous injection Methods 0.000 description 1
- 150000002505 iron Chemical class 0.000 description 1
- 238000005304 joining Methods 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 235000013372 meat Nutrition 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 229920001282 polysaccharide Polymers 0.000 description 1
- 239000005017 polysaccharide Substances 0.000 description 1
- 235000015277 pork Nutrition 0.000 description 1
- 239000003755 preservative agent Substances 0.000 description 1
- 230000008929 regeneration Effects 0.000 description 1
- 238000011069 regeneration method Methods 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 231100000161 signs of toxicity Toxicity 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 239000006188 syrup Substances 0.000 description 1
- 235000020357 syrup Nutrition 0.000 description 1
- 230000001131 transforming effect Effects 0.000 description 1
- 210000003462 vein Anatomy 0.000 description 1
Landscapes
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Polysaccharides And Polysaccharide Derivatives (AREA)
Abstract
The invention belongs to the saccharide derivative preparation technology field, in particular to an isomaltose oligosaccharide and preparation process. Isomaltose oligosaccharide is a novel stable chelating biological iron, and has the excellent effect of preventing iron-deficiency anemia of a human or livestock. The productive process comprises firstly preparing or choosing activating isomaltose oligosaccharide whose average molecular weight is 1500-2000 to be purified, conducting complex reaction of activating isomaltose oligosaccharide and iron hydroxide under a certain condition, wherein iron hydroxide is formed by the reaction of inorganic ferric salt and alkali before or in the process of the complex reaction, thirdly, purifying complex compound through adopting the methods such as organic solvent deposition, ion exchange, dialysis or hyper-filtration and the like. The product which is prepared by the invention has wider iron content range from 5% to 30% compared with current iron dextran products at home and abroad, and is superior to other product in other aspects such as flowability, viscosity, stability and absorption effect.
Description
Technical field
The invention belongs to the preparing technical field of carbohydrate derivative, be specially a kind of iron isomaltum oligosaccharide and preparation method thereof.
Background technology
Hypoferric anemia is not only human a kind of common disease, and is in animal kingdom, especially also very general in the domestic animal.For example piggy almost all has anaemia in various degree more than 95%.This be since piggy from breast milk and feed the institute getable iron amount also the deficiency its physiological requirements amount 10%.Cause growth retardation therefrom, lose weight, disease incidence rate and mortality ratio increase.Science proves, injecting disposable high-quality Iron Dextran for nascent piggy can make above-mentioned situation improve fully, therefore most developed countries are in the forbidding hormone medicine, supporting and encourageing people adopts this scientific methods to promote the health of pig, improve the quality and the output of pork, a large amount of facts have proved adopts this method can make the meat productivity of pig increase more than 20%.
Regrettably, present dextran iron product, no matter in quality with quantitatively also can not satisfy the needs of world market far away, major cause is owing to produce that required raw material--the quality and the synthetic technology of dextran remain in some problems.Traditional fermentation hydrolysis method is generally all adopted in the preparation of existing dextran.The shortcoming of this method is: the first, at the industrial ultra-low molecular volume production product that can't produce below 3000; The second, the product molecular weight distribution a wider range after the hydrolysis, and small molecular weight impurity is difficult to eliminate; The 3rd, this method yield is very low, causes cost up.This has just caused the quality of the Iron Dextran produced superior inadequately, and price is also higher relatively, and this is constituting some disadvantageous effects aspect its large-scale promotion, make further developing of this field be very restricted.
Summary of the invention
The objective of the invention is to requirement according to the world market, research and develop a kind of more more superior than existing dextran weight of iron, and medicine---the iron isomaltum oligosaccharide of the control people, animal hypoferric anemia that price is more cheap is in the hope of becoming the regeneration product of conventional medicament; Simultaneously, the present invention also will provide the preparation method of this iron isomaltum oligosaccharide.
Technical scheme of the present invention is as follows: iron isomaltum oligosaccharide forms the biological iron of chelating of nonionic key by dextrinosan and inorganic iron complexing, and its chemical general formula is (C
6H
10O
5)
nHOOFe, wherein, n=6～18.
