CN107991425A

CN107991425A - A kind of detection method for the Chinese medicine composition for treating traumatic injury

Info

Publication number: CN107991425A
Application number: CN201711291628.1A
Authority: CN
Inventors: 田涵雯; 王士青
Original assignee: CHANGCHUN RENMIN PHARMACEUTICAL GROUP Co Ltd
Current assignee: CHANGCHUN RENMIN PHARMACEUTICAL GROUP Co Ltd
Priority date: 2017-12-07
Filing date: 2017-12-07
Publication date: 2018-05-04
Anticipated expiration: 2037-12-07
Also published as: CN107991425B

Abstract

The present invention relates to a kind of detection method for the Chinese medicine composition for treating traumatic injury, belong to Chinese medicine, natural medicine technical field.On the basis of the indentification by TLC and aconitine limit of Angelica sinensis, borneol, cape jasmine, pseudo-ginseng and vinegar corydalis tuber check, the thin layer of frankincense and chrysanthemum differentiates, using high-efficient liquid phase chromatogram technology, its monarch drug in a prescription pseudo-ginseng content assaying method is improved；The content assaying method of toxicity medicinal material cinnabar has been created at the same time.The present invention solves said composition due to the relatively low caused unilateral fakement phenomena using single extract substitution valuable medicinal of control method, improve clinical safety and validity at the same time, this method is applicable not only to overall quality control of the industrialized production to this kind of composition, also provides safe and effective guarantee for clinical application.

Description

A kind of detection method for the Chinese medicine composition for treating traumatic injury

Technical field

The invention belongs to Chinese medicine, natural medicine technical field, and in particular to a kind of Traditional Chinese medical composition for treatment of traumatic injury for the treatment of Detection method.

Background technology

Traumatic injury refer mainly to because fall flutter, impact etc. caused by soft tissue injury, wound swelling pain, skin and flesh breakage go out Blood, also including tumble injury etc..Its main pathology is stopped up stagnant for extravasated blood, and blood is held one's breath resistance, therefore using pain, swelling as main clinical manifestation.With Promoting blood circulation and removing blood stasis, swelling and pain relieving for curative effect Chinese medicine composition be by pseudo-ginseng, wild aconite root, aconiti preparata,radix, lotus nut, Angelica sinensis, scald The rhizome of davallia, grass-leaved sweetflag, dandelion, field thistle, stir-baked OLIBANUM with vinegar, stir-baked MYRRHA with vinegar, hairyvein agrimony, borneol, safflower, chrysanthemum, cape jasmine, Paris polyphylla, cinnabar, Ten nine traditional Chinese medicines material prescription of vinegar corydalis tuber forms.Pseudo-ginseng is reused in side, can hemostasia and dissipation blood stasis, swelling and pain relieving is monarch drug in a prescription；It is equipped with system grass Crow, aconiti preparata,radix, Angelica sinensis, field thistle, stir-baked OLIBANUM with vinegar, stir-baked MYRRHA with vinegar, hairyvein agrimony, eight taste of safflower can principal drug assistance it is promoting blood circulation and removing blood stasis, hemostasia and detumescence, altogether For ministerial drug.Borneol energy clearing away heat to and alleviating pain, fragrance are walked to alter, sensible channels and collaterals；Lotus nut, dandelion, chrysanthemum, cape jasmine can clearing heat-fires；Stone Calamus, Paris polyphylla, cinnabar can tranquilizing and allaying excitement, alleviate traumatic injury caused by pain, be altogether adjutant.All medicines share, plays promoting blood circulationization altogether The effect of stasis of blood, swelling and pain relieving.

Said composition derives from folk remedy, is initially applied to clinic with powder form, coating is made by modern study Tablet (Changchun red tablet) and hard capsule (Changchun Hongyao capsule) are widely used in clinic.Changchun red tablet quality standard is received It is loaded in the 3rd (standard No. of ministry standard Traditional Chinese medicine historical preparation：WS3-B-0504-91), the standard is except character and disintegration time limited inspection Test outside project, without other control items, standard controllability is poor.And Changchun Hongyao capsule is the another of the enterprise development on the basis of tablet One dosage type low temperature product, performs the national registration standard (standard No. that State Food and Drug Administration issues： WS-11024(ZD- 1024)-2002-2013Z)；The thin-layer qualitative that 5 taste medicinal materials are added in standard differentiates, in aconitine limit inspection and pseudo-ginseng Single component ginsenoside Rg₁Quantitative control；Compared with tablet, its quality control has larger improvement, but the controlling party Method is only applicable to the quality control of capsule, is not restricted for other formulations.Said composition flavour of a drug are more, wherein monarch drug in a prescription pseudo-ginseng Medicinal material effective ingredient is more complicated, and the quantitative control of single component can not fully demonstrate the quality condition of whole preparation；And group The cinnabar of medicinal material containing toxicity in polymer components, current standard do not carry out it quantitative control, and security cannot ensure very well.In order to Preferably control this kind of composition quality, is avoided valuable medicinal in preparation process from being substituted using single extract, runs counter to original group Square combination principle phenomenon occurs.Enterprise has re-established the new detection method of said composition by technological transformation.

The content of the invention

The present invention provides a kind of detection method for treating Traditional Chinese medical composition for treatment of traumatic injury, to solve piece existing for said composition Face substitutes the fraud problem of valuable medicinal using single extract.

The present invention adopts the technical scheme that：

The Chinese medicine composition is obtained by the following steps：12.1 parts of fine grindings of cinnabar are into impalpable powder；24.2 parts of borneol is ground Wear into fine powder；121.2 parts of pseudo-ginseng, 12.12 parts of aconiti preparata,radix, 20.2 parts of Angelica sinensis, 20.2 parts of stir-baked OLIBANUM with vinegar, 20.2 parts of stir-baked MYRRHA with vinegar, stone Chang 8.1 parts of Pu is ground into fine powder, with Cinnabar facing-up, sieves, mixes；40.4 parts of remaining cape jasmine, 12.12 parts of wild aconite root, scald bone Broken 40.4 parts of benefit, 60.6 parts of dandelion, 6.7 parts of field thistle, 60.6 parts of hairyvein agrimony, 40.4 parts of safflower, 40.4 parts of chrysanthemum, lotus nut 12.12 parts, 13.5 parts of Paris polyphylla, vinegar corydalis tuber 20.2 etc. ten simply medicinal material add water to cook it is secondary, every time 2 it is small when, filter, close by several times And filtrate, it is 1.23~1.25 thick pastes to survey relative density when being condensed into 70~75 DEG C, is mixed with the above-mentioned powder in addition to borneol, It dry, pulverize, mix, add borneol fine powder, mix；

Detection method includes：Angelica sinensis, borneol, cape jasmine, the indentification by TLC and aconitine limit of pseudo-ginseng and vinegar corydalis tuber Inspection method, it is characterised in that further include：

(1) thin layer of stir-baked OLIBANUM with vinegar differentiates, comprises the following steps that：

(a) preparation of test solution：This product 2.5g is taken, is mixed, adds 20~40ml of ethanol, is ultrasonically treated 20~30 points Clock, filtration, filtrate volatilize, and residue adds ethanol 1ml, as test solution；

(b) preparation of control medicinal material solution：Frankincense control medicinal material 0.2g is taken, control is made by test solution preparation method Medicinal material solution；

(c) preparation of negative sample solution：The prescription medicinal material of scarce stir-baked OLIBANUM with vinegar is taken, is pressed in prescription ratio and preparation method for trying Negative controls are made in product solution manufacturing method；

(d) chromatographic condition and operation：Test, draw above-mentioned according to thin-layered chromatography (Chinese Pharmacopoeia version general rule 0502 in 2015) Each 5~10 μ l of three kinds of solution, put in same silica G F respectively₂₅₄On lamellae, with 60 DEG C~90 DEG C petroleum ethers：Thiacyclohexane：Acetic acid Ethyl ester：Formic acid=10:30:15:1 is solvent, is unfolded, and takes out, dries, put and inspected under 254nm ultraviolet lamps；

(e) result feature：In test sample chromatography, on position corresponding with control medicinal material chromatography, the spot of same color is shown Point；

(2), chrysanthemum thin layer differentiates, comprises the following steps that：

