CN102743742B - A kind of glycoprotein compositions of low solvent residue and its production and use - Google Patents
A kind of glycoprotein compositions of low solvent residue and its production and use Download PDFInfo
- Publication number
- CN102743742B CN102743742B CN201110102073.8A CN201110102073A CN102743742B CN 102743742 B CN102743742 B CN 102743742B CN 201110102073 A CN201110102073 A CN 201110102073A CN 102743742 B CN102743742 B CN 102743742B
- Authority
- CN
- China
- Prior art keywords
- preparation
- glycoprotein
- water
- organic solvent
- vacuum drying
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 102000003886 Glycoproteins Human genes 0.000 title claims abstract description 83
- 108090000288 Glycoproteins Proteins 0.000 title claims abstract description 83
- 239000000203 mixture Substances 0.000 title claims abstract description 72
- 239000002904 solvent Substances 0.000 title claims abstract description 50
- 238000004519 manufacturing process Methods 0.000 title description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 57
- 239000003960 organic solvent Substances 0.000 claims abstract description 44
- 238000002360 preparation method Methods 0.000 claims abstract description 40
- 239000002244 precipitate Substances 0.000 claims abstract description 33
- 238000001291 vacuum drying Methods 0.000 claims abstract description 25
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N ethanol Chemical group CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 58
- 239000002994 raw material Substances 0.000 claims description 21
- 239000000243 solution Substances 0.000 claims description 20
- 239000000463 material Substances 0.000 claims description 15
- 238000006297 dehydration reaction Methods 0.000 claims description 14
- 229940088597 Hormone Drugs 0.000 claims description 11
- 239000005556 hormone Substances 0.000 claims description 11
- 239000007788 liquid Substances 0.000 claims description 11
- 238000002156 mixing Methods 0.000 claims description 11
- CSCPPACGZOOCGX-UHFFFAOYSA-N acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims description 8
- UIIMBOGNXHQVGW-UHFFFAOYSA-M buffer Substances [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 claims description 8
- OKKJLVBELUTLKV-UHFFFAOYSA-N methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 8
- 229940015047 Chorionic Gonadotropin Drugs 0.000 claims description 6
- 108010062540 Chorionic Gonadotropin Proteins 0.000 claims description 6
- 102000011022 Chorionic Gonadotropin Human genes 0.000 claims description 6
- 108010073521 Luteinizing Hormone Proteins 0.000 claims description 5
- 102000009151 Luteinizing Hormone Human genes 0.000 claims description 5
- 238000001556 precipitation Methods 0.000 claims description 5
- 238000004090 dissolution Methods 0.000 claims description 4
- 239000007983 Tris buffer Substances 0.000 claims description 3
- 239000002131 composite material Substances 0.000 claims description 3
- 239000007864 aqueous solution Substances 0.000 claims description 2
- 230000003139 buffering Effects 0.000 claims description 2
- 238000001223 reverse osmosis Methods 0.000 claims description 2
- 239000007974 sodium acetate buffer Substances 0.000 claims description 2
- 239000008215 water for injection Substances 0.000 claims description 2
- 239000008367 deionised water Substances 0.000 claims 1
- 238000004817 gas chromatography Methods 0.000 claims 1
- 239000008363 phosphate buffer Substances 0.000 claims 1
- 238000003756 stirring Methods 0.000 description 14
- QTBSBXVTEAMEQO-UHFFFAOYSA-N acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 12
- 238000000034 method Methods 0.000 description 9
- 230000014759 maintenance of location Effects 0.000 description 8
- 238000004108 freeze drying Methods 0.000 description 7
- 239000000047 product Substances 0.000 description 7
- 235000001014 amino acid Nutrition 0.000 description 6
- 150000001413 amino acids Chemical class 0.000 description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- 239000008213 purified water Substances 0.000 description 5
- 208000000509 Infertility Diseases 0.000 description 4
- 229960004249 Sodium Acetate Drugs 0.000 description 4
- 238000001035 drying Methods 0.000 description 4
- 230000036512 infertility Effects 0.000 description 4
- 231100000535 infertility Toxicity 0.000 description 4
- IJGRMHOSHXDMSA-UHFFFAOYSA-N nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 4
- VMHLLURERBWHNL-UHFFFAOYSA-M sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 4
- 239000001632 sodium acetate Substances 0.000 description 4
- 235000017281 sodium acetate Nutrition 0.000 description 4
- 239000007787 solid Substances 0.000 description 4
- 241000894007 species Species 0.000 description 4
- 102000006771 Gonadotropins Human genes 0.000 description 3
- 108010086677 Gonadotropins Proteins 0.000 description 3
- DCXYFEDJOCDNAF-REOHCLBHSA-N L-asparagine Chemical compound OC(=O)[C@@H](N)CC(N)=O DCXYFEDJOCDNAF-REOHCLBHSA-N 0.000 description 3
- 230000000052 comparative effect Effects 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 239000002622 gonadotropin Substances 0.000 description 3
- 239000012535 impurity Substances 0.000 description 3
- 210000004246 Corpus Luteum Anatomy 0.000 description 2
- VEXZGXHMUGYJMC-UHFFFAOYSA-N HCl Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 229920003013 deoxyribonucleic acid Polymers 0.000 description 2
- 238000005265 energy consumption Methods 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 238000011067 equilibration Methods 0.000 description 2
- 239000007789 gas Substances 0.000 description 2
- 238000010438 heat treatment Methods 0.000 description 2
- 230000002779 inactivation Effects 0.000 description 2
- 230000009245 menopause Effects 0.000 description 2
