CN102375004B - Method for evaluating clinical compatibility of injections - Google Patents

Abstract

The invention discloses a method for evaluating the clinical compatibility of injections. The method comprises the following steps of: preparing a sample solution; carrying out titration; generating a reaction activity spectrum; calculating corresponding thermodynamic parameters, such as enthalpy change Delta H, reaction constant K, entropy change Delta S and the like; judging whether the reaction type is an enthalpy drive reaction or an entropy drive reaction according to the relation between Delta H and the T Delta S, wherein T is the reaction temperature; and further judging the clinical compatibility of the injections. Compared with the prior art, the method disclosed by the invention has the advantages of strong universality, high sensitivity, simpleness and convenience for operation, less interference factors and good repeatability; the system can rapidly reach a basic line balance during online titration by adopting an instrument isothermal power compensating system, so as to adapt to continuous titration and increase the measurement accuracy; the reaction type can be judged by parameters without the need of knowing all ingredients in the solution, therefore the method is especially suitable for Chinese medicine injections of which the material bases are undefined; and the method disclosed by the invention can be used for effectively remedying the limitation of incapability of detection of the conventional chemical analysis method caused by no comparison products and difficulty in determining ingredient characteristic absorption spectrum or absorption wavelength and has better universality and reaction activity fingerprint spectrum characteristics.

Description

A kind of method of estimating injection clinical compatibility compatibility

Technical field

The present invention relates to a kind of method of estimating injection clinical compatibility compatibility, belong to clinical drug safety assessment technique field.

Background technology

Injection is extensive use of in clinical, for satisfying treatment needs such as underlying diseases and complication, often needs drug combination.But because the difference of different pharmaceutical character, drug combination may cause the change of original material base, drug effect to reduce, even bad reaction takes place.Now the method for detection of the drug combination security mainly contains visual inspection, particulate matter, pH value, osmotic pressure, high performance liquid chromatography (HPLC) etc.Use these methods that physicochemical property before and after the drug combination is detected, be worth to reference data for clinical drug use, but these method complicated operations, can not monitor by real-time online, can not take into account physics, chemical information simultaneously, and sensitivity is not high, universality is not strong, has therefore limited its application in clinical drug compatibility compatibility detects.So demand setting up a kind of method of taking into account physics, chemical feature information change simultaneously urgently, estimate injection clinical compatibility compatibility rapidly with real-time online, sensitivity.

Summary of the invention

The objective of the invention is in order to overcome the detection method of existing injection clinical compatibility compatibility, complicated operation, can not monitor by real-time online, can not take into account physics, chemical information simultaneously, and the limitation that sensitivity is not high, universality is not strong, (Isothermal Titration Calorimetry ITC) attempts applicating evaluating injection clinical compatibility compatibility, in the hope of setting up a kind of rapid sensitive, detection method that universality is strong to introduce the isothermal titration calorimetry in physical chemistry field.

ITC is a kind of micro-by the use of thermal means, is function with the variation of participation reactive material total concentration and the thermal change of reaction, in conjunction with certain mathematical model, obtains thermodynamic parameter.ITC can the on-line monitoring fast response, is mainly used in the interaction between big molecule and/or little branch at present, and environment is to research fields such as the influence of reaction and enzyme kineticss, but Shang Weijian is used for this method to estimate the relevant report of injection clinical compatibility compatibility.

For achieving the above object, technical scheme of the present invention is a kind of method of estimating injection clinical compatibility compatibility, and this method may further comprise the steps:

(1) the pattern medicine of preparation compatibility to be detected and the sample liquid of drug combination;

(2) sample liquid of (1) step preparation is carried out titration at the isothermal titration calorimeter;

(3) utilize the thermal change that produces in the isothermal titration calorimeter real time record drop reaction process, reaction of formation activity profile;

(4) basis (3) goes on foot the reactivity spectrum of the thermal change that obtains, analytical calculation thermodynamic parameter enthalpy change Δ H and reaction constant K;

(5) according to (4) thermodynamic parameter enthalpy change Δ H and the reaction constant K in step, calculate the Entropy Changes Δ S of reaction;

(6) according to (4), (5) thermodynamic parameter in step, judge reaction type, if | Δ H|＞T| Δ S| is that enthalpy drives reaction, illustrates that these two kinds of injections after fused chemical reaction have taken place, and is incompatible, can not drug combination, and wherein T is temperature; If | Δ H|＜T| Δ S|, for entropy drives reaction, illustrate that physical reactions only takes place the fused of these two kinds of injections, chemical compatibility is good.