(a) preparation of test solution：This product 6g is taken, it is finely ground, add 30~50ml of chloroform, be heated to reflux 30 minutes, Filtration, discards filtrate, and the dregs of a decoction volatilize solvent, add 80~100ml of water, are heated to reflux 40~60 minutes, and centrifugation, takes supernatant, add 0.5~1.0ml of hydrochloric acid, secondary with ethyl acetate shaking extraction, each 40ml, combined ethyl acetate liquid, is evaporated, residue adds methanol 1ml makes solution, as test solution；

(b) preparation of control medicinal material solution：Chrysanthemum control medicinal material 0.5g separately is taken, is made pair by test solution preparation method According to medicinal material solution；

(c) preparation of negative sample solution：The prescription medicinal material of scarce chrysanthemum is taken, test sample is pressed in prescription ratio and preparation process Negative controls are made in solution manufacturing method；

(d) chromatographic condition and operation：Test, draw above-mentioned according to thin-layered chromatography (Chinese Pharmacopoeia version general rule 0502 in 2015) Each 2~5 μ l of three kinds of solution, put on same polyamide film, make into strips, with butyl acetate respectively：Glacial acetic acid：Water=: 2.5:The upper solution of (2.5~3) is solvent, is unfolded, and takes out, dries, put and inspected under 365nm ultraviolet lamps；

(e) result feature：In test sample chromatography, on position corresponding with control medicinal material chromatography, identical phosphor strip is shown Spot；

(3) content assaying method of pseudo-ginseng, step are as follows：

(a) chromatographic condition and system suitability are using octadecylsilane chemically bonded silica as filler；Using acetonitrile as stream Dynamic phase A, using water as Mobile phase B, according to the form below carries out gradient elution；Detection wavelength is 203nm, and number of theoretical plate presses Panax Notoginseng saponin R₁Peak 4000 should be not less than by calculating；

(b) the preparation precision of reference substance solution weighs ginsenoside Rg₁Reference substance, ginsenoside Rb₁Reference substance and pseudo-ginseng soap Glycosides R₁Appropriate reference substance, adds methanol that every 1ml is made and contains ginsenoside Rg₁0.4mg, ginsenoside Rb₁0.4mg, notoginsenoside R₁The mixed solution of 0.1mg, to obtain the final product；

(c) preparation of test solution takes this product, finely ground, takes 0.7g, accurately weighed, puts in apparatus,Soxhlet's, adds diethyl ether 100ml, when low-temperature heat refluxing extraction 3 is small, discards ether solution, the dregs of a decoction volatilize solvent, then add methanol 100ml, continue reflux and carry Take 4 it is small when, let cool, collect extracting solution, merged with methanol 10ml gradation washing container and the dregs of a decoction, washing lotion with extracting solution, be evaporated, it is residual Slag adds methanol to dissolve, and is transferred in 10ml measuring bottles, adds methanol to shake up to scale, filter, take subsequent filtrate, to obtain the final product；

(d) determination method is accurate respectively draws reference substance solution and each 10 μ l of test solution, injects liquid chromatograph, surveys It is fixed, containing ginsenoside Rg₁(C₄₂H₇₂O₁₄), ginsenoside Rb₁(C₅₄H₉₂O₂₃), Panax Notoginseng saponin R₁(C₄₇H₈₀O₁₈) total amount must not lack In 4.8mg；

(4) cinnabar assay, is measured using precipitation titration, and step is as follows：This product is taken, it is finely ground, 5g is taken, precision claims It is fixed, put in 250ml kjeldahl flasks, add 20~40ml of sulfuric acid and 7~9g of potassium nitrate, be heated to closely colourless, let cool, be transferred to 250ml In conical flask, with water 50ml washing flasks by several times, washing lotion is incorporated in solution, adds 1% liquor potassic permanganate to aobvious pink, two points Do not disappear in clock, then be added dropwise 2% ferrous sulfate solution to red disappear after, add ammonium ferric sulfate indicator solution 2ml, dripped with ammonium thiocyanate Determine liquid (0.1mol/L) titration, per mercuric sulphide HgS of the 1ml ammonium thiocyanates titrating solution equivalent to 11.63mg, this product every contains Zhu Sand is 9.5~13.5mg in terms of mercuric sulphide HgS.

Angelica sinensis indentification by TLC of the present invention comprises the following steps that：

The preparation of test solution：This product content 1.75g is taken, add diethyl ether 20ml, shakes 10 minutes, filtration, and filter residue is standby With；Filtrate is concentrated into 5ml, as test solution；

The preparation of control medicinal material solution：Angelica sinensis control medicinal material 0.1g is taken, comparison medicine is made by test solution preparation method Material solution；

The preparation of negative sample solution：The prescription medicinal material of scarce Angelica sinensis is taken, it is molten by test sample in prescription ratio and preparation process Negative controls are made in liquid and preparation method thereof；

TLC test：Draw above-mentioned each 6 μ l of three kinds of solution, put respectively on same silica gel g thin-layer plate, with just oneself Alkane：Ethyl acetate=5: 1 is solvent, are unfolded, and take out, dry, put and inspected under 365nm ultraviolet lamps；

As a result：In test sample chromatography, on position corresponding with control medicinal material chromatography, the fluorescence spot of same color is shown, And negative sample is noiseless.

Borneol indentification by TLC of the present invention comprises the following steps that：

The preparation of test solution：Angelica sinensis thin layer is taken to differentiate the test solution under item, as test solution；

The preparation of reference substance solution：Borneol reference substance 0.1g is taken, the 5ml that adds diethyl ether makes dissolving, as reference substance solution；

The preparation of negative sample solution：The prescription medicinal material of scarce borneol is taken, it is molten by test sample in prescription ratio and preparation process Negative controls are made in liquid and preparation method thereof；

TLC test：Above-mentioned each 3 μ l of three kinds of solution are drawn, are put respectively on same silica gel g thin-layer plate, with 60~ 90 DEG C of petroleum ethers：Ethyl acetate=17: 3, it is solvent, is unfolded, takes out, dry, spray with 5% vanillin-sulfuric acid solution, 105 It is clear DEG C to be heated to spot development；

As a result：In test sample chromatography, on position corresponding with reference substance chromatography, the spot of same color is shown, and it is negative Sample is noiseless.

Cape jasmine indentification by TLC of the present invention comprises the following steps that：

The preparation of test solution：Take Angelica sinensis to differentiate the dregs of a decoction under item, wave most ether, add methanol 20ml, be ultrasonically treated 30 Minute, filtration, filtrate is concentrated into 2ml, as test solution；

The preparation of reference substance solution：Gardenoside reference substance is taken, adds methanol that solution of every 1ml containing 2mg is made, as reference substance Solution；

The preparation of negative sample solution：The prescription medicinal material of scarce cape jasmine is taken, it is molten by test sample in prescription ratio and preparation process Negative controls are made in liquid and preparation method thereof；

TLC test：Each 4 μ l of above two solution are drawn, are put respectively on same silica gel g thin-layer plate, with three chloromethanes Alkane：Methanol=3: 1, it is solvent, is unfolded, takes out, dry, spray with 5% ethanol solution of sulfuric acid, being heated to spot at 105 DEG C shows Color is clear；

Pseudo-ginseng indentification by TLC of the present invention comprises the following steps that：

The preparation of test solution：This product is taken, it is finely ground, 0.7g is taken, it is accurately weighed, put in apparatus,Soxhlet's, add diethyl ether 100ml, when low-temperature heat refluxing extraction 3 is small, discards ether solution, the dregs of a decoction volatilize solvent, then add methanol 100ml, continue reflux and carry Take 4 it is small when, let cool, collect extracting solution, merged with methanol 10ml gradation washing container and the dregs of a decoction, washing lotion with extracting solution, be evaporated, it is residual Slag adds methanol to dissolve, and is transferred in 10ml measuring bottles, adds methanol to shake up to scale, filter, take subsequent filtrate, to obtain the final product；

The preparation of control medicinal material solution：Pseudo-ginseng control medicinal material 0.25g separately is taken, adds methanol 20ml, is ultrasonically treated 30 minutes, filter Cross, filtrate is evaporated, and residue adds methanol 5ml to make dissolving, as control medicinal material solution；

The preparation of negative sample solution：The prescription medicinal material of scarce pseudo-ginseng is taken, it is molten by test sample in prescription ratio and preparation process Negative controls are made in liquid and preparation method thereof；