- 229910000403 monosodium phosphate Inorganic materials 0.000 description 2
- 235000019799 monosodium phosphate Nutrition 0.000 description 2
- 229910052757 nitrogen Inorganic materials 0.000 description 2
- 235000018102 proteins Nutrition 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- AJPJDKMHJJGVTQ-UHFFFAOYSA-M sodium dihydrogen phosphate Chemical compound [Na+].OP(O)([O-])=O AJPJDKMHJJGVTQ-UHFFFAOYSA-M 0.000 description 2
- 239000008399 tap water Substances 0.000 description 2
- 235000020679 tap water Nutrition 0.000 description 2
- 206010002198 Anaphylactic reaction Diseases 0.000 description 1
- 208000003455 Anaphylaxis Diseases 0.000 description 1
- 229940057854 Gonal F Drugs 0.000 description 1
- 229940084986 Human Chorionic Gonadotropin Drugs 0.000 description 1
- GUBGYTABKSRVRQ-UUNJERMWSA-N Lactose Natural products O([C@@H]1[C@H](O)[C@H](O)[C@H](O)O[C@@H]1CO)[C@H]1[C@@H](O)[C@@H](O)[C@H](O)[C@H](CO)O1 GUBGYTABKSRVRQ-UUNJERMWSA-N 0.000 description 1
- 229940040129 Luteinizing Hormone Drugs 0.000 description 1
- NCYVXEGFNDZQCU-UHFFFAOYSA-N Nikethamide Chemical compound CCN(CC)C(=O)C1=CC=CN=C1 NCYVXEGFNDZQCU-UHFFFAOYSA-N 0.000 description 1
- 210000004681 Ovum Anatomy 0.000 description 1
- 108010020661 Profasi Proteins 0.000 description 1
- CXQXSVUQTKDNFP-UHFFFAOYSA-N Simethicone Chemical compound C[Si](C)(C)O[Si](C)(C)O[Si](C)(C)C CXQXSVUQTKDNFP-UHFFFAOYSA-N 0.000 description 1
- GSEJCLTVZPLZKY-UHFFFAOYSA-N Tris Chemical compound OCCN(CCO)CCO GSEJCLTVZPLZKY-UHFFFAOYSA-N 0.000 description 1
- 210000002700 Urine Anatomy 0.000 description 1
- 238000007792 addition Methods 0.000 description 1
- 230000036783 anaphylactic response Effects 0.000 description 1
- 108010066058 beta Subunit Luteinizing Hormone Proteins 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 230000005540 biological transmission Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 239000000969 carrier Substances 0.000 description 1
- 239000012159 carrier gas Substances 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 239000004205 dimethyl polysiloxane Substances 0.000 description 1
- 235000013870 dimethyl polysiloxane Nutrition 0.000 description 1
- 238000007599 discharging Methods 0.000 description 1
- 239000002552 dosage form Substances 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 235000013601 eggs Nutrition 0.000 description 1
- 239000000284 extract Substances 0.000 description 1
- 235000011194 food seasoning agent Nutrition 0.000 description 1
- 238000005755 formation reaction Methods 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- UFHFLCQGNIYNRP-UHFFFAOYSA-N hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 238000010921 in-depth analysis Methods 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 238000009114 investigational therapy Methods 0.000 description 1
- GUBGYTABKSRVRQ-XLOQQCSPSA-N lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 230000001294 luteotrophic Effects 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 230000005012 migration Effects 0.000 description 1
- 238000004806 packaging method and process Methods 0.000 description 1
- 229910052698 phosphorus Inorganic materials 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- OAICVXFJPJFONN-UHFFFAOYSA-N phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 1
- XNGIFLGASWRNHJ-UHFFFAOYSA-L phthalate(2-) Chemical compound [O-]C(=O)C1=CC=CC=C1C([O-])=O XNGIFLGASWRNHJ-UHFFFAOYSA-L 0.000 description 1
- 229920000435 poly(dimethylsiloxane) Polymers 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 230000035935 pregnancy Effects 0.000 description 1
- 239000003755 preservative agent Substances 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 231100000486 side effect Toxicity 0.000 description 1
- 239000000344 soap Substances 0.000 description 1
- 238000001694 spray drying Methods 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 230000000576 supplementary Effects 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
Abstract
The invention discloses compositions of glycoprotein of a kind of low solvent residue and preparation method thereof, in described glycoprotein compositions, organic solvent content is less than 0.5%；Described method includes step: (a) by the precipitate of glycoprotein compositions with water system 0 25 DEG C of common vacuum drying；B () continues the compositions that vacuum drying obtains the glycoprotein of low solvent residue after removing water system.
Description
Technical field
The present invention relates to the purification art of pharmaceutical grade protein.Particularly relate to the dry side containing the glycoprotein treating infertility
Method.
Background technology
The glycoprotein for the treatment of infertility is the material that a class formation is close, including chorionic-gonadotropin hormone (HCG), menopause
Phase gonadotropin (HMG), follicule-stimulating hormone (FSH) (FSH), lutropin (LH), wherein HMG is to sting containing follicle
Hormone and lutropin and the mixture of both proportional (1: 0.1-1).
HCG, FSH, LH are to be combined by the form of non-covalent bond by α chain and two subunits of β chain, and wherein their α is sub-
Base is identical, has 92 aminoacid, and molecular weight is about 14500D, and the agedoite on the 52nd and 78 positions is that N-sugar occurs
The aminoacid of base.The β subunit of HCG has 145-147 aminoacid, molecular weight 22200-39000D, wherein the 13rd, on 30 positions
Agedoite and the 121st, 127,132,145 positions be occur glycosylated place.The β subunit of FSH is by 111 aminoacid
Composition, molecular weight is about 18000D, and wherein the agedoite on the 7th and 24 positions is that the glycosylated aminoacid of N-occurs.And LH
β subunit be made up of 121 aminoacid, molecular weight is about 14800D.