In the step (1), if pattern medicine to be detected and drug combination are parenteral solutions, then the sample liquid of step (1) can directly be measured configuration from the parenteral solution of pattern medicine and drug combination; If pattern medicine to be detected and drug combination are freeze-dried powder, then get an amount of pattern medicine and drug combination respectively, be made into pattern drug solns and the drug combination solution of clinical working concentration.

Step (2) comprises following step at the titration operation that the isothermal titration calorimeter carries out:

(1) inject titrate in sample cell, vs can be pattern medicine sample liquid or drug combination sample liquid, or both exchanges;

(2) titrate constant temperature to 25 ℃, stir speed (S.S.) is 100～300rmin ^-1Under be stirred to the instrument self-poise after, the output power curve of register instrument is as baseline, be 30～60s writing time;

(3) with vs titrate is carried out titration, 20～50 of Continuous Titration, every 2～10 μ L, be 200～300s interval time;

(4) in the titration process, adopt Launch ITCRun software to carry out data acquisition.

The reactivity spectrum that generates according to the thermal change in the titration process in the step (3) is made up of course of reaction spectrum and thermal change spectrum, wherein the course of reaction spectrum is the power-time spectrum that obtains two kinds of parenteral solution titration processes by the thermal change that records drop reaction generation each time, and the thermal change spectrum is to carry out heat-titration number of times spectrum that integration obtains by the titration peak to the course of reaction spectrum.

Thermodynamic parameter enthalpy change Δ H in the step (4) and reaction constant K are according to the reactivity spectrum, calculate by following formula:

$Q=V\·\mathrm{\ΔH}\·C_M^b---\left(1\right)$

$\frac{Q}{n_L}={\mathrm{\Δ}}_r{H}_M^{\mathrm{\θ}}\·\frac{C_M^b}{C_L^0}---\left(2\right)$

$\frac{\left[\mathrm{ML}\right]}{C_L^f\·C_M^f}=K---\left(4\right)$

Wherein Q is thermal change each time, and V is volume, Be molar reaction enthalpy, n _LBe the total amount of substance of reactant in the sample cell, With

The total concentration of two kinds of injections in the difference acid-base titration pond, representative sample pond,

Be with sample cell in the vs concentration of example reaction,

With

The unreacted concentration of difference representative sample pond acid-base titration liquid, K is this equilibrium constant of reaction; [ML] is the total concentration of resultant of reaction,

Can be got by formula (4):

$\frac{C_M^b}{(C_M^0-C_M^b)\·(C_L^0-C_M^b)}=K---\left(5\right)$

Can be got by formula (5):

$K{\left(C_M^b\right)}^2-(C_L^{0}K+C_M^{0}K+1)C_M^b+K{C}_L^0{\mathrm{<figure-callout\; id="0"\; label="C\; M"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_M^{}=0---\left(6\right)$

$C_M^b=\frac{({\mathrm{<figure-callout\; id="0"\; label="C\; L"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_L^{}+C_M^0+\frac{1}{K})-\sqrt{{({\mathrm{<figure-callout\; id="0"\; label="C\; L"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_L^{}+{\mathrm{<figure-callout\; id="0"\; label="C\; M"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_M^{}+\frac{1}{K})}^2-4C_L^0{\mathrm{<figure-callout\; id="0"\; label="C\; M"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_M^{}}}{2}---\left(7\right)$

With (6) formula substitution (2) Shi Kede:

$\frac{Q}{n_L}={\mathrm{\&Delta;}}_r{H}_m^{\mathrm{\&theta;}}\&CenterDot;\frac{({\mathrm{<figure-callout\; id="0"\; label="C\; L"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_L^{}+C_M^0+\frac{1}{K})-\sqrt{{({\mathrm{<figure-callout\; id="0"\; label="C\; L"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_L^{}+{\mathrm{<figure-callout\; id="0"\; label="C\; M"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_M^{}+\frac{1}{K})}^2-4C_L^0{C}_M^0}}{2C_L^0}---\left(8\right)$

$=\frac{{\mathrm{\&Delta;}}_r{H}_m^{\mathrm{\&theta;}}}{2}\&CenterDot;[(1+\frac{C_M^0}{{\mathrm{<figure-callout\; id="0"\; label="C\; L"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_L^{}}+\frac{1}{C_L^{0}K})-\sqrt{{(1+\frac{C_M^0}{{\mathrm{<figure-callout\; id="0"\; label="C\; L"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_L^{}}+\frac{1}{C_L^{0}K})}^2-4\frac{C_M^0}{C_L^0}}]---\left(9\right)$

In the step (5), the calculating of Entropy Changes Δ S calculates by following formula:

ΔG＝-RTlnK (10)

ΔG＝ΔH-TΔS (11)

Wherein, Δ G is Gibbs free energy.

The present invention compared with prior art has following beneficial effect: (1) ITC has characteristics such as highly sensitive, real-time online, and has the differential thermal power compensating system, and balance is to adapt to Continuous Titration fast; (2) need not to know that whole compositions can be judged reaction type by parameter in the solution, especially suitable to material base indefinite (as traditional Chinese medicine); (3) the ITC method can remedy effectively that no reference substance, composition characteristics absorption spectra or absorbing wavelength are difficult to determine and the limitation that causes the conventional chemical analytical approach to detect, and has universality and reactivity dactylogram feature preferably.

Description of drawings

Fig. 1 is the fused power-time spectrum of 5% glucose injection and Yiqi and vein recovery freeze-dried powder;

Fig. 2 is the fused heat of 5% glucose injection and Yiqi and vein recovery freeze-dried powder-titration number of times spectrum;

Fig. 3 is the fused power-time spectrum of 5% vitamin C injection and Yiqi and vein recovery freeze-dried powder;

Fig. 4 is the fused heat of 5% vitamin C injection and Yiqi and vein recovery freeze-dried powder-titration number of times spectrum;

Fig. 5 is the comparison of thermodynamic parameter;

Fig. 6 is finger-print and Yiqi and vein recovery freeze-dried powder and the mixed chemical fingerprint of glucose injection of Yiqi and vein recovery freeze-dried powder;

Fig. 7 is Yiqi and vein recovery freeze-dried powder and the mixed chemical fingerprint of vitamin C injection;

Fig. 8 is the enlarged drawing of Fig. 7's (a) part collection of illustrative plates;

Fig. 9 is the enlarged drawing of Fig. 7's (b) part collection of illustrative plates;

Figure 10 is the enlarged drawing of Fig. 7 (c) part collection of illustrative plates;

Figure 11 is the enlarged drawing of Fig. 7 (d) part collection of illustrative plates.

Embodiment

For ease of the understanding of technical solution of the present invention, be introduced below in conjunction with concrete embodiment.

Experiment is the pattern medicine with Yiqi and vein recovery freeze-dried powder (YQFM), select vitamin C injection (Vc) and 5% glucose injection (5%GS) for share injection according to its clinical drug combination commonly used, employing ITC method share injection to two kinds and the fused after heat measure feature information of Yiqi and vein recovery freeze-dried powder aqueous solution is gathered, obtain the reactivity spectrum, obtain corresponding thermodynamic parameter, judge the type of reaction, (HPLC) verified with high performance liquid chromatography.