TLC test：Draw above-mentioned test solution and control medicinal material solution and each 10 μ l of cloudy sample solution, compare 5 μ l of product solution, put on same silica gel g thin-layer plate, with chloroform respectively：Methanol：Water=65: 35:10,10 DEG C with decentralization Put 12 it is small when more than lower floor's solution be solvent, be unfolded, take out, dry, spray with 5% ethanol solution of sulfuric acid, 105 DEG C heating It is clear to spot development；

Vinegar corydalis tuber indentification by TLC of the present invention comprises the following steps that：

The preparation of test solution：This product 35.0g is taken, is put in conical flask with cover, add diethyl ether 100ml, shakes 10 minutes, adds Ammonia solution 10ml, shakes 30 minutes, place 4 it is small when, filtration, filtrate put in separatory funnel, and the dregs of a decoction wash 2 times with ether, every time 30ml, washing lotion are incorporated in separatory funnel, are extracted 4 times, each 30ml, 30ml, 20ml with hydrochloric acid solution (2 → 100) shaking, 20ml, merges acid solution, adjusts pH value to 10~11 with ammonium hydroxide, with ether shaking extraction 3 times, each 30ml, 30ml, 20ml, closes And ether solution, it is evaporated, residue adds absolute ethyl alcohol 0.5ml to make dissolving, as test solution；

The preparation of control medicinal material solution：Corydalis tuber control medicinal material 2g is taken, adds appropriate amount of water, when decoction 2 is small, filtration, filtrate is used Ammonium hydroxide adjusts pH value to 10~11, is shaken and extracted with ether 40ml, divided and take ether layer, volatilize, it is molten that residue adds absolute ethyl alcohol 1ml to make Solution, as control medicinal material solution；

The preparation of negative sample solution：The prescription medicinal material of scarce vinegar corydalis tuber is taken, is pressed in prescription ratio and preparation process for trying Negative controls are made in product solution manufacturing method；

TLC test：Draw above-mentioned each 5 μ l of three kinds of solution, put respectively on same silica gel g thin-layer plate, with just oneself Alkane：Ethyl acetate=1: 1, be solvent, with ammonia steam presaturation 10 minutes, be unfolded, take out, dry, put smoked in iodine vapor to Spot development is clear, waves the iodine of most absorption, puts and is inspected under ultraviolet lamp (365nm).

Aconitine limit detection of the present invention comprises the following steps that：

The preparation of test solution：This product 35.0g is taken, is put in conical flask with cover, add diethyl ether 100ml, shakes 10 minutes, adds Ammonia solution 10ml, shakes 30 minutes, place 4 it is small when, filtration, filtrate put in separatory funnel, and the dregs of a decoction wash 2 times with ether, every time 30ml, washing lotion are incorporated in separatory funnel, and extraction 4 times (30ml, 30ml, 20ml, 20ml) is shaken with hydrochloric acid solution (2 → 100), Merge acid solution, adjust pH value to 10~11 with ammonium hydroxide, extract 3 times (30ml, 30ml, 20ml) with ether shaking and merge ether solution, It is evaporated, residue adds absolute ethyl alcohol 0.5ml to make dissolving, as test solution；

The preparation of reference substance solution：Separately take aconitine reference substance appropriate, it is weighed, add absolute ethyl alcohol that every 1ml is made and contain 2.0mg Solution, as reference substance solution；

The preparation of negative sample solution：The prescription medicinal material of scarce aconiti preparata,radix, wild aconite root is taken, is pressed in prescription ratio and preparation process Negative controls are made in test solution preparation method；

TLC test：Each 12ul of above-mentioned test solution and negative controls solution, reference substance solution 5ul are drawn, Put respectively on same silica gel g thin-layer plate, with n-hexane：Ethyl acetate=1:1 is solvent, puts the expansion cylinder of ammonia saturated with vapor In, it is unfolded, takes out, dry, with same solvent second outspread, takes out, dry, spray with potassium iodide iodine test solution：Bismuth potassium iodide tries Liquid=1:1 mixed solution.

As a result：In test sample chromatography, the spot occurred on position corresponding with reference substance chromatography should be less than reference substance Spot occurs without spot.

Combination dosage form of the present invention includes：Powder, granule, tablet and hard capsule.

The method of inspection of the present invention using TLC technique to Angelica sinensis, borneol, cape jasmine, pseudo-ginseng, corydalis tuber in primary standard Verified；The thin-layer qualitative detection method of frankincense, chrysanthemum is created at the same time；Three have been reconstructed using high-efficient liquid phase chromatogram technology Seven quantitative detecting methods, multi-objective control is transformed into by single norm controlling, while establishes the quantitative detecting method of cinnabar, is carried High clinical safety and validity.This method is applicable not only to overall quality control of the industrialized production to this kind of composition, Also safe and effective guarantee is provided for clinical application.

The beneficial effects of the invention are as follows：

(1) thin layer of newly-increased medicinal material frankincense of the present invention differentiates, its feature is：Establish with silica G F₂₅₄Lamellae For carrier, with petroleum ether (60~90 DEG C)：Thiacyclohexane：Ethyl acetate：Formic acid=10:30:15:1 is solvent, in ultraviolet lamp The chromatography of fluorescence spot is seen under the conditions of 254nm；Change using silica gel g thin-layer plate as carrier in routine, with ring ring or normal hexane： Ethyl is solvent, the chromatographic condition to be developed the color in the sunlight with 5% vanillin-sulfuric acid solution.

(2) thin layer of newly-increased medicinal material chrysanthemum of the present invention differentiates, its feature is：Establish using hydrochloric acid hydrolysate to grind Object is studied carefully, using polyamide film as carrier, with butyl acetate：Glacial acetic acid：Water=7:2.5:The upper solution of (2.5~3.0) is Solvent, sees fluorescence dark spots under the conditions of ultraviolet lamp 365nm；Change using principal component chlorogenic acid in chrysanthemum as research pair The normal experiment of elephant, conventional method have negative interference in this preparation.

(3) content assaying method of newly-increased medicinal material cinnabar of the present invention, its feature are：Energy in titration experiments is determined The amount of the enough concentrated sulfuric acid potassium nitrate reacted completely, mercuric sulphide in cinnabar is oxidized to mercury ion and sulfuric acid by the two in a heated condition Radical ion, the content of mercuric sulphide is calculated finally by the ammonium thiocyanate of consumption.

(4) content assaying method of medicinal material pseudo-ginseng of the present invention, its feature are：Using high performance liquid chromatography, sample Product processing is cleaned using Soxhlet ether, the processing mode of methanol effective component extracting, and mobile phase then utilizes the mode of gradient elution To its active ingredient ginsenoside Rg₁, ginsenoside Rb₁, Panax Notoginseng saponin R₁Three kinds of index components carry out separation in fact for index Test, pass through condition selection and methodology validation, it was demonstrated that its method has feasibility；Routine is changed by detection of single component to refer to Target control method.

Detection method of the present invention is simple and easy to do, is suitable for detection of the industrialized production to its quality, detection side Method specificity is strong, favorable reproducibility, and through methodology validation, its accuracy is high, while to ensure that industrialized production quality provides one Kind science, accurate detection method.

Brief description of the drawings

Fig. 1 is that Angelica sinensis differentiates thin-layer chromatogram, and 1,3,5 be test sample in figure, and 2,4 be Angelica sinensis control medicinal material, and 6 feminine genders are right According to；

Fig. 2 is that borneol differentiates thin-layer chromatogram, and 1,3,5 be test sample in figure, and 2,4 be borneol control medicinal material, and 6 feminine genders are right According to；

Fig. 3 is that cape jasmine differentiates thin-layer chromatogram, and 1,3,5 be test sample in figure, 2,4 jasminoidin reference substances, 6 negative controls；

Fig. 4 is that pseudo-ginseng differentiates thin-layer chromatogram, and 1,3,5 be test sample, 2, pseudo-ginseng control medicinal material, 4, ginsenoside Rg₁、 Rb₁Panax Notoginseng saponin R₁Reference substance, 6 negative controls；

Fig. 5 is that corydalis tuber differentiates thin-layer chromatogram, and 1,3,5 be test sample, 2,4 corydalis tuber control medicinal materials, 6 negative controls；

Fig. 6 is that frankincense differentiates thin-layer chromatogram, and 1,3,5 be test sample, 2,4 frankincense control medicinal materials, 6 negative controls；

Fig. 7 is that chrysanthemum differentiates thin-layer chromatogram, and 1,3,5 be test sample, 2,4 chrysanthemum control medicinal materials, 6 negative controls；

Fig. 8 is that aconitine limit checks thin-layer chromatogram, and 1,3,5 be test sample, and 2,4 be aconitine reference substance, and 6 is negative Control；

Fig. 9 is ginsenoside Rg₁Ultraviolet maximum absorption spectrum figure；

Figure 10 is ginsenoside Rb₁Ultraviolet maximum absorption spectrum figure；

Figure 11 is Panax Notoginseng saponin R₁Ultraviolet maximum absorption spectrum figure；

Figure 12 is Panax Notoginseng saponin R₁Content standard curve map；

Figure 13 is ginsenoside Rg₁Content standard curve map；

Figure 14 is ginsenoside Rb₁Content standard curve map；

Figure 15 is ginsenoside Rg₁, ginsenoside Rb₁, Panax Notoginseng saponin R₁Blank solution chromatogram；

Figure 16 is ginsenoside Rg₁, ginsenoside Rb₁, Panax Notoginseng saponin R₁Reference substance chromatogram；

Figure 17 is ginsenoside Rg₁, ginsenoside Rb₁, Panax Notoginseng saponin R₁Content sample chromatogram figure；

Figure 18 is ginsenoside Rg₁, ginsenoside Rb₁, Panax Notoginseng saponin R₁Content negative control chromatogram.