HCG, HMG, FSH, LH are mainly used in treating infertility and external supplementary reproduction clinically.They can be from specific
The urine of women's (pregnancy period or menopause) extracts, it is possible to prepared by DNA recombinant technique.
The first generation product of glycoprotein for the treatment of infertility be the Serono company sixties in last century Profasi (HCG),
Pergonal (HMG), their purity is the most relatively low, containing a large amount of impurity.After the eighties in last century, Serono company pushes away respectively
Having gone out Metrodin-HP, it is the highly purified FSH preparation that a kind of impurity content is the lowest；Release the most again and utilize DNA restructuring skill
Gonal-F (rFSH), the Luveris (rLH) etc. that art produces, it addition, Ferring company is also proposed high-purity menopausal rush property
Parathyrine-Menopur.
As can be seen here, the most all it is being devoted to the development and application of high-purity glycoprotein, is substituting common low
Purity product, with the anaphylaxis overcoming the foreign protein in common low-purity product to cause.
CN1309567A discloses the liquid preparation containing FSH or its variant.But, because liquid form must preserve
Below-20 DEG C, otherwise glycoprotein easily inactivates, and liquid form is not easy storage and transport；Liquid form freezes owing to running into
Knot-course of dissolution, the most repeatedly freeze-course of dissolution, it is easier to cause glycoprotein to inactivate；Liquid form further encounters bag
Packaging container is at low temperatures close to the fragility crackly danger of point etc..And for finished dosage form, liquid form is not except allowing
Easily outside transport, also run into the less stable of the glycoprotein of liquid form and it is necessary to adding preservative agent guarantee to exist
In effect duration, microorganism will not be grown, and brings the hidden danger of safety to clinical practice.
Therefore, the glycoprotein compositions of solid form is more suitable for industrialized production.And being mainly obtained of solid form is logical
Cross lyophilization and vacuum drying means, but highly purified glycoprotein is easy to occur degeneration to lose during lyophilization
Living, its deactivation mainly shows as entire molecule and is degraded into α chain and two subunits of β chain, and these subunits are also impurity, they
It is the isomeric compound maybe may having side effects that is of no curative effect, the amount of 10-30% will be added when feeding intake, to offset lyophilization
The inactivation of process, and finished product also can produce a certain amount of subunit, reduce the purity of product.CN101347613A is public
Open a kind of freeze drying process preparing glycoprotein compositions and can prepare the highly purified glycoprotein lyophilizing group being practically free of subunit
Compound, but the effect with this method removal organic solvent is unsatisfactory.
And be vacuum dried and be typically fully to mix organic solvent and glycoprotein solution, it is dehydrated again, then after forming precipitation
Precipitation is vacuum dried, although this method can reduce glycoprotein deactivation, but in dried glycoprotein
Organic solvent content is higher, and weight/mass percentage composition is more than 1%.Organic solvent is the most harmful, Chinese Pharmacopoeia in 2010
In clear stipulaties medicine, ethanol limit is 0.5%.
Therefore, this area a kind of can reduce the vacuum drying side of organic solvent residual in glycoprotein in the urgent need to developing
Method, and it is derived from the glycoprotein compositions of low organic solvent residual.
Summary of the invention
It is desirable to provide the glycoprotein compositions of a kind of low organic solvent residual, and obtain the combination of above-mentioned glycoprotein
The vacuum drying method of thing.
One aspect of the present invention provides the glycoprotein compositions of a kind of low organic solvent content, wherein organic solvent quality hundred
Divide content less than 0.5%, preferably more than 0.3%, be more preferably less than 0.1%.
Wherein, described organic solvent is ethanol, acetone or methanol.
Wherein, described glycoprotein is selected from chorionic-gonadotropin hormone, HMG, follicule-stimulating hormone (FSH), corpus luteum
Generate element or its mixing.
Wherein, described chorionic-gonadotropin hormone be Urina Hominis source and/or restructuring human chorionic gonadotropin or its
Variant；Described HMG be Urina Hominis source and/or restructuring human menopausal gonadotropin or its variant；Described ovum
Bubble stimulin be Urina Hominis source and/or restructuring HFSH or its variant；Described lutropin is Urina Hominis source
And/or restructuring human luteinizing hormone or its variant.
Another aspect of the present invention, it is provided that the side of a kind of glycoprotein compositions simply preparing low solvent residue
Method.Inventor, through further investigation, finds, by glycoprotein compositions and water system altogether same vacuum drying, can effectively remove organic
Solvent, obtains the glycoprotein compositions with the low solvent residue of good stability.
The drying principles of the compositions of the glycoprotein of low solvent residue
Owing to glycoprotein material at high temperature changeableness inactivates, therefore spray drying or the mode of other heating should not be used
Being dried, and cryodesiccated mode there is also certain deactivation phenomenom, and freeze drying equipment is expensive, dry amount is little,
Energy consumption is big.Use boulton process (being not added with aqueous systems to be dried) product degraded inactivation inconspicuous, but organic solvent residual is high.Invention
The reason that its dissolvent residual is exceeded standard by people conducts in-depth analysis, at sample vacuum drying early stage organic solvent and water with necessarily
Ratio jointly volatilize, but along with organic solvent and the reduction of water content in sample, the amount of volatilization is more and more less, when in sample
Water content when no longer reducing, the amount of organic solvent the most no longer reduces.Therefore the residual quantity of organic solvent contains with the water in sample
Measure relevant.Inventor, it is surprisingly found that place in vacuum desiccator and can discharge the material of water vapour, is not only able to obtain low
The compositions of the glycoprotein of dissolvent residual, and there is not degraded deactivation phenomenom in sample.