The preparation method of iron isomaltum oligosaccharide, step specific as follows:
(1) dextrinosan is dissolved in water, be mixed with concentration and be 15～20% solution, carry out fractional precipitation with ethanolic soln then, or carry out ultrafiltration with ultra-filtration membrane, or employing dextran gel chromatography separation method, be prepared into molecular-weight average and be 1500～2000 dextrinosan, high-pressure liquid phase is unimodal, area 99～100%, adopt conventional method to activate then, obtain reducing sugar and be lower than 0.1% activation dextrinosan, gained is activated dextrinosan carry out purifying, remove salt and other low molecular impurities;
(2) get an amount of purified iron hydroxide solution, measure wherein ferro element weight, the concentration that adds equivalent weight then is 20～25% above-mentioned activation dextrinosan solution, temperature is at 25～30 ℃, liquid phase mixes, and dripping 25% yellow soda ash or 8N sodium hydroxide solution is 11 to pH, heat temperature raising to 90～95 ℃ then, the continuously stirring reaction is finished until complex reaction;
(3) product with above-mentioned complex reaction carries out washing of precipitate, dialysis or ultra-filtration by organic solvent, removes salt and low molecular impurity.
The preparation method of iron isomaltum oligosaccharide as mentioned above, wherein, the process of ethanol fractional precipitation is as follows in the step (1): in 15～20% dextrinosan solution, add 95% ethanol, and slowly stir, reach at 38% o'clock to the concentration of ethanol in solution and stop, treating that precipitation fully after, change supernatant liquor over to another container, in supernatant liquor, add 95% ethanol again, and stir, reach at 55% o'clock to alcohol concn and stop, treating that precipitation fully after, supernatant liquor is removed in suction, the collecting precipitation thing, and with a small amount of 95% washing with alcohol twice, then throw out is dissolved in the pure water.
The preparation method of iron isomaltum oligosaccharide as mentioned above, wherein, the purification process of activation dextrinosan comprises in the step (1): organic solvent deposit washing method, dialysis method, ultra-filtration membrane and nanofiltration membrane separation method, and ion exchange method.
The preparation method of iron isomaltum oligosaccharide as mentioned above, wherein, the preparation method of iron hydroxide solution is in the step (2): the preparation iron level is 5～7% inorganic molysite solution, 25～30 ℃ of temperature, drip 25% yellow soda ash or 8N sodium hydroxide solution, fully stir, make pH reach 1.5～1.8, filter disgorging then; Further with filtrate by dialysis or ion-exchange or ultrafiltration, remove the salt that produces in the neutralization reaction and ion to obtain the iron hydroxide solution of purifying.
The preparation method of iron isomaltum oligosaccharide as mentioned above, wherein, the organic solvent in the step (3) is ethanol or Virahol.
The another kind of preparation method of iron isomaltum oligosaccharide, step specific as follows:
(1) dextrinosan is dissolved in water, be mixed with concentration and be 15～20% solution, carry out branch's precipitation with ethanolic soln then, or carry out ultrafiltration with ultra-filtration membrane, or employing dextran gel chromatography separation method, be prepared into molecular-weight average and be 1500～2000 dextrinosan, high-pressure liquid phase is unimodal, area 99～100%, adopt conventional method to activate then, obtain reducing sugar and be lower than 0.1% activation dextrinosan, gained is activated dextrinosan carry out purifying, remove salt and other low molecular impurities;
(2) an amount of inorganic molysite being mixed with concentration is 5% solution, measure wherein ferro element weight, be that 20～25% above-mentioned activation dextrinosan solution directly mixes then with itself and the concentration of equivalent weight, temperature is at 25～30 ℃, slowly drip 25% yellow soda ash or 8N sodium hydroxide solution, to pH be 6.0～6.5, after dropwising, be warming up to 45～50 ℃, continue reaction 4 hours;
(3) carry out washing of precipitate, dialysis or ultra-filtration by organic solvent, remove a large amount of salts and the mineral ion that produce in the above-mentioned reaction product;
(4) complex compound behind the above-mentioned desalination is dissolved in water, regulate pH to 10, be warming up to 80～90 ℃ of reactions 2 hours then, after high-pressure liquid phase detection complexing is finished, can cool the temperature to room temperature and pH is transferred to 6.0, and then add isopyknic 95% ethanolic soln precipitation, washing once, and throw out is dissolved in the pure water again, add heat extraction ethanol.