Embodiment

Embodiment 1

The preparation method and detection method of Changchun Hongyao capsule

The capsule of Chinese medicine composition of the present invention is made of the Chinese medicine of following parts by weight

121.2 parts of pseudo-ginseng, 12.12 parts of wild aconite root, 12.12 parts of aconiti preparata,radix, 12.12 parts of lotus nut, 20.2 parts of Angelica sinensis, scald 40.4 parts of the rhizome of davallia, 8.1 parts of grass-leaved sweetflag, 60.6 parts of dandelion, 6.7 parts of field thistle, 20.2 parts of stir-baked OLIBANUM with vinegar, 20.2 parts of stir-baked MYRRHA with vinegar, celestial being 60.6 parts of crane grass, 24.2 parts of borneol, 40.4 parts of safflower, 40.4 parts of chrysanthemum, 40.4 parts of cape jasmine, 13.5 parts of Paris polyphylla, cinnabar 12.1 Part, 20.2 parts of vinegar corydalis tuber

Capsule preparation method thereof

19 taste of the above, cinnabar fine grinding is into impalpable powder；Borneol and grinding is into fine powder；Pseudo-ginseng, aconiti preparata,radix, Angelica sinensis, frankincense, do not have Medicine, grass-leaved sweetflag are ground into fine powder, with Cinnabar facing-up, sieve, mix；Medicinal material adds water to cook two to remaining cape jasmine etc. ten simply It is secondary, every time 2 it is small when, by several times filter, merging filtrate, be condensed into relative density be 1.23~1.25 (70~75 DEG C) thick paste.With Above-mentioned powder (removing borneol) mixing, dry, pulverize, and addition starch is appropriate, mixes, and adds borneol, mixes, and loads capsule, is made 1000, to obtain the final product.

The detection method of capsule is as follows：

Angelica sinensis indentification by TLC comprises the following steps that：

As a result：In test sample chromatography, on position corresponding with control medicinal material chromatography, the fluorescence spot of same color is shown, And negative sample is noiseless, illustrate that this method possesses the feature of specificity.See attached drawing 1.

Borneol indentification by TLC comprises the following steps that：

As a result：In test sample chromatography, on position corresponding with reference substance chromatography, the spot of same color is shown, and it is negative Sample is noiseless, illustrates that this method possesses the feature of specificity.See attached drawing 2.

Cape jasmine indentification by TLC comprises the following steps that：

As a result：In test sample chromatography, on position corresponding with reference substance chromatography, the spot of same color is shown, and it is negative Sample is noiseless, illustrates that this method possesses the feature of specificity.See attached drawing 3.

Pseudo-ginseng indentification by TLC comprises the following steps that：

As a result：In test sample chromatography, on position corresponding with reference substance chromatography, the spot of same color is shown, and it is negative Sample is noiseless, illustrates that this method possesses the feature of specificity.See attached drawing 4.

Vinegar corydalis tuber indentification by TLC comprises the following steps that：

TLC test：Draw above-mentioned each 5 μ l of three kinds of solution, put respectively on same silica gel g thin-layer plate, with just oneself Alkane：Ethyl acetate=1: 1, be solvent, with ammonia steam presaturation 10 minutes, be unfolded, take out, dry, put smoked in iodine vapor to Spot development is clear, waves the iodine of most absorption, puts and is inspected under 365nm ultraviolet lamps.

As a result：In test sample chromatography, on position corresponding with control medicinal material chromatography, the fluorescence spot of same color is shown, And negative sample is noiseless, illustrate that this method possesses the feature of specificity.See attached drawing 5.

Stir-baked OLIBANUM with vinegar indentification by TLC comprises the following steps that：

The preparation of test solution：This product content 2.5g is taken, it is finely ground, add ethanol 20ml, be ultrasonically treated 20 minutes, filter Cross, filtrate volatilizes, and residue adds ethanol 1ml, as test solution；

The preparation of control medicinal material solution：Frankincense control medicinal material 0.2g is taken, comparison medicine is made by test solution preparation method Material solution；

The preparation of negative sample solution：The prescription medicinal material of scarce stir-baked OLIBANUM with vinegar is taken, test sample is pressed in prescription ratio and preparation process Negative controls are made in solution manufacturing method；

TLC test：Above-mentioned each 5 μ l of three kinds of solution are drawn, are put respectively in same silica G F₂₅₄On lamellae, with 60 DEG C petroleum ether：Thiacyclohexane：Ethyl acetate：Formic acid=10:30:15:1 is solvent, is unfolded, and takes out, dries, it is ultraviolet to put 254nm Inspected under light lamp；

As a result：In test sample chromatography, on position corresponding with control medicinal material chromatography, the spot and feminine gender of same color are shown Sample is noiseless, illustrates that this method possesses the feature of specificity.See attached drawing 6.

Chrysanthemum indentification by TLC comprises the following steps that：

The preparation of test solution：This product content 6g is taken, it is finely ground, add chloroform 30ml, be heated to reflux 30 minutes, filter Cross, discard filtrate, the dregs of a decoction volatilize solvent, add water 80ml, be heated to reflux 1 it is small when, centrifugation, take supernatant, add hydrochloric acid 0.5ml, use Ethyl acetate shaking extraction is secondary, and each 40ml, combined ethyl acetate liquid, is evaporated, and residue adds methanol 1ml to make solution, as confession Test sample solution；

The preparation of control medicinal material solution：Chrysanthemum control medicinal material 0.5g separately is taken, control is made by test solution preparation method Medicinal material solution；

The preparation of negative sample solution：The prescription medicinal material of scarce chrysanthemum is taken, it is molten by test sample in prescription ratio and preparation process Negative controls are made in liquid and preparation method thereof；

TLC test：Above-mentioned each 2 μ l of three kinds of solution are drawn, puts on same polyamide film, makes into strips respectively, With butyl acetate：Glacial acetic acid：Water=7:2.5:2.5 upper solution is solvent, is unfolded, and takes out, dries, it is ultraviolet to put 365nm Inspected under light lamp；

As a result：In test sample chromatography, on position corresponding with control medicinal material chromatography, aobvious identical fluorescence streak and feminine gender Sample is noiseless, illustrates that this method possesses the feature of specificity.See attached drawing 7.

Toxic component aconitine limit inspection comprises the following steps that：

The preparation of test solution：This product content 35.0g is taken, is put in conical flask with cover, add diethyl ether 100ml, shaking 10 Minute, test solution 10ml is ammoniated, is shaken 30 minutes, when placement 4 is small, filtration, filtrate is put in separatory funnel, and the dregs of a decoction are washed 2 times with ether, Each 30ml, washing lotion are incorporated in separatory funnel, with hydrochloric acid solution (2 → 100) shaking extraction 4 times (30ml, 30ml, 20ml, 20ml), merge acid solution, adjust pH value to 10~11 with ammonium hydroxide, extract 3 times (30ml, 30ml, 20ml) with ether shaking and merge Ether solution, is evaporated, and residue adds absolute ethyl alcohol 0.5ml to make dissolving, as test solution；

As a result：In test sample chromatography, the spot occurred on position corresponding with reference substance chromatography should be less than reference substance Spot occurs without spot, illustrates that this method possesses the feature of specificity.See attached drawing 8.

Pseudo-ginseng assay is comprised the following steps that using high effective liquid chromatography for measuring：

Chromatographic condition is with system suitability using octadecylsilane chemically bonded silica as filler；Using acetonitrile as mobile phase A, using water as Mobile phase B, the regulation according to the form below carries out gradient elution；Detection wavelength is 203nm, and number of theoretical plate presses notoginsenoside R₁Peak, which calculates, should be not less than 4000；

The preparation of reference substance solution：Precision weighs ginsenoside Rg₁Reference substance, ginsenoside Rb₁Reference substance and notoginsenoside R₁Appropriate reference substance, adds methanol that every 1ml is made and contains ginsenoside Rg₁0.4mg, ginsenoside Rb₁0.4mg, notoginsenoside R₁The mixed solution of 0.1mg, to obtain the final product；

The preparation of test solution：This product content under content uniformity item is taken, it is finely ground, 0.7g is taken, it is accurately weighed, put rope In family name's extractor, add diethyl ether 100ml, when low-temperature heat refluxing extraction 3 is small, discards ether solution, the dregs of a decoction volatilize solvent, then add methanol 100ml, when continuation refluxing extraction 4 is small, lets cool, and collects extracting solution, with methanol 10ml gradation washing container and the dregs of a decoction, washing lotion is with carrying Take liquid to merge, be evaporated, residue adds methanol to dissolve, and is transferred in 10ml measuring bottles, adds methanol to shake up to scale, filter, take continuous filter Liquid, to obtain the final product；

Accurate absorption reference substance solution and each 10 μ l of test solution respectively, injection liquid chromatograph, measure, to obtain the final product,

This product every contains ginsenoside Rg₁(C₄₂H₇₂O₁₄), ginsenoside Rb₁(C₅₄H₉₂O₂₃), Panax Notoginseng saponin R₁ (C₄₇H₈₀O₁₈) total amount must not be less than 4.8mg.

The methodological study of pseudo-ginseng detection method of content

Instrument：1260 high performance liquid chromatographs of Agilent (G13llC quaternarys gradient pump, G1329B autosamplers, G1316A column ovens, G13l4F Variable wavelength UV detectors, G2170B chem workstations), Sai Duolisi BP211D electronics day It is flat.

Acetonitrile (chromatographically pure), methanol (chromatographically pure), water (purified water), other reagents are that analysis is pure.

Ginsenoside Rg₁Lot number：110703-201529, ginsenoside Rb₁Lot number：110704-201424, notoginsenoside R₁Lot number：110745-201318, derives from National Institute for Food and Drugs Control.

The selection of measure wavelength takes ginsenoside Rg₁Reference substance, ginsenoside Rb₁Reference substance and Panax Notoginseng saponin R₁Control Appropriate product, add methanol that ginsenoside Rg is respectively prepared₁Reference substance solution, ginsenoside Rb₁Reference substance solution and Panax Notoginseng saponin R₁ Reference substance solution, using solvent as blank, is scanned, it is 203nm to determine this product Detection wavelength at 200~400nm wavelength.Attached drawing 9-11。

The investigation of linear relationship takes ginsenoside Rg respectively₁, ginsenoside Rb₁And Panax Notoginseng saponin R₁Appropriate reference substance, adds first Alcohol is configured to contain ginsenoside Rg₁1200ug/ml, ginsenoside Rb₁1200ug/ml and Panax Notoginseng saponin R₁The control of 240ug/ml Product solution, precision draw this solution 2ml, 6ml, 10ml, 12ml, 14ml, 18ml, 20ml respectively to 20ml volumetric flasks, add methanol It is diluted to scale.It is accurate again to draw 10ul, liquid chromatograph is injected separately into, records chromatogram, is sat using integrating peak areas value to be vertical Mark, with ginsenoside Rg₁, ginsenoside Rb₁And Panax Notoginseng saponin R₁Sample size (μ g) draws standard curve for abscissa.The result is shown in Table 1,2,3, attached drawing 12-14.

1 Panax Notoginseng saponin R of table₁Standard curve determination result

2 ginsenoside Rg of table₁Standard curve determination result

3 ginsenoside Rb of table₁Standard curve determination result

The result shows that Panax Notoginseng saponin R₁Concentration is in the range of 24~240 μ g/ml；Ginsenoside Rg₁Concentration 120~ In the range of 1200 μ g/ml；Ginsenoside Rb₁Concentration is in the range of 120~1200 μ g/ml, integrating peak areas value and notoginsenoside R₁, ginsenoside Rg₁, ginsenoside Rb₁Sample size is in good linear relationship, and regression equation is respectively：Panax Notoginseng saponin R₁Y= 507.88x-5.0117 correlation coefficient r=0.99993；Ginsenoside Rg₁Y=208.38x+4.4034, correlation coefficient r= 0.99998；Ginsenoside Rb₁Y=230.35x-5.9035, correlation coefficient r=0.99997.

The measure of precision is accurate to draw same each 10 μ l of test liquid, replication 6 times, each index components integrating peak areas The RSD values difference of value：Panax Notoginseng saponin R₁RSD values are 0.61%, ginsenoside Rg₁RSD values are 0.42%, ginsenoside Rb₁RSD It is worth for 0.28%.It the results are shown in Table 4.

4 precision measurement result of table

The result shows that：Panax Notoginseng saponin R₁, ginsenoside Rg₁With ginsenoside Rb₁Precision it is good.

Reappearance test selects same batch of sample, prepares 6 parts of test solutions, each index components content RSD values point respectively Not：Panax Notoginseng saponin R₁RSD values are 1.79%, ginsenoside Rg₁RSD values are 0.84%, ginsenoside Rb₁RSD values are 1.44%. Ginsenoside Rg₁, ginsenoside Rb₁With Panax Notoginseng saponin R g₁The RSD values of total amount are 1.13%.It the results are shown in Table 5.

5 reappearance measurement result of table

Test result indicates that：The reappearance of this product is good.

Stability test takes same test solution, with 0,2,4,6,8,12,24 for time interval sample introduction, each sample introduction 10 μ l, record integrating peak areas value, calculate each index components content RSD values difference：Panax Notoginseng saponin R₁RSD values be 1.72%, people Join saponin(e Rg₁RSD values for 0.72%, ginsenoside Rb₁RSD values be 0.78%.It the results are shown in Table 6.

6 stability of solution measurement result of table

The result shows that this assay method test solution internal stability when 24 is small is good.

Recovery test takes this product content appropriate, finely ground, weighs 6 parts, every part of about 0.7g, accurately weighed, puts 6 respectively In filtration paper cylinder, then ginsenoside Rg is weighed respectively₁Reference substance about 6.6mg, ginsenoside Rb₁Reference substance about 4.1mg and pseudo-ginseng soap Glycosides R₁Reference substance about 1.0mg, it is accurately weighed, put in above-mentioned filtration paper cylinder and mixed with sample respectively, then the processing method by test sample Handled with method, calculate the rate of recovery and RSD values.The rate of recovery=[measured amount (mg)-formulation content (mg)]/addition reference substance Measure (mg) × 100%.It the results are shown in Table 7,8,9.

7 Panax Notoginseng saponin R of table₁Recovery test result

8 ginsenoside Rg of table₁Recovery test result

9 ginsenoside Rb of table₁Recovery test result

The result shows that：This method sample recovery rate is good.

Control experiment takes the negative sample solution prepared in prescription ratio and preparation process with method, reference substance solution, for examination Product solution, solvent used each 10 μ l of sample introduction under identical experiment condition.Test result indicates that：The negative control of this product with Ginsenoside Rg₁Reference substance, ginsenoside Rb₁Reference substance, Panax Notoginseng saponin R₁Without miscellaneous on the corresponding position of reference substance solution chromatographic peak Mass peak occurs, and illustrates that the blank assay of this product is noiseless, this method has specificity.The result is shown in Figure 15, Figure 16, Figure 17, Figure 18.

Cinnabar assay uses complexometric titration, comprises the following steps that：

This product under weight differential item is taken, it is finely ground, 5g is taken, it is accurately weighed, put in 250ml kjeldahl flasks, add sulfuric acid 30ml With potassium nitrate 8g, it is heated near colourless, lets cool, be transferred in 250ml conical flasks, washing flask, washing lotion are incorporated to by several times with water 50ml In solution, 1% liquor potassic permanganate is added not disappear in two minutes, then 2% ferrous sulfate solution is added dropwise to red to aobvious pink After disappearance, add ammonium ferric sulfate indicator solution 2ml, titrated with ammonium thiocyanate titrating solution (0.1mol/L).Per 1ml ammonium thiocyanate titrating solutions The mercuric sulphide (HgS) of (0.1mol/L) equivalent to 11.63mg.