Its drying principles is conducted in-depth research after obtaining the compositions of glycoprotein of low solvent residue by inventor.?
Vacuum desiccator is placed and can discharge the material of water vapour in order to control in process of vacuum drying in sample is organic
Solvent and the volatilization ratio of water.In process of vacuum drying, organic solvent volatilizees jointly with the water in sample, in vacuum drying oven,
Organic solvent and steam respectively account for a certain proportion of, but along with dry carrying out, moisture is limited, in order to maintain this to put down
Weighing apparatus, we put into the material that can discharge water vapour in dry system, artificially increase vapour pressure in vacuum drying oven, can suppress
The volatilization of the water in sample, can promote again the volatilization of organic solvent in sample simultaneously.Thus effectively reduce in sample organic
The residual of solvent, and the water content in sample is also maintained at reduced levels, therefore there is not degraded deactivation phenomenom in sample.
Its dry run is goed deep into after obtaining the compositions of glycoprotein of low solvent residue by inventor once more
Research, again it has surprisingly been found that when organic solvent be dried qualified after, then remove the material discharging water vapour, continue to be vacuum dried
Certain time, the moisture in sample can be vacuum dried in acceptability limit, and organic solvent can also reduce further, sample
Have good stability in the range of this moisture content.Therefore the dry run of the present invention be divided into two benches, first stage be by sample with
Aqueous systems is vacuum dried removing organic solvent jointly, and second stage is to remove aqueous systems to be vacuum dried that sample moisture content is qualified is
Only.
Preparation method
A kind of method that the invention provides glycoprotein compositions preparing low organic solvent content, described method include with
Lower step:
A the solution of organic solvent with the composition material containing glycoprotein is sufficiently mixed precipitation by (), collect precipitate
(1), after dehydration of organic solvent, continue to collect the composition precipitates of glycoprotein, it is thus achieved that containing the precipitate of glycoprotein compositions
(2)；With
B (), by the precipitate (2) containing glycoprotein compositions and water system altogether same vacuum drying, obtains what the present invention provided
The glycoprotein compositions of low organic solvent residual.
In another preference, in step (a), described organic solvent temperature-20 to 25 DEG C, preferably-20 to 0 DEG C.
In another preference, in step (a), described organic solvent is selected from ethanol, acetone or methanol.
In another preference, in step (a), described dehydration organic solvent amount is moist precipitate thing (1) weight 0 to 10
Times, preferably 1 to 4 times.
In another preference, in step (b), described baking temperature at 0 to 25 DEG C, preferably 0 to 20 DEG C.
In another preference, in step (b), described water system include solid-state, liquid or gaseous state tap water, go from
Sub-water, reverse osmosis water, pure water and water for injection, can discharge the object of water vapour, and aqueous solution, preferably purified water.
In another preference, in step (b), described drying time is 1 to 48 hour, preferably 10 to 24 hours.
In another preference, the solution containing glycoprotein compositions raw material described in step a can be by following steps system
Standby:
By the material dissolution containing glycoprotein compositions in water or buffer, preparation is containing glycoprotein compositions raw material
Solution.
In another preference, the described raw material containing glycoprotein compositions is solid.
In another preference, described buffer is selected from: pH value buffering range 3 to 11 buffer；It preferably is selected from；Phosphorus
Phthalate buffer, Tris buffer, sodium-acetate buffer.
In another preference, described solution temperature at 5 to 20 DEG C, preferably 10 to 16 DEG C.
In another preference, the pH value of the described solution containing glycoprotein compositions raw material is 3 to 11, preferably 4 to 10.
In another preference, following steps can also be contained below in step (b):
C () continues the combination that vacuum drying obtains the glycoprotein of the low solvent residue that the present invention provides after removing water system
Thing.
In step (c), described drying time is 0 to 48 hour, preferably 5 to 24 hours.
In a preference of the present invention, the method for the glycoprotein compositions preparing low organic solvent content comprises following
Step:
(a ') it is sufficiently mixed precipitation with the solution of the organic solvents of-20～0 DEG C with the composition material containing glycoprotein, receive
Taking precipitate (1)；
(b ') by precipitate (1), after dehydration of organic solvent, continue to collect the composition precipitates of glycoprotein, it is thus achieved that contain
The precipitate (2) of glycoprotein compositions；
(c ') by the composition precipitates thing (2) of glycoprotein and water system 0～25 DEG C of common vacuum drying 1～48 hours,
Obtain the compositions with the low solvent glycoprotein of good stability；With
(d '), after step (c '), removes water system, continues, in 0～25 DEG C, the low solvent that vacuum drying step (c ') prepares
The compositions 0 of glycoprotein～48 hours, obtain the compositions of the glycoprotein of more low solvent residue.
As used herein, " glycoprotein " is selected from one or more following mixing: chorionic-gonadotropin hormone
(chorionic gonadotropin, CG), follicule-stimulating hormone (FSH) (follicule-stimulatinghormone, FSH), corpus luteum
Generation element (luteotropic hormone, LH), HMG (human menopausal gonadotropin,
HMG)。
As used herein, " glycoprotein compositions raw material " refers to the compositions containing glycoprotein, including glycoprotein, or
Person includes acceptable carrier in glycoprotein and pharmaceutically/bromatology.The existing glycoprotein of preparing in this area can be used to combine
The glycoprotein compositions that the method for thing is obtained, as raw material, can be solid or liquid, such as but not limited to, Chinese patent
The highly purified HMG that preparation method disclosed in CN1246332C obtains；Preparation method disclosed in Chinese patent CN1958603B obtains
The HCG obtained；And the FSH that method disclosed in CN101317103A obtains.