Experimental apparatus and material:

NANO ITC ^2GIsothermal titration calorimeter (U.S. TA company).The instrument lowest detection is limited to 0.1nJ, and baseline stability is 20nW, and 25 ℃ is that temperature stability is 0.0002 ℃, and working temperature is 2～80 ℃, corresponding time 12s.High performance liquid chromatograph 1200 (U.S. Anjelen Sci. ﹠ Tech. Inc).Injection Yiqi and vein recovery freeze-dried powder (lot number: 090702, Tianjin TianShiLi ZhiJiao Medicine Co., Ltd), sterile water for injection (lot number: 09110118, Yongkang, Beijing pharmaceutcal corporation, Ltd), 5% glucose injection (lot number: 091028257, Shijiazhuang Siyao Co., Ltd), vitamin C injection (lot number: 1001301, Tianjin gold credit amino acid company limited), acetonitrile (Fisher).Reference substance ginsenoside Rb ₁(1110704-200420) available from Nat'l Pharmaceutical ﹠ Biological Products Control Institute.

Experimental technique and result

1. isothermal titration calorimetry

1.1 the preparation of sample liquid

It is an amount of to get YQFM, adds the sterile water for injection dissolving, and making its final concentration is 0.052gmL ^-15%GS, Vc uncork are namely used.

1.2 titration

In sample cell, fill with titrated solution, constant temperature to 25 ℃, the control stir speed (S.S.) is 300rmin ^-1, treat the instrument self-poise after, the record 60s, the beginning titration, 25 of Continuous Titration, every 4 μ L, be 300s interval time.Launch ITCRun software carries out data acquisition.

1.3 the generation of reactivity spectrum

The isothermal titration calorimeter obtains the fused course of reaction spectrum (power-time spectrum) of two kinds of parenteral solutions by the thermal change that records drop reaction generation each time, if the collection of illustrative plates peak upwards, then fuses and emits thermal change figure (heat-titration number of times spectrum) in the process; If the collection of illustrative plates peak is downward, then absorb heat in the fused process.Respectively each titration peak is carried out integration, then obtain the collection of illustrative plates of thermal change in the Continuous Titration process.Course of reaction spectrum and thermal change spectrum are referred to as the reactivity spectrum.

Use the ITC method and investigate the compatibility of YQFM and 5%GS, Vc respectively, reactivity is composed as Fig. 1, Fig. 2, Fig. 3, shown in Figure 4.The I curve is two kinds of injection titration results among Fig. 1, Fig. 3; Be the influence of deduction vs to solvent in the sample cell, investigated the drop reaction of 5%GS, Vc and sterilized water, shown in II curve among Fig. 1, Fig. 3.After the influence of deduction vs to solvent, the titration peak is carried out integration, the titration process thermal change collection of illustrative plates that obtains such as Fig. 2, shown in Figure 4.

1.4 the calculating of thermodynamic parameter enthalpy change Δ H and reaction constant K

Titration each time all can be accompanied by the variation of heat, and reaches balance.Suppose that thermal change each time is Q, then obtain corresponding accounting equation:

$Q=V\&CenterDot;\mathrm{\&Delta;H}\&CenterDot;C_M^b---\left(1\right)$

$\frac{Q}{n_L}={\mathrm{\&Delta;}}_r{H}_M^{\mathrm{\&theta;}}\&CenterDot;\frac{C_M^b}{C_L^0}---\left(2\right)$

$\frac{\left[\mathrm{ML}\right]}{C_L^f\&CenterDot;C_M^f}=K---\left(4\right)$

More than various in, V is volume,

Be molar reaction enthalpy, n _LBe the total amount of substance of reactant in the sample cell,

With

The total concentration of two kinds of injections in the difference acid-base titration pond, representative sample pond,

Be with sample cell in the vs concentration of example reaction,

With

The unreacted concentration of difference representative sample pond acid-base titration liquid, K is this equilibrium constant of reaction; [ML] is the total concentration of resultant of reaction,

Can be got by formula (4):

$\frac{C_M^b}{(C_M^0-C_M^b)\&CenterDot;(C_L^0-C_M^b)}=K---\left(5\right)$

By (5) Shi Kede:

$K{\left(C_M^b\right)}^2-(C_L^{0}K+C_M^{0}K+1)C_M^b+K{C}_L^0{\mathrm{<figure-callout\; id="0"\; label="C\; M"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_M^{}=0---\left(6\right)$

$C_M^b=\frac{({\mathrm{<figure-callout\; id="0"\; label="C\; L"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_L^{}+{\mathrm{<figure-callout\; id="0"\; label="C\; M"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_M^{}+\frac{1}{K})-\sqrt{{({\mathrm{<figure-callout\; id="0"\; label="C\; L"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_L^{}+{\mathrm{<figure-callout\; id="0"\; label="C\; M"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_M^{}+\frac{1}{K})}^2-4C_L^0{\mathrm{<figure-callout\; id="0"\; label="C\; M"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_M^{}}}{2}---\left(7\right)$

With (7) formula substitution (2) Shi Kede:

$\frac{Q}{n_L}={\mathrm{\&Delta;}}_r{H}_m^{\mathrm{\&theta;}}\&CenterDot;\frac{({\mathrm{<figure-callout\; id="0"\; label="C\; L"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_L^{}+{\mathrm{<figure-callout\; id="0"\; label="C\; M"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_M^{}+\frac{1}{K})-\sqrt{{({\mathrm{<figure-callout\; id="0"\; label="C\; L"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_L^{}+{\mathrm{<figure-callout\; id="0"\; label="C\; M"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_M^{}+\frac{1}{K})}^2-4C_L^0{\mathrm{<figure-callout\; id="0"\; label="C\; M"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_M^{}}}{2C_L^0}---\left(8\right)$

$=\frac{{\mathrm{\&Delta;}}_r{H}_m^{\mathrm{\&theta;}}}{2}\&CenterDot;[(1+\frac{C_M^0}{{\mathrm{<figure-callout\; id="0"\; label="C\; L"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_L^{}}+\frac{1}{C_L^{0}K})-\sqrt{{(1+\frac{C_M^0}{{\mathrm{<figure-callout\; id="0"\; label="C\; L"\; filenames="HSA00000245997400011.png,HSA00000245997400021.png"\; state="\{\{state\}\}">C</figure-callout>}}_L^{}}+\frac{1}{C_L^{0}K})}^2-4\frac{C_M^0}{C_L^0}}]---\left(9\right)$

Launch NanoAnalyze software can use above-mentioned formula that the data of experiment gained are analyzed, and will test with the sample concentration input, wherein with ginsenoside Rb ₁The concentration of concentration (seeing 2.4 following concentration calculating) metering Yiqi and vein recovery freeze-dried powder aqueous solution.Software is analyzed the reactivity spectrum automatically, obtains two kinds of injections fused thermodynamic parameter reaction enthalpy change Δ H and reaction constant K through over-fitting.

1.5 the calculating of Entropy Changes Δ S

Try to achieve Gibbs free energy Δ G and the Entropy Changes Δ S of reaction according to thermodynamic parameter relational expression (10), (11), wherein R is 8.314KJ/mol.

Δ G=-RTlnK formula (10)

Δ G=Δ H-T Δ S formula (11)

The result of calculation of reaction enthalpy change Δ H, reaction constant K, Gibbs free energy Δ G and Entropy Changes Δ S is summed up as shown in table 1.

The thermodynamic parameter that the different parenteral solutions of table 1 are fused

1.6 the judgement of reaction type

The peak type of composing the course of reaction spectrum that YQFM as can be seen and two kinds of injection titration produce from reactivity has larger difference.Wherein the collection of illustrative plates gradually changeable with 5%GS titration generation is not strong, and average every liberated heat is about 40 μ J in the thermal change collection of illustrative plates, and the molar reactive enthalpy change (| Δ H|=0.04kJ/mol) less than 20kJ/mol, in the fused process of prompting YQFM and 5%GS physical reactions has taken place mainly.The course of reaction spectrum gradually changeable of YQFM and Vc titration is strong, rule is obvious, and every liberated heat is all greater than 1800 μ J in the thermal change spectrum, and the molar reactive enthalpy change (| Δ H|=50.00kJ/mlo) greater than 40kJ/mol, in prompting YQFM and the Vc titration process, generation or the fracture of chemical bond may be arranged.

As can be seen from Table 1, after the effect of deduction titration injection to solvent, the Δ G of 5%GS, Vc and YQFM reaction is negative value, can judge fusing of two kinds of parenteral solutions and Yiqi and vein recovery freeze-dried powder thus spontaneous reaction has all taken place.Formula (11) show Δ G by Δ H and-T Δ S two parts form, these two parts have determined the type of reacting to the size of Gibbs free energy contribution.

If | Δ H|＞T| Δ S|, then this reacts for enthalpy drives reaction, is chemical reaction.

If | Δ H|＜T| Δ S|, then this reacts for entropy drives reaction, is physical reactions.

Δ G, the Δ H that titration produces between different injections ,-T Δ S sees Fig. 5.The fused process that can intuitive judgment from Fig. 5 goes out between Vc and the YQFM is chemical reaction, and the fused process between 5%GS and the YQFM is physical reactions.Can infer: after Vc and YQFM were fused, transformation had taken place in material base.Therefore, GS can mix use with the Yiqi and vein recovery freeze-dried powder, but before illustrating the influence of material base change to body, should avoid Vc to mix use with the Yiqi and vein recovery freeze-dried powder in the clinical application.

2. efficient liquid-phase chromatography method

2.1 test sample preparation

It is an amount of to get the YQFM content, is dissolved in the sterile water for injection, as mother liquor.Getting the mother liquor portion adds aqua sterilisa and is diluted to 0.026gmL ^-1Other gets the mother liquor portion and adds sterilized water and be diluted to 0.052gmL ^-1, get dilution back solution respectively and mix with equal-volume 5%GS, Vc, be prepared into mixed solution.

2.2 standard items preparation

Precision takes by weighing ginsenoside Rb ₁Reference substance 6mg, methanol constant volume is in the 50mL volumetric flask, as the standard items mother liquor.It is 10,20,30,40,50 μ gmL that the standard items mother liquor is diluted ^-1

2.3 liquid-phase condition

Chromatographic column HALO C18, (4.6mm * 75mm, 2.7 μ m).Flowing is A mutually, water; B, acetonitrile; A-B (85:15).Gradient elution: 0-5min, 15%B; 5-10min, 15%-17%B; 10-20min, 17%-27%B; 20-25min, 27%-33.5%B; 25-45min, 33.5%-60%B; 45-50min, 60%-50%B.Flow velocity 0.25mL/min detects wavelength 203nm, 30 ℃ of column temperatures, sample size 5 μ L.

2.4 saponin content is measured

According to 2.3 following liquid-phase conditions, the chemical fingerprint of working sample, as shown in Figure 6.Try to achieve and contain ginsenoside Rb in the test sample ₁Concentration is 24.8mgmL ^-1, try to achieve test sample with ginsenoside Rb according to formula (12), (13) ₁The volumetric molar concentration of metering is 44.8mM.

Molal quantity n=content (g)/molecular weight (g/mol) formula (12)

Volumetric molar concentration (M)=molal quantity (mol)/volume (L) formula (13)

2.5 the efficient liquid phase collection of illustrative plates that different injections mix

The efficient liquid phase collection of illustrative plates that 5%GS and YQFM are fused, as shown in Figure 6.As we can see from the figure, after two kinds of injections are fused, there is not the variation of tangible chromatographic peak.

The finger-print that Vc and Yiqi and vein recovery freeze-dried powder fuse as shown in Figure 7.As we can see from the figure, after two kinds of injections were fused, variation had taken place in the liquid phase collection of illustrative plates.

With after Vc, 5%GS mix, visual inspection does not have significant change to YQFM respectively.After HPLC measured, liquid phase collection of illustrative plates and former collection of illustrative plates after fused to YQFM and 5%GS, Vc respectively compared, and find that mixed collection of illustrative plates has three kinds of variation tendencies, and the first, variation has taken place in the number at peak, as chromatographic peak minimizing among Fig. 8, chromatographic peak increase among Fig. 9; The second, variation has taken place in retention time, as shown in figure 10; The 3rd, variation has taken place in relative peak area, as shown in figure 11.Use the included angle cosine method and carry out similarity analysis, the result shows, the fused back of YQFM and 5%GS and YQFM collection of illustrative plates similarity be 0.90 and YQFM and Vc fused after and YQFM collection of illustrative plates similarity be 0.56.As seen YQFM and Vc fused after, make the YQFM collection of illustrative plates that significant variation take place, and 5%GS and YQFM fused after, collection of illustrative plates is significantly change not.Prompting thus: Vc can make the YQFM chemical constitution change.

This experimental result is proved ITC method gained result, demonstrates accuracy and reliability that the ITC method qualitatively judges.

Claims (4)

1. a method of estimating injection clinical compatibility compatibility is characterized in that, this method comprises following step:

(1) the pattern medicine of preparation compatibility to be detected and the sample liquid of drug combination;

(2) sample liquid of (1) step preparation is carried out titration at the isothermal titration calorimeter;

(3) utilize the thermal change that produces in the isothermal titration calorimeter real time record drop reaction process, reaction of formation activity profile;

(4) basis (3) goes on foot the reactivity spectrum of the thermal change that obtains, and calculates thermodynamic parameter enthalpy change Δ H and reaction constant K by following formula,

（1），

（2），

（3），

（4），

（5），

（6），

（7），

（8），

（9）；

Wherein Q is thermal change each time, and V is volume, Be molar reaction enthalpy,

Be the total amount of substance of reactant in the sample cell,

With

The total concentration of two kinds of injections in the difference acid-base titration pond, representative sample pond,

Be with sample cell in the vs concentration of example reaction,

The unreacted concentration of representative sample liquid,

Representative not with sample cell in the vs concentration of example reaction, K is this equilibrium constant of reaction;

Be the total concentration of resultant of reaction,

=

,

=

-

=

-

(5) according to (4) thermodynamic parameter enthalpy change Δ H and the reaction constant K in step, calculate the Entropy Changes Δ S of reaction, Entropy Changes Δ S calculates by following formula:

ΔG=﹣RTlnK （10）

ΔG=ΔH﹣TΔS （11）

Δ G in the above-mentioned formula is Gibbs free energy;

(6) according to the thermodynamic parameter in (4), (5) step, judge reaction type, if ＞

, be that enthalpy drives reaction, illustrate that these two kinds of injections after fused chemical reaction have taken place, incompatible, can not drug combination, wherein T is temperature; If ＜

, for entropy drives reaction, illustrating that physical reactions only takes place the fused of these two kinds of injections, chemical compatibility is good.

2. the method for evaluation injection clinical compatibility compatibility according to claim 1, it is characterized in that, if pattern medicine to be detected and drug combination are parenteral solutions, then the sample liquid of step (1) can directly be measured configuration from the parenteral solution of pattern medicine and drug combination; If pattern medicine to be detected and drug combination are freeze-dried powder, then get an amount of pattern medicine and drug combination respectively, be made into pattern drug solns and the drug combination solution of clinical working concentration.

3. the method for evaluation injection clinical compatibility compatibility according to claim 1 is characterized in that, the titration operation that carries out at the isothermal titration calorimeter in the step (2) comprises following step:

(1) vs is pattern medicine sample liquid or drug combination sample liquid;

(2) titrate constant temperature to 25 ℃ is at 100 ~ 300rmin ^-1Stir speed (S.S.) under be stirred to the instrument self-poise after, the output power curve of register instrument is as baseline, be 30 ~ 60s writing time;

(3) with vs titrate is carried out titration, 20 ~ 50 of Continuous Titration, every 2 ~ 10 μ L, be 200 ~ 300s interval time;

(4) in the titration process, adopt Launch ITCRun software to carry out data acquisition.

4. the method for evaluation injection clinical compatibility compatibility according to claim 1, it is characterized in that, the reactivity spectrum that generates according to the thermal change in the titration process in the step (3) is made up of course of reaction spectrum and thermal change spectrum, wherein the course of reaction spectrum is the power-time spectrum that obtains two kinds of parenteral solution titration processes by the thermal change that records drop reaction generation each time, and the thermal change spectrum is to carry out heat-titration number of times spectrum that integration obtains by the titration peak to the course of reaction spectrum.

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