This product every in terms of mercuric sulphide (HgS), should be 9.5~13.5mg containing cinnabar.

The methodological study of cinnabar detection method of content

Instrument：The triumphant formula flasks of 250ml, 10ml browns acid buret, 50ml graduated cylinders, Electric stove, Sai Duolisi balances BP211D

Reagent reagent：1% liquor potassic permanganate, 2% ferrous sulfate solution, 0.1mol/L ammonium thiocyanates titrating solution, sulfuric acid Iron ammonium indicator solution, potassium nitrate (analysis is pure), sulfuric acid (analysis is pure).

Reappearance test selects same batch of sample, and precision weighs 6 parts of sample, is carried out respectively according to cinnabar content assaying method Vulcanize sclera remodeling, RSD=1.44%, measurement result is shown in Table 10.

10 reappearance measurement result of table

Test result indicates that：The reappearance of this method is good.

Recovery test takes the sample of known content (33.14mg/g), finely ground, weighs 6 parts, every part of about 5g, accurately weighed, Weigh cinnabar fine powder (being 98.2% containing HgS) about 170mg of known content respectively again, it is accurately weighed, put 250mL kjeldahl flasks In, handled by cinnabar content assaying method with method, calculate the rate of recovery and RSD values.The rate of recovery=[measured amount (mg)-preparation Content (mg)]/add reference substance amount (mg) × 100%.It the results are shown in Table 11.

11 cinnabar recovery test result of table

Test result indicates that：This method sample recovery rate is good.

Control experiment takes the negative sample prepared in prescription ratio and preparation process with method, same by cinnabar content assaying method Method operates, the consumption of its titrating solution shows that quantitative determination of the negative sample to cinnabar has no significant effect, we in below 0.05mL Method has specificity.

The preparation method and detection method of 2 Changchun red tablet of embodiment

Tablet is made of the Chinese medicine of following parts by weight, with embodiment 1

Method for preparing tablet thereof

19 taste of the above, cinnabar fine grinding is into impalpable powder；Borneol and grinding is into fine powder；Pseudo-ginseng, aconiti preparata,radix, Angelica sinensis, frankincense, do not have Medicine, grass-leaved sweetflag are ground into fine powder, with Cinnabar facing-up, sieve, mix；Medicinal material adds water to cook two to remaining cape jasmine etc. ten simply It is secondary, every time 2 it is small when, by several times filter, merging filtrate, be condensed into relative density be 1.23~1.25 (70~75 DEG C) thick paste.With Above-mentioned powder (removing borneol) mixing, dry, pulverize, and addition starch is appropriate, mixes, and adds borneol, mixes, is pressed into 1000, To obtain the final product.

Angelica sinensis, borneol, cape jasmine, pseudo-ginseng and the indentification by TLC of vinegar corydalis tuber and the inspection method of aconitine limit, together Embodiment 1；Further include：

(a) preparation of test solution：This product 2.5g is taken, is mixed, adds ethanol 30ml, is ultrasonically treated 25 minutes, is filtered, filter Liquid volatilizes, and residue adds ethanol 1ml, as test solution；

(c) preparation of negative sample solution：The prescription medicinal material of vinegar hypogalactia perfume is taken, is pressed in prescription ratio and preparation method for trying Negative controls are made in product solution manufacturing method；

(d) chromatographic condition and operation：Test, draw above-mentioned according to thin-layered chromatography (Chinese Pharmacopoeia version general rule 0502 in 2015) Each 8 μ l of three kinds of solution, put in same silica G F respectively₂₅₄On lamellae, with 75 DEG C of petroleum ethers：Thiacyclohexane：Ethyl acetate：Formic acid =10:30:15:1 is solvent, is unfolded, and takes out, dries, put and inspected under 254nm ultraviolet lamps；

(a) preparation of test solution：This product 6g is taken, it is finely ground, add chloroform 40ml, be heated to reflux 30 minutes, filter, Filtrate is discarded, the dregs of a decoction volatilize solvent, add 80~100ml of water, are heated to reflux 50 minutes, and centrifugation, takes supernatant, add hydrochloric acid 0.8ml, Secondary with ethyl acetate shaking extraction, each 40ml, combined ethyl acetate liquid, is evaporated, and residue adds methanol 1ml to make solution, as Test solution；

(d) chromatographic condition and operation：Test, draw above-mentioned according to thin-layered chromatography (Chinese Pharmacopoeia version general rule 0502 in 2015) Each 4 μ l of three kinds of solution, put on same polyamide film, make into strips, with butyl acetate respectively：Glacial acetic acid：Water=:2.5: 2.7) upper solution is solvent, is unfolded, and takes out, dries, put and inspected under 365nm ultraviolet lamps；

(e) result feature:In test sample chromatography, on position corresponding with control medicinal material chromatography, identical phosphor strip is shown Spot.

(3) content assaying method of pseudo-ginseng, step are as follows：

(4) cinnabar assay, is measured using precipitation titration, and step is as follows：This product is taken, it is finely ground, 5g is taken, precision claims It is fixed, put in 250ml kjeldahl flasks, add sulfuric acid 20ml and potassium nitrate 7g, be heated to closely colourless, let cool, be transferred to 250ml conical flasks In, with water 50ml washing flasks by several times, washing lotion is incorporated in solution, adds 1% liquor potassic permanganate to aobvious pink, in two minutes not Disappear, then be added dropwise 2% ferrous sulfate solution to red disappear after, add ammonium ferric sulfate indicator solution 2ml, with ammonium thiocyanate titrating solution (0.1mol/L) is titrated, and per mercuric sulphide HgS of the 1ml ammonium thiocyanates titrating solution equivalent to 11.63mg, this product every is containing cinnabar with sulphur Change mercury HgS meters, be 9.5~13.5mg.

Embodiment 3

The preparation method and detection method that the red medicine in Changchun dissipates

Powder is made of the Chinese medicine of following parts by weight, with embodiment 1

Powder preparation method

19 taste of the above, cinnabar fine grinding is into impalpable powder；Borneol and grinding is into fine powder；Pseudo-ginseng, aconiti preparata,radix, Angelica sinensis, frankincense, do not have Medicine, grass-leaved sweetflag are ground into fine powder, with Cinnabar facing-up, sieve, mix；Medicinal material adds water to cook two to remaining cape jasmine etc. ten simply It is secondary, every time 2 it is small when, by several times filter, merging filtrate, be condensed into relative density be 1.23~1.25 (70~75 DEG C) thick paste.With Above-mentioned powder (removing borneol) mixing, dry, pulverize, and addition starch is appropriate, mixes, and adds borneol, mixes, packing, to obtain the final product.

Angelica sinensis, borneol, cape jasmine, pseudo-ginseng and the indentification by TLC of vinegar corydalis tuber and the inspection method of aconitine limit, together Embodiment 1, further includes：

(a) preparation of test solution：This product 2.5g is taken, is mixed, adds ethanol 40ml, is ultrasonically treated 30 minutes, is filtered, filter Liquid volatilizes, and residue adds ethanol 1ml, as test solution；

(d) chromatographic condition and operation：Test, draw above-mentioned according to thin-layered chromatography (Chinese Pharmacopoeia version general rule 0502 in 2015) Each 10 μ l of three kinds of solution, put in same silica G F respectively₂₅₄On lamellae, with 90 DEG C of petroleum ethers：Thiacyclohexane：Ethyl acetate：Formic acid =10:30:15:1 is solvent, is unfolded, and takes out, dries, put and inspected under 254nm ultraviolet lamps；

(a) preparation of test solution：This product 6g is taken, it is finely ground, add chloroform 50ml, be heated to reflux 30 minutes, filter, Filtrate is discarded, the dregs of a decoction volatilize solvent, add water 100ml, are heated to reflux 40 minutes, and centrifugation, takes supernatant, add hydrochloric acid 1.0ml, use second Acetoacetic ester shaking extraction is secondary, and each 40ml, combined ethyl acetate liquid, is evaporated, and residue adds methanol 1ml to make solution, as trying Product solution；

(d) chromatographic condition and operation：Test, draw above-mentioned according to thin-layered chromatography (Chinese Pharmacopoeia version general rule 0502 in 2015) Each 5 μ l of three kinds of solution, put on same polyamide film, make into strips, with butyl acetate respectively：Glacial acetic acid：Water=:2.5:3 Upper solution be solvent, be unfolded, take out, dry, put and inspected under 365nm ultraviolet lamps；

(e) result feature：In test sample chromatography, on position corresponding with control medicinal material chromatography, identical phosphor strip is shown Spot；Through methodology validation, its negative sample is noiseless, illustrates that this method possesses the feature of specificity.

(3) content assaying method of pseudo-ginseng, step are as follows：

(4) cinnabar assay, is measured using precipitation titration, and step is as follows：This product is taken, it is finely ground, 5g is taken, precision claims It is fixed, put in 250ml kjeldahl flasks, add sulfuric acid 40ml and potassium nitrate 9g, be heated to closely colourless, let cool, be transferred to 250ml conical flasks In, with water 50ml washing flasks by several times, washing lotion is incorporated in solution, adds 1% liquor potassic permanganate to aobvious pink, in two minutes not Disappear, then be added dropwise 2% ferrous sulfate solution to red disappear after, add ammonium ferric sulfate indicator solution 2ml, with ammonium thiocyanate titrating solution (0.1mol/L) is titrated, and per mercuric sulphide HgS of the 1ml ammonium thiocyanates titrating solution equivalent to 11.63mg, this product every is containing cinnabar with sulphur Change mercury HgS meters, be 9.5~13.5mg.

Embodiment 4

The preparation method and method of quality control of the red medicine granule in Changchun

Granule is made of the Chinese medicine of following parts by weight, with embodiment 1

The process for producing granula of composition

19 taste of the above, cinnabar fine grinding is into impalpable powder；Borneol and grinding is into fine powder；Pseudo-ginseng, aconiti preparata,radix, Angelica sinensis, frankincense, do not have Medicine, grass-leaved sweetflag are ground into fine powder, with Cinnabar facing-up, sieve, mix；Medicinal material adds water to cook two to remaining cape jasmine etc. ten simply It is secondary, every time 2 it is small when, by several times filter, merging filtrate, be condensed into relative density be 1.23~1.25 (70~75 DEG C) thick paste.With Above-mentioned powder is uniformly mixed, and particle is made, dry, whole grain, packing, to obtain the final product.

Granule detection method, with embodiment 1.

Claims

1. a kind of detection method for the Chinese medicine composition for treating traumatic injury, the Chinese medicine composition is obtained by the following steps 's：12.1 parts of fine grindings of cinnabar are into impalpable powder；24.2 parts of borneol is ground into fine powder；121.2 parts of pseudo-ginseng, 12.12 parts of aconiti preparata,radix, Angelica sinensis 20.2 parts, 20.2 parts of stir-baked OLIBANUM with vinegar, 20.2 parts of stir-baked MYRRHA with vinegar, 8.1 parts of grass-leaved sweetflag be ground into fine powder, with Cinnabar facing-up, sieve, mix It is even；40.4 parts of remaining cape jasmine, 12.12 parts of wild aconite root, 40.4 parts of Rhizoma drynariae preparata, 60.6 parts of dandelion, 6.7 parts of field thistle, hairyvein agrimony 60.6 parts, 40.4 parts of safflower, 40.4 parts of chrysanthemum, 12.12 parts of lotus nut, 13.5 parts of Paris polyphylla, the medicine simply of vinegar corydalis tuber 20.2 etc. ten Material add water to cook it is secondary, every time 2 it is small when, filter by several times, merging filtrate, surveyed when being condensed into 70~75 DEG C relative density for 1.23~ 1.25 thick pastes, mix with the above-mentioned powder in addition to borneol, dry, pulverize, and mix, and add borneol fine powder, mix；

Detection method includes：Angelica sinensis, borneol, cape jasmine, pseudo-ginseng and vinegar corydalis tuber indentification by TLC and aconitine limit inspection Checking method, it is characterised in that further include：

(a) preparation of test solution：This product 2.5g is taken, is mixed, adds 20~40ml of ethanol, is ultrasonically treated 20~30 minutes, filter Cross, filtrate volatilizes, and residue adds ethanol 1ml, as test solution；

(b) preparation of control medicinal material solution：Frankincense control medicinal material 0.2g is taken, control medicinal material is made by test solution preparation method Solution；

(c) preparation of negative sample solution：The prescription medicinal material of vinegar hypogalactia perfume is taken, it is molten by test sample in prescription ratio and preparation method Negative controls are made in liquid and preparation method thereof；

(d) chromatographic condition and operation：Tested according to thin-layered chromatography (Chinese Pharmacopoeia version general rule 0502 in 2015), draw above-mentioned three kinds Each 5~10 μ l of solution, put in same silica G F respectively₂₅₄On lamellae, with 60 DEG C~90 DEG C petroleum ethers：Thiacyclohexane：Acetic acid second Ester：Formic acid=10:30:15:1 is solvent, is unfolded, and takes out, dries, put and inspected under 254nm ultraviolet lamps；

(e) result feature：In test sample chromatography, on position corresponding with control medicinal material chromatography, the spot of same color is shown；Through Its negative sample of methodology validation is noiseless, illustrates that this method possesses the feature of specificity；

(a) preparation of test solution：This product 6g is taken, it is finely ground, add 30~50ml of chloroform, be heated to reflux 30 minutes, filter, Filtrate is discarded, the dregs of a decoction volatilize solvent, add 80~100ml of water, are heated to reflux 40~60 minutes, and centrifugation, takes supernatant, add hydrochloric acid 0.5~1.0ml, secondary with ethyl acetate shaking extraction, each 40ml, combined ethyl acetate liquid, is evaporated, and residue adds methanol 1ml Make solution, as test solution；

(b) preparation of control medicinal material solution：Chrysanthemum control medicinal material 0.5g separately is taken, comparison medicine is made by test solution preparation method Material solution；

(c) preparation of negative sample solution：The prescription medicinal material of scarce chrysanthemum is taken, test solution is pressed in prescription ratio and preparation process Negative controls are made in preparation method；

(d) chromatographic condition and operation：Tested according to thin-layered chromatography (Chinese Pharmacopoeia version general rule 0502 in 2015), draw above-mentioned three kinds Each 2~5 μ l of solution, put on same polyamide film, make into strips, with butyl acetate respectively：Glacial acetic acid：Water=:2.5: The upper solution of (2.5~3) is solvent, is unfolded, and takes out, dries, put and inspected under 365nm ultraviolet lamps；

(e) result feature：In test sample chromatography, on position corresponding with control medicinal material chromatography, identical fluorescence streak is shown；Through Its negative sample of methodology validation is noiseless, illustrates that this method possesses the feature of specificity.

(3) content assaying method of pseudo-ginseng, step are as follows：

(a) chromatographic condition and system suitability are using octadecylsilane chemically bonded silica as filler；Using acetonitrile as mobile phase A, using water as Mobile phase B, according to the form below carries out gradient elution；Detection wavelength is 203nm, and number of theoretical plate presses Panax Notoginseng saponin R₁Peak calculates 4000 should be not less than；

(b) the preparation precision of reference substance solution weighs ginsenoside Rg₁Reference substance, ginsenoside Rb₁Reference substance and Panax Notoginseng saponin R₁ Appropriate reference substance, adds methanol that every 1ml is made and contains ginsenoside Rg₁0.4mg, ginsenoside Rb₁0.4mg, Panax Notoginseng saponin R₁0.1mg Mixed solution, to obtain the final product；

(d) determination method is accurate respectively draws reference substance solution and each 10 μ l of test solution, injects liquid chromatograph, measure, contains Ginsenoside Rg₁C₄₂H₇₂O₁₄, ginsenoside Rb₁C₅₄H₉₂O₂₃, Panax Notoginseng saponin R₁C₄₇H₈₀O₁₈Total amount must not be less than 4.8mg；

(4) cinnabar assay, is measured using precipitation titration, and step is as follows：This product is taken, it is finely ground, 5g is taken, it is accurately weighed, put In 250ml kjeldahl flasks, add 20~40ml of sulfuric acid and 7~9g of potassium nitrate, be heated near colourless, let cool, be transferred to 250ml conical flasks In, with water 50ml washing flasks by several times, washing lotion is incorporated in solution, adds 1% liquor potassic permanganate to aobvious pink, in two minutes not Disappear, then be added dropwise 2% ferrous sulfate solution to red disappear after, add ammonium ferric sulfate indicator solution 2ml, with 0.1mol/L ammonium thiocyanates Titrating solution titrates, and per mercuric sulphide HgS of the 1ml ammonium thiocyanates titrating solution equivalent to 11.63mg, this product every is containing cinnabar with mercuric sulphide HgS is counted, and is 9.5~13.5mg.

A kind of 2. detection method for the Chinese medicine composition for treating traumatic injury according to claim 1, it is characterised in that Angelica sinensis Indentification by TLC comprises the following steps that：

The preparation of test solution：This product content 1.75g is taken, add diethyl ether 20ml, shakes 10 minutes, filtration, and filter residue is spare；Filter Liquid is concentrated into 5ml, as test solution；

The preparation of control medicinal material solution：Angelica sinensis control medicinal material 0.1g is taken, it is molten that control medicinal material is made by test solution preparation method Liquid；

The preparation of negative sample solution：The prescription medicinal material of scarce Angelica sinensis is taken, test solution system is pressed in prescription ratio and preparation process Negative controls are made in Preparation Method；

TLC test：Above-mentioned each 6 μ l of three kinds of solution are drawn, are put respectively on same silica gel g thin-layer plate, with n-hexane：Second Acetoacetic ester=5: 1 is solvent, are unfolded, and take out, dry, put and inspected under 365nm ultraviolet lamps；

As a result：In test sample chromatography, on position corresponding with control medicinal material chromatography, the fluorescence spot of same color is shown, and it is cloudy Property sample is noiseless.

A kind of 3. detection method for the Chinese medicine composition for treating traumatic injury according to claim 2, it is characterised in that borneol Indentification by TLC comprises the following steps that：

The preparation of negative sample solution：The prescription medicinal material of scarce borneol is taken, test solution system is pressed in prescription ratio and preparation process Negative controls are made in Preparation Method；

TLC test：Above-mentioned each 3 μ l of three kinds of solution are drawn, are put respectively on same silica gel g thin-layer plate, with 60~90 DEG C of stones Oily ether：Ethyl acetate=17: 3, it is solvent, is unfolded, take out, dry, spray with 5% vanillin-sulfuric acid solution, in 105 DEG C of heating It is clear to spot development；

As a result：In test sample chromatography, on position corresponding with reference substance chromatography, the spot of same color, and negative sample are shown It is noiseless.

A kind of 4. detection method for the Chinese medicine composition for treating traumatic injury according to claim 2, it is characterised in that cape jasmine Indentification by TLC comprises the following steps that：

The preparation of test solution：Take Angelica sinensis to differentiate the dregs of a decoction under item, wave most ether, add methanol 20ml, be ultrasonically treated 30 minutes, Filtration, filtrate is concentrated into 2ml, as test solution；

The preparation of reference substance solution：Gardenoside reference substance is taken, adds methanol that solution of every 1ml containing 2mg is made, it is molten as reference substance Liquid；

The preparation of negative sample solution：The prescription medicinal material of scarce cape jasmine is taken, test solution system is pressed in prescription ratio and preparation process Negative controls are made in Preparation Method；

TLC test：Each 4 μ l of above two solution are drawn, are put respectively on same silica gel g thin-layer plate, with chloroform： Methanol=3: 1, it is solvent, is unfolded, takes out, dry, spray with 5% ethanol solution of sulfuric acid, it is clear to be heated to spot development at 105 DEG C It is clear；

A kind of 5. detection method for the Chinese medicine composition for treating traumatic injury according to claim 1, it is characterised in that pseudo-ginseng Indentification by TLC comprises the following steps that：

The preparation of test solution：This product is taken, it is finely ground, 0.7g is taken, it is accurately weighed, put in apparatus,Soxhlet's, add diethyl ether 100ml, When low-temperature heat refluxing extraction 3 is small, ether solution is discarded, the dregs of a decoction volatilize solvent, then add methanol 100ml, and it is small to continue refluxing extraction 4 When, let cool, collect extracting solution, merged with methanol 10ml gradation washing container and the dregs of a decoction, washing lotion with extracting solution, be evaporated, residue adds Methanol dissolves, and is transferred in 10ml measuring bottles, adds methanol to shake up to scale, filter, take subsequent filtrate, to obtain the final product；

The preparation of control medicinal material solution：Pseudo-ginseng control medicinal material 0.25g separately is taken, adds methanol 20ml, is ultrasonically treated 30 minutes, filtration, Filtrate is evaporated, and residue adds methanol 5ml to make dissolving, as control medicinal material solution；

The preparation of negative sample solution：The prescription medicinal material of scarce pseudo-ginseng is taken, test solution system is pressed in prescription ratio and preparation process Negative controls are made in Preparation Method；

TLC test：Draw above-mentioned test solution and control medicinal material solution and each 10 μ l of cloudy sample solution, reference substance are molten 5 μ l of liquid, put on same silica gel g thin-layer plate, with chloroform respectively：Methanol：Water=65: 35:It is 10,10 DEG C arranged below 12 small When more than lower floor's solution be solvent, be unfolded, take out, dry, spray with 5% ethanol solution of sulfuric acid, spot is heated at 105 DEG C Colour developing is clear；

6. a kind of detection method for the Chinese medicine composition for treating traumatic injury according to claim 1, it is characterised in that vinegar prolongs Hu rope indentification by TLC comprises the following steps that：

The preparation of test solution：This product 35.0g is taken, is put in conical flask with cover, add diethyl ether 100ml, shakes 10 minutes, ammonification examination Liquid 10ml, shakes 30 minutes, and when placement 4 is small, filtration, filtrate is put in separatory funnel, and the dregs of a decoction are washed 2 times with ether, and each 30ml, is washed Liquid is incorporated in separatory funnel, with hydrochloric acid solution (2 → 100) shaking extraction 4 times, each 30ml, 30ml, 20ml, 20ml, merges acid Liquid, adjusts pH value to 10~11 with ammonium hydroxide, with ether shaking extraction 3 times, each 30ml, 30ml, 20ml, merges ether solution, steam Dry, residue adds absolute ethyl alcohol 0.5ml to make dissolving, as test solution；

The preparation of control medicinal material solution：Corydalis tuber control medicinal material 2g is taken, adds appropriate amount of water, when decoction 2 is small, filtration, filtrate ammonium hydroxide PH value is adjusted to 10~11, is shaken and extracted with ether 40ml, divided and take ether layer, volatilize, residue adds absolute ethyl alcohol 1ml to make dissolving, makees For control medicinal material solution；

The preparation of negative sample solution：The prescription medicinal material of scarce vinegar corydalis tuber is taken, it is molten by test sample in prescription ratio and preparation process Negative controls are made in liquid and preparation method thereof；

TLC test：Above-mentioned each 5 μ l of three kinds of solution are drawn, are put respectively on same silica gel g thin-layer plate, with n-hexane：Second Acetoacetic ester=1: 1, it is solvent, with ammonia steam presaturation 10 minutes, is unfolded, takes out, dry, puts to smoke to spot in iodine vapor and show Color is clear, waves the iodine of most absorption, puts and is inspected under 365nm ultraviolet lamps；

A kind of 7. detection method for the Chinese medicine composition for treating traumatic injury according to claim 1, it is characterised in that the rhizome of Chinese monkshood Alkali limit detection comprises the following steps that：

The preparation of reference substance solution：Separately take aconitine reference substance appropriate, it is weighed, add absolute ethyl alcohol that every 1ml is made containing the molten of 2.0mg Liquid, as reference substance solution；

The preparation of negative sample solution：The prescription medicinal material of scarce aconiti preparata,radix, wild aconite root is taken, is pressed in prescription ratio and preparation process for trying Negative controls are made in product solution manufacturing method；

TLC test：Each 12ul of above-mentioned test solution and negative controls solution, reference substance solution 5ul are drawn, respectively Point is on same silica gel g thin-layer plate, with n-hexane：Ethyl acetate=1:1 is solvent, in the expansion cylinder for putting ammonia saturated with vapor, Expansion, takes out, dries, and with same solvent second outspread, takes out, dries, spray with potassium iodide iodine test solution：Bismuth potassium iodide test solution= 1:1 mixed solution；

As a result：In test sample chromatography, the spot occurred on position corresponding with reference substance chromatography should be less than the spot of reference substance Or occur without spot.