The acquisition of above glycoprotein compositions raw material is only citing, and glycoprotein compositions raw material of the present invention should not
It is so limited.
In the method for the glycoprotein compositions obtaining low solvent residue of present invention offer, described " will be containing sugar egg
The precipitate (2) of white compositions and water system are altogether with being vacuum dried " refer to place the precipitate (2) containing glycoprotein compositions
Generally place the place of sample at vacuum desiccator, and (such as bottom) is placed to contain and can be discharged water and steam around vacuum desiccator
The container opened wide of the material of vapour, the described material that can discharge water vapour is selected from tap water, pure water or mixture of ice and water.
In the method for the glycoprotein compositions obtaining low solvent residue of present invention offer, described " removing aqueous systems "
Outside referring to contain the vacuum drying environment of container migration opened wide of the material that can discharge water vapour.
Main advantages of the present invention are:
1, the invention provides the compositions of the glycoprotein of a kind of new low solvent residue.
2, the invention provides the preparation method of the compositions of the glycoprotein of a kind of new low solvent residue.
3, the present invention has mild condition, operating process is simple, degraded deactivation phenomenom is little, and product stability is good, and energy consumption is low,
Alleviate technological operation degree-of-difficulty factor to a great extent, reduce production cost.
Accompanying drawing explanation
Fig. 1 shows the GC collection of illustrative plates of the standard substance that ethanol content is 0.5%；Wherein the retention time of solvent, peak area and
Mass content is as shown in the following chart:
| Retention time (minute) | Peak area | Mass content (%) | Solvent species |
| 3.175 | 1040.5 | 0.5 | Ethanol |
Fig. 2 shows the ethanol content GC collection of illustrative plates in comparative example 2 in HMG sample；The wherein retention time of solvent, peak area
With mass content as shown in the following chart:
| Retention time (minute) | Peak area | Mass content (%) | Solvent species |
| 3.175 | 2368.5 | 1.1 | Ethanol |
Fig. 3 shows the ethanol content GC collection of illustrative plates in embodiment 6 in HMG sample；The wherein retention time of solvent, peak area
With mass content as shown in the following chart:
| Retention time (minute) | Peak area | Mass content (%) | Solvent species |
| 3.175 | 69.6 | 0.03 | Ethanol |
Fig. 4 shows the ethanol content GC collection of illustrative plates in embodiment 9 in HCG sample；The wherein retention time of solvent, peak area
With mass content as shown in the following chart:
| Retention time (minute) | Peak area | Mass content (%) | Solvent species |
| 3.175 | 414.1 | 0.20 | Ethanol |
Detailed description of the invention
The compositions organic solvent residual of glycoprotein measures
In embodiments of the present invention, GC is used to record the residual solvent of sample prepared by the inventive method, described
GC detection method is as follows:
Chromatographic column: Varian CP-Select 624 (94% cyanogen propyl group phenyl-6% dimethyl polysiloxane)
Post specification: 30m × 0.32mm × 1.8 μm
Injector temperature: 200 DEG C
Detector temperature: 260 DEG C
Flow velocity: 2.5ml/ minute, constant current
Furnace temperature:
Carrier gas: nitrogen
Split ratio: 1: 1 (work station), actual split ratio: 17: 1 (soap film flowmeter measured values)
Detector gas: air mass flow: 400ml/ minute
Hydrogen flowing quantity: 40ml/ minute
Make-up gas flow: 25ml/ minute (nitrogen)
Head-space sampler condition
Heating equilibrium temperature: 80 DEG C；Quantitative loop temperature: 85 DEG C；Transmission line temperature: 90 DEG C；Equilibration time: 20 minutes；Add
The pressure time: 0.5 minute；Fill the sample time: 0.2 minute；Quantitative loop equilibration time: 0.10 minute；Sample injection time: 1 minute；Whole fortune
The row time: 30 minutes；Sampling volume: 1ml；Shaking pattern: slight vibration.
Below in conjunction with specific embodiment, the present invention is expanded on further.Should be understood that these embodiments are merely to illustrate the present invention
Rather than restriction the scope of the present invention.The experimental technique of unreceipted actual conditions in the following example, generally according to conventional strip
Part, or according to the condition proposed by manufacturer, unless otherwise indicated, otherwise percentage ratio and number are calculated by weight.
Unless otherwise defined, the meaning that all specialties used in literary composition are familiar with one skilled in the art with scientific words
Justice is identical.Additionally, any method sill similar or impartial to described content all can be applicable in the inventive method.Wen Zhong
Described preferable implementation only presents a demonstration with material and is used.
Below in conjunction with specific embodiment, the present invention is expanded on further.Should be understood that these embodiments are merely to illustrate the present invention
Rather than restriction the scope of the present invention.The experimental technique of unreceipted actual conditions in the following example, generally according to conventional strip
Part or according to the condition proposed by manufacturer.Unless otherwise indicated, otherwise percentage ratio and number are calculated by weight.
Unless otherwise defined, the meaning that all specialties used in literary composition are familiar with one skilled in the art with scientific words
Justice is identical.Additionally, any method similar or impartial to described content and material all can be applicable in the present invention.Described in literary composition
Preferable implementation only present a demonstration with material and be used.