Raw material--the dextrinosan that the present invention adopts has fundamentally overcome dextran existing shortcoming on preparation technology.Be in particular in: the first, identical with dextran on its structure, but molecular weight significantly reduces, and is 1500～2000 only, points out it to have how active terminal group, this will improve itself and the complex ability of iron and the stability and the result of treatment of product greatly; In addition, by the product of low molecular weight feedstocks preparation, viscosity reduces, and this not only makes injection more easy, and provides possibility for the product of preparation high Fe content (for example 20～30%).The second, the dextrinosan molecular weight distribution that the present invention adopts is extremely narrow, and high-pressure liquid phase Zhong ， Mine Kuan is Yu the high ratio of Mine only is half of traditional zymotic hydrolysis method, and purity reaches 99～100%, and this more guarantees the high purity and the hypotoxicity of product after the complexing.The 3rd, the productive rate that this method obtains is about three times of traditional method.Use technology of the present invention institute synthetic quality product and be far superior to current international other dextran iron products, price then significantly reduces.
Embodiment
Dextrinosan system by glucose with α 1-6 glycosidic link mode be combined into, its structure and dextran are basic identical, just carbochain is significantly on the low side, thereby have more for the terminal group of transforming, these terminal group (hydroxyl) are after oxidation, become active carboxyl, can with the ferro element complexing, form biological iron---the iron isomaltum oligosaccharide of chelating of nonionic key.
Iron isomaltum oligosaccharide forms the biological iron of chelating of nonionic key by dextrinosan and inorganic iron complexing, and its chemical general formula is (C
6H
10O
5)
nHOOFe, wherein, n=6～18.
Iron isomaltum oligosaccharide is the ideal medicament of control people, animal hypoferric anemia, it is characterized in that:
(1) the iron level scope is wider, can be from 5%, 10% to 20%, even 30%, be suitable for preparing the product of all size.
(2) with existing Iron Dextran relatively, iron isomaltum oligosaccharide solution has good flowability and very low viscosity (only be general Iron Dextran 1/4th), thereby is more suitable in muscle or intravenous injection.
(3) stability of iron isomaltum oligosaccharide and absorptivity are extremely good, and acute toxicity test is also qualified fully.
The introduction of relevant dextrinosan can be with reference to Chinese patent application " a kind of dextrinosan-fructose syrup and preparation method thereof " (03100280.3) and " preparation method of IMOS (IMOS) " (200510002141.8).
Embodiment 1
Neutralization reaction was carried out before the complexing process, i.e. hydroxide iron processes, and concrete operations are as follows:
(1) preparation of ironic hydroxide
Weigh analytical pure iron trichloride (FeCl
36H
2O) 242 grams are put in the 2000ml there-necked flask, add water and stir and to make moltenly, and volume are transferred to 1000ml.Attemperation is at 25～30 ℃, and stirring velocity is 100 rev/mins, begins to drip 25%Na
2CO
3Solution, rate of addition are the 3ml/ branch, are 1.5～1.8 can stop to pH.
Reaction solution is packed in the dialysis tubing, pure water is dialysed, rise to more than 3.0 as pH after 48 hours, and cl content is lower than at 0.1% o'clock, can stop dialysis, solution is filtered standby, should measure iron level before using.This routine iron level is 6.8%.
(2) activation dextrinosan solution
The activation dextrinosan is provided by Shenzhen Herbon Polysaccharide Bio-technology Co., Ltd., its process for preparation is that dextrinosan is dissolved in water, be mixed with concentration and be 15～20% solution, carry out fractional precipitation with ethanol then, or carry out ultrafiltration with ultra-filtration membrane, and or adopt dextran gel chromatography separation method, it is unimodal to be prepared into molecular-weight average and to be 1500～2000 dextrinosan (25 ℃ of intrinsic viscosities 0.035～0.040) high-pressure liquid phase, purity is more than 99%, and reducing sugar is lower than 0.01%.Get this product 40 gram, add water and make moltenly, and volume transferred to 200ml, filter standby (fresh preparation) then.Ultra-filtration membrane ultrafiltration process and dextran gel chromatography separation method are known technology.The process of ethanol fractional precipitation is as follows: in 15～20% dextrinosan solution, add 95% ethanol, and slowly stirring (30～50 rev/mins), reach at 38% o'clock to the concentration of ethanol in solution and stop, leaving standstill about 2 hours, treat that precipitation fully after, change supernatant liquor over to another container, in supernatant liquor, add 95% ethanol again, and stir, reach at 55% o'clock to alcohol concn and stop, left standstill about 4 hours, after treating that precipitation fully, inhale and remove supernatant liquor, collecting precipitation thing, and with twice of a small amount of 95% washing with alcohol, then throw out is dissolved in the pure water, detects through high-pressure liquid phase, this part dextrinosan molecular weight is in 1500～2000 scopes.