Comparative example 1
The lyophilization of FSH
Preparation 45mL 0.01M sodium dihydrogen phosphate (adjusts pH about 6.5 with NaOH), adds 5ml ethanol, adds 2g lactose,
After stirring and dissolving, filter with 0.22 μm filter.Take 10mL filtered solution, add (the big life in sky, Shanghai of above-mentioned high-purity FSH of 10.0mg
Thing pharmaceutical Co. Ltd provides, and biological value is 8817 ius/mg), after being completely dissolved, sabot is put into freeze dryer and (is purchased from
Virtis), in, carry out lyophilization, outlet by the method for CN101347613A embodiment 1, obtain 391mg freeze-dried powder.
After measured, the biological value result of FSH is as follows:
Comparative example 2
Traditional vacuum seasoning prepares HMG
The sodium acetate solution of preparation 10ml 0.1mol/L, regulates pH6.5 with acetic acid, is cooled to 10 DEG C, adds 250mgHMG
Raw material (is provided by Shanghai Tianwei Biological Pharmaceutical Corp., biological value is 405IU/mg in terms of FSH), stirring and dissolving.Add-20
DEG C ethanol 50ml, stirring and evenly mixing, centrifugal to obtain moist precipitate 931mg, to use 3ml ethanol dehydration, centrifugal to obtain HMG moist precipitate 901mg, 20
It is vacuum dried 96 hours at DEG C, obtains 249mgHMG sample.After measured, biological value (in terms of FSH) and solvent result
As follows:
Embodiment 3
Prepare low solvent residue HMG
The sodium acetate solution of preparation 10ml 0.1mol/L, regulates pH4.0 with acetic acid, is cooled to 16 DEG C, adds 250mgHMG
Raw material (is provided by Shanghai Tianwei Biological Pharmaceutical Corp., biological value is 405IU/mg in terms of FSH), stirring and dissolving.Add-20
DEG C ethanol 100ml, stirring and evenly mixing, centrifugal to obtain HMG moist precipitate 936mg, with the aqueous systems one being placed with mixture of ice and water at 20 DEG C
Play vacuum drying 10 hours, obtain 257mgHMG sample.After measured, biological value (in terms of FSH) and solvent result are such as
Under:
Result is as follows:
Embodiment 4
Prepare low solvent residue HMG
The sodium acetate solution of preparation 10ml 0.1mol/L, regulates pH6.5 with acetic acid, is cooled to 10 DEG C, adds 250mgHMG
Raw material (is provided by Shanghai Tianwei Biological Pharmaceutical Corp., biological value is 405IU/mg in terms of FSH), stirring and dissolving.Add-20
DEG C ethanol 50ml, stirring and evenly mixing, centrifugal to obtain moist precipitate 956mg, to use 10ml ethanol dehydration, centrifugal to obtain HMG moist precipitate 916mg,
10 DEG C are vacuum dried together with the aqueous systems being placed with mixture of ice and water 24 hours, obtain 252mgHMG sample.After measured, biological effect
Valency (in terms of FSH) and solvent result are as follows:
Result is as follows:
Embodiment 5
Prepare low solvent residue HMG
The sodium acetate solution of preparation 10ml 0.1mol/L, regulates pH10 with acetic acid, is cooled to 15 DEG C, adds 250mgHMG
Raw material (is provided by Shanghai Tianwei Biological Pharmaceutical Corp., biological value is 405IU/mg in terms of FSH), stirring and dissolving.Add-10
DEG C ethanol 100ml, stirring and evenly mixing, centrifugal to obtain HMG moist precipitate 953mg, 0 DEG C together with the aqueous systems being placed with purified water vacuum do
Dry 24 hours, removing water system, 0 DEG C is continued to be dried 5 hours, obtains 246mgHMG sample.After measured, biological value is (with FSH
Meter) and solvent result as follows:
Result is as follows:
Embodiment 6
Prepare low solvent residue HMG
The sodium dihydrogen phosphate of preparation 10ml 0.1mol/L, regulates pH7.0 by NaOH solution, is cooled to 10 DEG C, adds
250mgHMG raw material (is provided by Shanghai Tianwei Biological Pharmaceutical Corp., biological value is 405IU/mg in terms of FSH), stirs molten
Solve.Add 0 DEG C of ethanol 50ml, stirring and evenly mixing, be centrifuged and to obtain moist precipitate 952mg, use 3ml ethanol dehydration, be centrifuged and to obtain HMG moist precipitate
932mg, is vacuum dried 24 hours at 0 DEG C together with the aqueous systems being placed with purified water, removes water system, and 20 DEG C to continue to be dried 24 little
Time, obtain 244mgHMG sample.After measured, biological value (in terms of FSH) and solvent result are as follows:
Result is as follows:
Embodiment 7
Prepare low solvent residue HCG
The Tris solution of preparation 10ml 0.1mol/L, regulates pH7.5 with hydrochloric acid, is cooled to 10 DEG C, adds 300mgHCG former
Material (being provided by Shanghai Tianwei Biological Pharmaceutical Corp., biological value 5702IU/mg), stirring and dissolving.Add 0 DEG C of ethanol
50ml, stirring and evenly mixing, it is centrifuged and to obtain moist precipitate 1172mg, use 11ml ethanol dehydration, be centrifuged and to obtain HCG moist precipitate 1022mg, at 10 DEG C
Being vacuum dried together with the aqueous systems being placed with mixture of ice and water 24 hours, remove water system, 20 DEG C are continued to be dried to obtain 24 hours,
To 295mgHCG sample.After measured, biological value and solvent result are as follows:
Result is as follows:
Embodiment 8
Prepare the FSH of low solvent residue
Take 10 DEG C of solution 1000ml containing FSH raw material (to be provided by Shanghai Tianwei Biological Pharmaceutical Corp., biological value
1958IU/ml), add-20 DEG C of ethanol 5000ml, stirring and evenly mixing, be centrifuged to obtain moist precipitate 15820mg, use 48ml ethanol dehydration, from
Gains in depth of comprehension FSH moist precipitate 14958mg, is vacuum dried together with the aqueous systems being placed with purified water 24 hours at 0 DEG C, removes water system,
20 DEG C are continued to be dried 24 hours, obtain 3995mg FSH sample.After measured, the biological value of FSH and solvent result are such as
Under:
Embodiment 9
Prepare the HCG of low solvent residue
Take 16 DEG C of solution 10L containing HCG raw material (to be provided by Shanghai Tianwei Biological Pharmaceutical Corp., biological value
68319IU/ml), add-10 DEG C of ethanol 50L, stirring and evenly mixing, be centrifuged and to obtain moist precipitate 620g, use 4L ethanol dehydration, be centrifuged and to obtain HCG
Moist precipitate 560g, is vacuum dried 20 hours at 20 DEG C together with the aqueous systems being placed with purified water, removes water system, and 20 DEG C are continued dry
Dry 20 hours, obtain the HCG sample of 122g.
After measured, biological value and the solvent result of HCG are as follows:
Embodiment 10
Prepare low solvent residue HCG
According to the step operation of embodiment 7, difference be with the ethanol in methanol alternate embodiment 7 raw material is precipitated,
Moist precipitate is prepared in dehydration, obtains 298mgHCG sample.After measured, biological value and solvent result are as follows:
Result is as follows:
Embodiment 11
Prepare low solvent residue HMG
According to the step operation of embodiment 6, difference be with the ethanol in acetone alternate embodiment 6 raw material is precipitated,
Moist precipitate is prepared in dehydration, obtains 240mgHMG sample.After measured, biological value (in terms of FSH) and solvent result are as follows:
Result is as follows:
The foregoing is only presently preferred embodiments of the present invention, be not limited to the substantial technological content model of the present invention
Enclosing, the substantial technological content of the present invention is broadly to be defined in the right of application, any technology that other people complete
Entity or method, if with the right of application defined in identical, also or the change of a kind of equivalence, all quilt
It is considered as being covered by among this right.
Claims (24)
1. the preparation method of the glycoprotein compositions of a low solvent residue, it is characterised in that described method includes step:
A the solution of organic solvent with the composition material containing glycoprotein is sufficiently mixed precipitation by (), collect precipitate (1), uses
After dehydration of organic solvent, continue to collect the composition precipitates of glycoprotein, it is thus achieved that the precipitate (2) containing glycoprotein compositions, institute
The glycoprotein stated is selected from chorionic-gonadotropin hormone, HMG, follicule-stimulating hormone (FSH), lutropin or its mixing；
B (), by the precipitate (2) containing glycoprotein compositions and water system altogether same vacuum drying, obtains described low solvent residue
Glycoprotein compositions, in the glycoprotein compositions of this low solvent residue, organic solvent weight/mass percentage composition is less than 0.5%；Described
The content of organic solvent is measured by GC method.
2. preparation method as claimed in claim 1, it is characterised in that in step a, described dehydration organic solvent amount is wet heavy
0 to 10 times of shallow lake thing (1) weight.
3. preparation method as claimed in claim 2, it is characterised in that in step a, described dehydration organic solvent amount is wet heavy
1 to 4 times of shallow lake thing (1) weight.
4. preparation method as claimed in claim 1, it is characterised in that in step a, described organic solvent is selected from ethanol, acetone
Or methanol.
5. preparation method as claimed in claim 1, it is characterised in that in step a, described organic solvent temperature is-20 to 25
℃。
6. preparation method as claimed in claim 5, it is characterised in that in step a, described organic solvent temperature is-20 to 0
℃。
7. preparation method as claimed in claim 1, it is characterised in that in step b, described vacuum drying temperature is 0 to 25
℃。
8. preparation method as claimed in claim 7, it is characterised in that in step b, described vacuum drying temperature is 0 to 20
℃。
9. preparation method as claimed in claim 1, it is characterised in that in step b, the described vacuum drying time be 1 to
48h。
10. preparation method as claimed in claim 9, it is characterised in that in step b, the described vacuum drying time be 10 to
24h。
11. preparation methoies as claimed in claim 1, it is characterised in that in stepb, the water in described water system is from the beginning
Water, deionized water, reverse osmosis water, pure water or water for injection.
12. preparation methoies as claimed in claim 11, it is characterised in that in step b, the water in described water system be liquid,
Solid-state, gaseous state or mixed style.
13. preparation methoies as claimed in claim 12, it is characterised in that in step b, described water system is aqueous solution.
14. preparation methoies as claimed in claim 12, it is characterised in that in step b, described water system is steamed for producing water
The object of gas.
15. preparation methoies as claimed in claim 1, it is characterised in that described in step a containing glycoprotein compositions raw material
Solution prepared by following steps:
By the material dissolution containing glycoprotein compositions in water or buffer, preparation is molten containing glycoprotein compositions raw material
Liquid.
16. preparation methoies as claimed in claim 15, it is characterised in that described in be dissolved at temperature 5 to 20 DEG C and carry out.
17. preparation methoies as claimed in claim 16, it is characterised in that described temperature is at 10 to 16 DEG C.