(3) complex reaction
Above-mentioned activatory dextrinosan liquid 200ml is mixed with 6.8% iron hydroxide solution 588ml, put in the 1000ml there-necked flask, attemperation is 25～30 ℃ of scopes, 50～100 rev/mins of stirring velocitys, begin to drip 8N sodium hydroxide to pH11, be heated to 90～95 ℃ of reactions 2 hours then, high-pressure liquid phase is measured and is shown that complex reaction is finished.
(4) purge process
Product after the complexing contains more salt and other lower-molecular substances, must be further purified.This example adopts the method for ethanol sedimentation washing.Process is as follows: the iron level of above-mentioned complex compound is transferred to 14～15%, under the agitation condition that (is lower than 50 rev/mins) at a slow speed, add 95% ethanol sedimentation of 1.2 times of complex compound volumes.After leaving standstill 30 minutes, inhale and abandon supernatant liquor, use small amount of ethanol washing precipitation secondary then.Again throw out is dissolved in the pure water, further adds heat extraction ethanol, sampling the carrying out mass analysis of cooling back, if free iron content is lower than 0.2%, ethanol eliminates, and cl content is lower than 0.8%, then solution can be filtered,, use 0.22 μ millipore filtration more earlier with 0.45 μ cardboard filter.
(5) adjusting of iron isomaltum oligosaccharide solution component
At first be the adjusting of concentration, can be by to the concentrating or dilution of solution behind the purifying, so that the solution iron level reaches predetermined level, for example 5%, 10%, 15%, 20% or 30%.
Next is pH, the cl content of regulator solution, makes it to reach the officinal requirement, and the adjusting of pH value is to adopt the conventional mode that adds acid or add alkali to realize.
The 3rd is to add suitable preservatives, and makes its content be controlled at the pharmacopeia specialized range.
Embodiment 2
Neutralization reaction is carried out in the complexing process, i.e. inorganic molysite method, and concrete operations are as follows:
(1) neutralization and complex reaction
Weigh analytical pure iron trichloride (FeCl
36H
2O) 193.6 grams add water and make moltenly, and volume are transferred to 800ml, pour into then in the there-necked flask of 2000ml, and the activation dextrinosan solution (joining method with embodiment 1) with 200ml 20% is added in the above-mentioned liquor ferri trichloridi again.Then there-necked flask is placed 25～30 ℃ water-soluble, turn on agitator makes rotating speed reach 50～100 rev/mins, after treating that solution temperature is constant, begin to drip 30% sodium carbonate solution till the pH 6.0～6.5, with temperature being increased to 45～50 ℃, and continue to stir 4 hours.
(2) purge process
Since simultaneous neutralization reaction in the above-mentioned complexing process, thereby have a large amount of salts to produce, so require to carry out effective purge process.Comprise organic solvent deposit, dialysis, ion-exchange or ultrafiltration membrance filter etc.This example adopts the method for ethanol sedimentation.Operate as follows: earlier the solution after the above-mentioned preliminary complexing is passed through 0.45 μ membrane filtration, then under (50 rev/mins) stir at a slow speed, careful 95% ethanol sedimentation that adds 1.5 times of volumes, left standstill 30～60 minutes, treat that layering is clear after, inhale and to abandon supernatant liquor, the lower sediment thing is used 60% washing with alcohol two to three times again, measure residual free iron and cl content in the throw out, finish, otherwise continue washing as the qualified purge process of then pointing out.
(3) strengthen complex reaction
In pure water, the regulator solution volume makes iron content in 5～8% scopes, uses 5N NaOH solution simultaneously with the resolution of precipitate behind the above-mentioned purifying, regulates pH to 8.0, is warming up to 85～90 ℃ then and also continues to stir (50 rev/mins) 2 hours.Cool the temperature to then below 30 ℃, and add 1.5 times of volumes, 95% ethanol redeposition once, after leaving standstill 30 minutes, after supernatant is abandoned in suction, precipitation is dissolved with pure water again, volume is controlled at iron content 5%～7%, and reheat to 90 ℃ is removed ethanol and is concentrated into desired concn (for example 10% or 20%).
(4) adjusting of dextrinosan anhydride solution component
With embodiment 1
Repeatability and stability test
1, replica test
10 batches 20% iron isomaltum oligosaccharide solution of method preparation of Application Example 1, and the main chemical compositions in the mensuration all samples, result's (mean+SD) is followed successively by: iron level 20.15 ± 0.81 (%), dextrinosan content 16.15 ± 1.45 (%), cl content 0.85 ± 0.13 (%), 25 ℃ relative viscosity are 5.36 ± 0.81, free iron content is 0.15 ± 0.05 (%).
The result of Application Example 2 gained and above-mentioned approaching points out its repeatability good.These samples have also been sent international how tame specialized company and research institution test acquisition their approval and favorable comment.
2, stability test
The method of Application Example 1 and embodiment 2 prepares 80 batch samples altogether, be sub-packed in the 100ml vial, be put in respectively after the sterilization cooling under 4 ℃, 25 ℃ and 50 ℃ of three temperature, observe the color and the precipitation situation of a sample every day, followed up a case by regular visits to 1 annual bearing and shown, color sample does not have any change when 4 ℃ and 25 ℃, does not have the obvious sediment generation yet, about 5% sample has the prompting of minute quantity precipitation to have good stability in the time of 50 ℃, meets the world market requirement fully.
Biology is measured
1, acute toxicity test
According to American Pharmacopeia, get 20 of male white mouses (about mean body weight 20 grams), be divided into two groups: A group and B group, 10 every group.By the dosage of per kilogram of body weight 200mg, from 20% prepared iron isomaltum oligosaccharide solution of A group and B group small white mouse tail vein injection embodiment 1 and embodiment 2, observed for 1 week respectively, all animals all do not have any signs of toxicity, and all survive.
2, muscle absorption test
Get 6 male rabbits, be divided into two groups of A, B at random, 3 every group, outside deep part muscle is injected 20% the iron isomaltum oligosaccharide solution of 1ml by embodiment 1 and embodiment 2 preparations respectively on two treated animal back legs.With sacrifice of animal, the absorbing state of anatomic observation injection portion iron, result show that the iron absorption of 6 animals is 100% after one week.
Claims (7)
1. an iron isomaltum oligosaccharide by dextrinosan and inorganic iron complexing, forms the biological iron of chelating of nonionic key, and its chemical general formula is (C
6H
10O
5)
nHOOFe, n=6～18.
2. the preparation method of an iron isomaltum oligosaccharide, step specific as follows:
(1) dextrinosan is dissolved in water, be mixed with concentration and be 15～20% solution, carry out fractional precipitation with ethanolic soln then, or carry out ultrafiltration with ultra-filtration membrane, or employing dextran gel chromatography separation method, be prepared into molecular-weight average and be 1500～2000 dextrinosan, high-pressure liquid phase is unimodal, area 99～100%, adopt conventional method to activate then, obtain reducing sugar and be lower than 0.1% activation dextrinosan, gained is activated dextrinosan carry out purifying, remove salt and other low molecular impurities;
(2) get an amount of purified iron hydroxide solution, measure wherein ferro element weight, the concentration that adds equivalent weight then is 20～25% above-mentioned activation dextrinosan solution, temperature is at 25～30 ℃, liquid phase mixes, and dripping 25% yellow soda ash or 8N sodium hydroxide solution is 11 to pH, heat temperature raising to 90～95 ℃ then, the continuously stirring reaction is finished until complex reaction;
(3) product with above-mentioned complex reaction carries out washing of precipitate, dialysis or ultra-filtration by organic solvent, removes salt and low molecular impurity.
3. the preparation method of iron isomaltum oligosaccharide as claimed in claim 2, it is characterized in that: the process of ethanol fractional precipitation is as follows in the step (1): in 15～20% dextrinosan solution, add 95% ethanol, and slowly stir, reaching at 38% o'clock to the concentration of ethanol in solution stops, after treating that precipitation fully, change supernatant liquor over to another container, in supernatant liquor, add 95% ethanol again, and stir, reaching at 55% o'clock to alcohol concn stops, after treating that precipitation fully, inhale and remove supernatant liquor, collecting precipitation thing, and, then throw out is dissolved in the pure water with a small amount of 95% washing with alcohol twice.
4. the preparation method of iron isomaltum oligosaccharide as claimed in claim 2, it is characterized in that: the purification process of activation dextrinosan comprises in the step (1): the organic solvent deposit washing method, dialysis method, ultra-filtration membrane and nanofiltration membrane separation method, and ion exchange method.
5. the preparation method of iron isomaltum oligosaccharide as claimed in claim 2, it is characterized in that: the preparation method of iron hydroxide solution is in the step (2): the preparation iron level is 5～7% inorganic molysite solution, 25～30 ℃ of temperature, drip 25% yellow soda ash or 8N sodium hydroxide solution, fully stir, make pH reach 1.5～1.8, filter disgorging then; Further with filtrate by dialysis or ion-exchange or ultrafiltration, remove the salt that produces in the neutralization reaction and ion to obtain the iron hydroxide solution of purifying.
6. the preparation method of iron isomaltum oligosaccharide as claimed in claim 2, it is characterized in that: the organic solvent in the step (3) is ethanol or Virahol.
7. the preparation method of an iron isomaltum oligosaccharide, step specific as follows:
(1) dextrinosan is dissolved in water, be mixed with concentration and be 15～20% solution, carry out fractional precipitation with ethanolic soln then, or carry out ultrafiltration with ultra-filtration membrane, or employing dextran gel chromatography separation method, be prepared into molecular-weight average and be 1500～2000 dextrinosan, high-pressure liquid phase is unimodal, area 99～100%, adopt conventional method to activate then, obtain reducing sugar and be lower than 0.1% activation dextrinosan, gained is activated dextrinosan carry out purifying, remove salt and other low molecular impurities;
(2) an amount of inorganic molysite being mixed with concentration is 5% solution, measure wherein ferro element weight, be that 20～25% above-mentioned activation dextrinosan solution directly mixes then with itself and the concentration of equivalent weight, temperature is at 25～30 ℃, slowly drip 25% yellow soda ash or 8N sodium hydroxide solution, to pH be 6.0～6.5, after dropwising, be warming up to 45～50 ℃, continue reaction 4 hours;
(3) carry out washing of precipitate, dialysis or ultra-filtration by organic solvent, remove a large amount of salts and the mineral ion that produce in the above-mentioned reaction product;
(4) complex compound behind the above-mentioned desalination is dissolved in water, regulate pH to 10, be warming up to 80～90 ℃ of reactions 2 hours then, after high-pressure liquid phase detection complexing is finished, can cool the temperature to room temperature and pH is transferred to 6.0, and then add isopyknic 95% ethanolic soln precipitation, washing once, and throw out is dissolved in the pure water again, add heat extraction ethanol.
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| EP1947120A1 (en) * | 2007-01-19 | 2008-07-23 | Vifor (International) Ag | Iron-carbohydrate complex compounds |
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| CA2891198A1 (en) * | 2012-11-30 | 2014-06-05 | Pharmacosmos A/S | Cryoprotecting agent, cryoprotecting and cryopreserved compositions, uses thereof, and methods of cryopreservation |
| CN106674367A (en) * | 2016-12-14 | 2017-05-17 | 烟台东诚药业集团股份有限公司 | Method for purifying and preparing isomalto-oligosaccharide and application |
| CN107712357A (en) * | 2017-11-21 | 2018-02-23 | 天峨县宏昌农机专业合作社 | A kind of complex compound of secondary Complexing Iron and preparation method thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1041762A (en) * | 1988-10-14 | 1990-05-02 | 广西化工研究所 | The preparation method of Iron-Dextrin Complex |
| CN1097989A (en) * | 1993-12-27 | 1995-02-01 | 广东太阳神集团有限公司 | Sucrose ferrum, its preparation method and preparation thereof |
| JP2006241067A (en) * | 2005-03-03 | 2006-09-14 | Showa Sangyo Co Ltd | Allergy inhibitor |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1041762A (en) * | 1988-10-14 | 1990-05-02 | 广西化工研究所 | The preparation method of Iron-Dextrin Complex |
| CN1097989A (en) * | 1993-12-27 | 1995-02-01 | 广东太阳神集团有限公司 | Sucrose ferrum, its preparation method and preparation thereof |
| JP2006241067A (en) * | 2005-03-03 | 2006-09-14 | Showa Sangyo Co Ltd | Allergy inhibitor |
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