18. preparation methoies as claimed in claim 15, it is characterised in that described buffer is selected from: pH value buffering range is 3
To the buffer of 11.
19. preparation methoies as claimed in claim 18, it is characterised in that described buffer is selected from: phosphate buffer,
Tris buffer, sodium-acetate buffer.
20. preparation methoies as claimed in claim 15, it is characterised in that the described solution containing glycoprotein compositions raw material
PH value controls 3 to 11.
21. preparation methoies as claimed in claim 20, it is characterised in that the described solution containing glycoprotein compositions raw material
PH value controls 4 to 10.
22. preparation methoies as claimed in claim 1, it is characterised in that face is possibly together with following steps after the stepb:
C () is continued vacuum drying after removing water system and is obtained the glycoprotein compositions of described low solvent residue.
23. preparation methoies as claimed in claim 22, it is characterised in that in step c, the described vacuum drying time be 0 to
48h。
24. preparation methoies as claimed in claim 23, it is characterised in that in step c, the described vacuum drying time be 5 to
24h。
Priority Applications (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201110102073.8A CN102743742B (en) | 2011-04-22 | A kind of glycoprotein compositions of low solvent residue and its production and use | |
| KR1020137031113A KR20140018970A (en) | 2011-04-22 | 2012-04-20 | Glycoprotein composition with low residual solvent level, and preparation method and use thereof |
| PCT/CN2012/074425 WO2012142961A1 (en) | 2011-04-22 | 2012-04-20 | Glycoprotein composition with low residual solvent level, and preparation method and use thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201110102073.8A CN102743742B (en) | 2011-04-22 | A kind of glycoprotein compositions of low solvent residue and its production and use |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN102743742A CN102743742A (en) | 2012-10-24 |
| CN102743742B true CN102743742B (en) | 2016-12-14 |
Family
ID=
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1302818A (en) * | 2000-12-12 | 2001-07-11 | 上海惠海生化制品厂 | Human chorionic gonadotropin and its preparing process |
| CN101307103A (en) * | 2007-09-11 | 2008-11-19 | 上海天伟生物制药有限公司 | Purification method of follicle stimulating hormone |
| CN101555279A (en) * | 2009-05-19 | 2009-10-14 | 上海天伟生物制药有限公司 | Urinary follicle stimulating hormone with high purity and preparation method thereof |
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1302818A (en) * | 2000-12-12 | 2001-07-11 | 上海惠海生化制品厂 | Human chorionic gonadotropin and its preparing process |
| CN101307103A (en) * | 2007-09-11 | 2008-11-19 | 上海天伟生物制药有限公司 | Purification method of follicle stimulating hormone |
| CN101555279A (en) * | 2009-05-19 | 2009-10-14 | 上海天伟生物制药有限公司 | Urinary follicle stimulating hormone with high purity and preparation method thereof |
Non-Patent Citations (1)
| Title |
|---|
| 高单位尿促性素研制初报;陈少雄;《中国生化药物杂志》;20011231;第22卷(第4期);201-202 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN100484525C (en) | Pantoprazole sodium freeze-dried powder injection and preparing method thereof | |
| CN102512466B (en) | Panax notoginseng saponins freeze-dried powder injection and preparation method thereof | |
| CN100506217C (en) | Ambroxol hydrochloride freeze-dried powder injection and preparing method thereof | |
| CN102219851A (en) | Preparation method for insulin glargine crystals | |
| CN113930472B (en) | Preparation method and product of donkey-hide gelatin peptide with low pro-inflammatory reaction | |
| CN101703484B (en) | Preparation method of hexadecadrol sodium phosphate freeze-dried powder injection | |
| CN102178656A (en) | HM-3 polypeptide freeze-dried powder preparation and preparation method thereof | |
| CN101947208B (en) | Fludarabine phosphate composition for injection and preparation method thereof | |
| CN103088096A (en) | Method for extracting collagen type II by taking agkistrodon as raw material | |
| CN102743742B (en) | A kind of glycoprotein compositions of low solvent residue and its production and use | |
| CN102058548B (en) | Ambroxol hydrochloride composition for injection and preparation method thereof | |
| CN102091046A (en) | Fludarabine phosphate freeze-dried powder injection and preparation method thereof | |
| CN101190232B (en) | Magnesium isoglycyrrhizinate freeze dried powder injection for injection and preparation method thereof | |
| CN102697740B (en) | Gemcitabine hydrochloride lyophilized powder for injection and preparation method thereof | |
| CN102846563B (en) | Injection composition of pemetrexed disodium and preparation method of injection composition | |
| CN100417381C (en) | Nicorandil freeze-drying powder preparation method | |
| CN102429942B (en) | Freeze-drying method of pseudo-ginseng total saponin freeze-dried injection powder with filling amount more than 500 mg | |
| CN114213512A (en) | Composition for enhancing phycobiliprotein photo-thermal stability and preparation method and application thereof | |
| CN102743742A (en) | Glycoprotein composition with low solvent residues, its preparation and its application | |
| CN103655490A (en) | Idarubicin hydrochloride pharmaceutical composition and preparation method thereof | |
| CN101317851A (en) | Freeze drying prescription containing glycyrrhizin and preparation method thereof | |
| CN103655489A (en) | Epirubicin hydrochloride pharmaceutical composition and preparation method thereof | |
| CN108047306A (en) | A kind of grifola frondosus selenium chelating peptide and preparation method thereof | |
| CN103495152B (en) | Thymopentin-containing medicine composition, as well as preparation and preparation method thereof | |
| CN103432084B (en) | Preparation method of thymopentin freeze-dried powder injection |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant |