CN105181866A - Method for rapid detection of benzalkonium chloride in eye drops - Google Patents
Method for rapid detection of benzalkonium chloride in eye drops Download PDFInfo
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- CN105181866A CN105181866A CN201510539781.6A CN201510539781A CN105181866A CN 105181866 A CN105181866 A CN 105181866A CN 201510539781 A CN201510539781 A CN 201510539781A CN 105181866 A CN105181866 A CN 105181866A
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- eye drops
- benzalkonium chloride
- dimethyl ammonium
- benzyl dimethyl
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Abstract
The invention discloses a method for rapid detection of benzalkonium chloride in eye drops. The method comprises the following steps: a pure signal of dodecyl dimethyl benzyl ammonium chloride which is a component to be detected is extracted from signals of a benzalkonium chloride mixture by a pure signal extraction method in a multi-variable system of oblique projection, and a standard spectrum database is established by virtue of least square fitting of the extracted pure signal and a corresponding concentration value. Then, the content of the component to be detected in an eye drop sample is calculated according to a spatial included angle criterion. According to the invention, benzalkonium chloride is directly quantified by means of a multi-wavelength ultraviolet spectrum, separation of benzalkonium chloride is not needed, and the method has the advantages of short sample measurement time and quantitative calculation time, and high analysis efficiency; spectrum signal extraction and quantitative analysis are realized by a software method, so that the analysis cost is reduced, and apparatus and reagent losses are reduced; the combination method of high performance liquid chromatography and ultraviolet spectrum is used, thereby improving analysis precision and applicability of the method; the method has good anti-interference capability, high identification degree and high robustness.
Description
Technical field
The invention belongs to drug ingredient detection technique field, particularly relate to the method for quick of benzalkonium chloride in a kind of eye drops.
Background technology
For preventing eye-drops preparations by microbial contamination in storage and use procedure, most of eye drops especially multiple dose eye drops often adds bacteriostatic agent.Benzalkonium chloride (BAC) because having the features such as good bactericidal effect, toxicity are little during its low concentration, by as bacteriostatic agent conventional in eye drops.The detection method of current benzalkonium chloride is mainly high performance liquid chromatography, high performance liquid chromatography can effectively measure benzalkonium chloride content, but its chromatographic peak easily trails, post effect is difficult to satisfy the demands, and the analyzing samples time is long, the analysis time of a sample reaches 20 minutes, even more of a specified duration, for the analysis operation of sample in enormous quantities, the equal reagent loss that flows is large, analysis cost is high, and manipulation strength is large.In addition, benzalkonium chloride is made up of the homolog of 3 kinds of nalka hydrocarbyl substituent dimethyl benzyl ammonium chlorides, and meeting is inconsistent because homolog in reference substance and eye drops product forms causes puzzlement to analyzing detection.In addition mass spectrometry is also for benzalkonium chloride assay, but its instrument is expensive, and popularization degree is little, and can failure analysis sample.
In recent years, the fast development of chemometrics method brings new life to substances of interest analysis in complex sample, but lacks very much for the chemometrics method analyzing antibacterial component in eye drops.Wherein space angle criterion is as a kind of new chemometrics method, obtains more application because it has the advantages such as specimen in use quantity is few, pre-service is simple, method is easy, quick.
Summary of the invention
The object of the present invention is to provide the method for quick of benzalkonium chloride in a kind of eye drops, be intended to solve traditional sensing techniques exist analysis efficiency low, manipulation strength is large and the high deficiency of analysis cost, and component pure spectrum signal to be measured be difficult to obtain problem.
The present invention realizes like this, a method for quick for benzalkonium chloride in eye drops, component dodecyl benzyl dimethyl ammonium chloride to be measured is extracted by pure signal extracting method in the multivariate system of oblique projection by described method for quick from benzalkonium chloride potpourri; The dodecyl benzyl dimethyl ammonium chloride pure signal of extraction and corresponding concentration value are carried out least square fitting Criterion spectra database; Then space angle criterion is adopted to try to achieve the content of component to be measured in eye drops sample.In the present invention, benzalkonium chloride content presses dodecyl benzyl dimethyl ammonium chloride note.
Specifically comprise the following steps:
The first step, Benza is prepared:
Pipette the benzalkonium chloride of 12.02mg/mL, use redistilled water constant volume, preparation series concentration is the benzalkonium chloride standard solution of 1 ~ 20 μ g/mL;
Second step, the foundation of benzalkonium chloride background spectra database:
Through efficient liquid phase chromatographic analysis after being filtered by the benzalkonium chloride standard solution of preparation, gather the multi-wavelength three-dimensional data of serial benzalkonium chloride standard solution through ultraviolet spectrometer (UVS); The light intensity data value gathering the multi-wavelength three-dimensional data of the benzalkonium chloride of gained changes into absorbance; The retention time of component dodecyl benzyl dimethyl ammonium chloride to be measured is determined according to the substance characteristics of dodecyl benzyl dimethyl ammonium chloride, myristyl benzyl dimethyl ammonium chloride in benzalkonium chloride potpourri, according to benzalkonium chloride chromatogram, from benzalkonium chloride standard solution three-dimensional data, deduct the spectroscopic data of component to be measured, remaining light modal data to be added and as benzalkonium chloride background spectra database H; The background spectra database H of gained is carried out svd, according to system major component number, background spectra database H is reconstructed, obtain the background spectra database H ' that dimension is suitable;
3rd step, according to the time point that the chromatogram of benzalkonium chloride finds the highest response of component to be measured corresponding, deducts component spectra signal to be measured as component pure signal S to be measured;
4th step, the benzalkonium chloride standard solution adopting fiber spectrometer obtained to the first step carries out spectral scan, obtains the ultraviolet spectrum data matrix y of benzalkonium chloride standard solution;
5th step, the benzalkonium chloride background spectra database H ' of gained, the pure analytic signal S of component to be measured and benzalkonium chloride standard solution ultraviolet spectrum data matrix y are imported computing platform, component dodecyl benzyl dimethyl ammonium chloride to be measured extracts based on pure signal extracting method in the multivariate system of oblique projection by application from benzalkonium chloride potpourri, obtains the stable component pure spectrum signal matrix S ' to be measured of the series concentration of 1 ~ 20 μ g/mL;
6th step, carries out least square fitting by concentration value corresponding with it for the component pure spectrum signal to be measured obtained, obtains the dodecyl benzyl dimethyl ammonium chloride spectroscopic data of unit concentration, stored in standard spectral data storehouse V; The standard spectrum storehouse V obtained can be used for next step and calculates.
Further, describedly specifically to comprise based on pure signal extracting method in the multivariate system of oblique projection:
Step one, the H ', the S that try to achieve are imported computing platform, and ask for pure signal disjunctive model, i.e. oblique projection operator, expression formula is:
E
sH=S (S
tp
h ⊥s)
-1s
tp
h ⊥, wherein P
h ⊥=I-P
h=I-H (H
th)
-1h
t;
In above formula, the transposition of subscript T representing matrix, I is and P
hthe unit matrix that dimension is identical;
Step 2, by obtain y and oblique projection operator E
s|Hbe multiplied, namely obtain the dodecyl benzyl dimethyl ammonium chloride pure spectrum signal S ' of the series concentration of 1 ~ 20 μ g/mL.
Further, need after described Criterion spectra database to build sample background spectra database by liquid phase-ultraviolet coupling; Adopt svd to choose system number of principal components to Fundamental database dimensionality reduction in conjunction with second order difference method, obtain the background spectra database that data volume is less; Obtain eye drops sample ultraviolet spectrum data.
Further, further comprising the steps:
Step one, pipettes eye drops sample in the volumetric flask of 10mL, dilutes suitable multiple with redistilled water, obtained eye drops dilution;
Step 2, the foundation of eye drops sample background spectra database:
Step 3, adopts fiber spectrometer to carry out spectral scan to obtained eye drops dilution, obtains the ultraviolet spectrum data matrix a of eye drops dilution.
Further, the foundation of described eye drops sample background spectra database specifically comprises:
The first step, obtains the liquid phase-ultraviolet coupling data of eye drops sample, through efficient liquid phase chromatographic analysis after being filtered by not diluted eye drops sample, gathers the multi-wavelength three-dimensional data of eye drops sample through ultraviolet spectrometer (UVS).
Second step, the light intensity data value of the eye drops sample multi-wavelength three-dimensional data gathering gained is changed into absorbance, the every a line of data after conversion represents the chromatographic data under a wavelength, inscribes the spectroscopic data eluting material when each row represents one;
3rd step, obtain not containing the bias light modal data of component to be measured, according to the retention time of the component dodecyl benzyl dimethyl ammonium chloride to be measured determined, from eye drops sample three-dimensional data, deduct the spectroscopic data of component to be measured, all the other spectroscopic datas to be added and as eye drops background spectra database H
1;
4th step, the dimensionality reduction of background spectra database, by the background spectra database H of gained
1carry out svd, according to system major component number by background spectra database H
1be reconstructed, obtain the background spectra database H that dimension is suitable
1'.
Further, in the dimensionality reduction of described local spectra database, system number of principal components object defining method is: by matrix H, H
1after carrying out variance criterion, add the interference that white noise shelters uneven noise and non-linear factor, judge its independent variable number with second order difference value sequence turning point, obtain system major component number N, N
1;
By background spectra database H, H
1the method be reconstructed is as follows:
Application svd program [US
1v
1]=svd (H) carries out svd dimensionality reduction to benzalkonium chloride background spectra database H, obtains m rank matrix with orthogonal rows U after decomposition, n rank row orthogonal matrix V
1with the eigenvalue matrix S of matrix H
1, the front N getting U arranges the benzalkonium chloride background spectra database H ' after being dimensionality reduction.
H is obtained with method
1'.
Further, need dodecyl benzyl dimethyl ammonium chloride assay in eye drops sample after described acquisition eye drops sample ultraviolet spectrum data, respectively by the Component Standard spectra database V to be measured of gained, eye drops background spectra database H
1' and eye drops sample ultraviolet spectrum data matrix a importing computing platform, application space angle criterion reduces the spectrum of component dodecyl benzyl dimethyl ammonium chloride to be measured from eye drops Sample dilution spectrum, draws the content of component dodecyl benzyl dimethyl ammonium chloride to be measured in eye drops dilution.
Further, further comprising the steps:
Step one, reduces step delta according to quantitative accuracy setting;
Step 2, at formula y
i=a
ix+b
iin bring larger x into
1value, obtains v
1;
Y
irepresent the absorbance of dodecyl benzyl dimethyl ammonium chloride under i wavelength, a
i, b
ibe constant, x represents the concentration of dodecyl benzyl dimethyl ammonium chloride, v
1represent in concentration to be x
1time dodecyl benzyl dimethyl ammonium chloride multi-wavelength absorbance, v
1for all y
ithe matrix of value composition;
Step 3, deducts v from eye drops dilution sample light modal data a
1/ Δ, the variable after deduction is designated as da; Background spectra database H
1' merge postscript for contrast space M with variable da, calculate and contrast space M and v
1angle;
Step 4, progressively deducts v from eye drops dilution sample light modal data a
1after, repeat step 3;
Step 5, after the dodecyl benzyl dimethyl ammonium chloride in eye drops dilution sample is deducted completely, the spectral vector v of comparison space M and dodecyl benzyl dimethyl ammonium chloride
1space angle value there will be maximal value θ
max, record space angle maximal value θ
maxcorresponding reduces step number λ, by the concentration x of dodecyl benzyl dimethyl ammonium chloride
1with the content Y reducing dodecyl benzyl dimethyl ammonium chloride in step number λ estimation eye drops dilution sample
1, calculating formula is Y
1=x
1/ Δ × λ, the Y obtained
1be the content value of dodecyl benzyl dimethyl ammonium chloride in eye drops dilution sample; If Y
1value and x
1value difference is comparatively large, then bring one and Y into
1be close and be greater than Y
1x
2recalculate;
Step 6, namely tries to achieve the content of dodecyl benzyl dimethyl ammonium chloride in actual eye drops sample according to extension rate.
The method for quick of benzalkonium chloride in eye drops provided by the invention, compared with prior art, has following advantage:
1. analysis efficiency is high, manipulation strength is little:
Because the present invention adopts multi-wavelength ultraviolet spectrum direct quantitative benzalkonium chloride, do not need to be separated, and sample is measured and quantitative computing time is short, its analysis efficiency is higher, and manipulation strength is less, and the express-analysis being highly suitable for similar sample in enormous quantities measures.
2. analysis cost is low:
Because the present invention does not propose particular/special requirement to instrument, only need simple setting just can continue to use common single order spectral instrument and export data, special requirement are not proposed to data dimension, after data acquisition, by the data processing step that the present invention proposes, namely realize dodecyl benzyl dimethyl ammonium chloride spectral signal by mathematics separation method to extract, and quantitative test.Thus manufacture difficulty and the production cost of instrument can not be improved.And reagent consumption significantly reduces, reduce analysis cost.
3. the Fundamental database of tested component is not contained without the need to accumulation:
Because eye drops is of a great variety, raw material is complicated, sets up complete Fundamental database workload large, the applicability of impact analysis process, analysis precision and method.And the present invention is by the method for high performance liquid chromatography-ultraviolet spectrum coupling, obtain the light intensity three-dimensional data of eye drops each moment of sample, each wavelength, and obtain not containing the Fundamental database of tested component.The method not only reduces working strength, and improves analysis precision, expands the applicability of method.
4. can lowering apparatus and reagent loss:
For similar sample tests, the present invention, after carrying out a chromatographic resolution, without the need to recycling at the bottom of coupling instrument cumulative volume, can realize quantitatively by means of only spectral measurement, thus reducing instrument and reagent loss.
5. robustness is good:
The present invention is compared with existing Single wavelength, dual wavelength and several wavelength method, and antijamming capability is strong, can identification degree high, robustness is good.
6. analysis method comparison of the present invention is simple, without complex steps, is easy to apply.
Accompanying drawing explanation
Fig. 1 is the method for quick process flow diagram of benzalkonium chloride in the eye drops that provides of the embodiment of the present invention;
Fig. 2 is the dodecyl benzyl dimethyl ammonium chloride ultraviolet spectrogram of the oblique projection extraction that the embodiment of the present invention provides;
Fig. 3 is the embodiment of the present invention space angle value provided and the curve map of reducing step number.
Embodiment
In order to make object of the present invention, technical scheme and advantage clearly understand, below in conjunction with embodiment, the present invention is further elaborated.Should be appreciated that specific embodiment described herein only in order to explain the present invention, be not intended to limit the present invention.
Below in conjunction with accompanying drawing, application principle of the present invention is explained in detail.
The present invention relates to the detection method of ingredient components in a kind of eye drops, adopt the dodecyl benzyl dimethyl ammonium chloride in eye drops in potpourri benzalkonium chloride to complete content detection.
As shown in Figure 1, in the eye drops of the embodiment of the present invention, the method for quick of benzalkonium chloride comprises the following steps:
S101: adopt pure signal extracting method in the multivariate system of oblique projection to be extracted from benzalkonium chloride potpourri by component dodecyl benzyl dimethyl ammonium chloride to be measured, Criterion spectra database;
S102: build sample background spectra database by liquid phase-ultraviolet coupling; Obtain eye drops sample ultraviolet spectrum data;
S103: last content of trying to achieve component dodecyl benzyl dimethyl ammonium chloride to be measured in eye drops sample according to space angle criterion, completes mensuration.
The concrete steps of the embodiment of the present invention are as follows:
The method for quick of benzalkonium chloride in a kind of eye drops of the embodiment of the present invention, comprises the following steps:
I, extraction dodecyl benzyl dimethyl ammonium chloride pure signal, Criterion spectra database.
A. Benza preparation:
Accurately pipette the benzalkonium chloride of appropriate 12.02mg/mL, use redistilled water constant volume, preparation series concentration is the benzalkonium chloride standard solution of 1 ~ 20 μ g/mL.
Described benzalkonium chloride 1 ~ 20 μ g/mL series concentration wherein benzalkonium chloride presses dodecyl benzyl dimethyl ammonium chloride note.
B. the foundation of benzalkonium chloride background spectra database:
SB1. liquid phase-ultraviolet coupling the data of benzalkonium chloride standard solution are obtained:
Through efficient liquid phase chromatographic analysis after benzalkonium chloride standard solution steps A prepared filters, gather the multi-wavelength three-dimensional data of serial benzalkonium chloride standard solution through ultraviolet spectrometer (UVS).
In step SB1, high-efficient liquid phase chromatogram condition is:
Chromatographic column: ODS-SP92689C18 chromatographic column (4.6mm*250mm)
Isocratic elution, mobile phase: acetonitrile: 0.02mol/L ammonium acetate solution (containing triethylamine 10mL in every L, regulating pH to 5.0 ± 0.5 with glacial acetic acid)=90: 10 (V/V)
Flow velocity: 1mL/min
Determined wavelength: 262nm
Sample size: 20uL
Column temperature: room temperature.
Step SB1 medium ultraviolet fiber spectrometer condition is:
Determined wavelength scope: 196.93 ~ 400.36nm; Spectrum interval: 0.46nm; Quartz colorimetric utensil light path: 1.0cm; Take redistilled water as reference sample; Integral time: 8ms.
SB2. conversion data form:
The light intensity data value of the multi-wavelength three-dimensional data gathering the benzalkonium chloride of gained in step SB1 is changed into absorbance, the every a line of data after conversion represents the chromatographic data under a wavelength, inscribes the spectroscopic data eluting material when each row represents one;
SB3. obtain not containing the bias light modal data of component to be measured:
The retention time of component dodecyl benzyl dimethyl ammonium chloride to be measured is determined according to the substance characteristics of dodecyl benzyl dimethyl ammonium chloride, myristyl benzyl dimethyl ammonium chloride in benzalkonium chloride potpourri, documents and materials and experimental implementation, wherein the retention time of dodecyl benzyl dimethyl ammonium chloride is 8.31 ~ 9.02min, and the retention time of myristyl benzyl dimethyl ammonium chloride is 11.35 ~ 11.79min.According to benzalkonium chloride chromatogram, from benzalkonium chloride standard solution three-dimensional data, deduct the spectroscopic data of component dodecyl benzyl dimethyl ammonium chloride to be measured, all the other spectroscopic datas to be added and as benzalkonium chloride background spectra database H;
SB4. the dimensionality reduction of Fundamental database:
The database H data amount obtained due to step SB3 is large, and calculated amount will be caused very large.Therefore the background spectra database H of gained in step SB3 is carried out svd, according to system major component number, background spectra database H is reconstructed, obtain the background spectra database H ' that dimension is suitable.
C. the pure signal of component to be measured is obtained:
According to the time point 8.49min that the chromatogram of benzalkonium chloride finds the highest response of component to be measured corresponding, from benzalkonium chloride standard solution three-dimensional data, deduct component spectra signal to be measured as component pure signal S to be measured.
D. benzalkonium chloride standard solution spectrum measuring:
Adopt fiber spectrometer to carry out spectral scan to the benzalkonium chloride standard solution that steps A obtains 1 ~ 20 μ g/mL, obtain the multi-wavelength ultraviolet spectrum data matrix y of benzalkonium chloride standard solution.
E. component pure signal to be measured is extracted:
The benzalkonium chloride background spectra database H ' of step SB4, C and D gained, the pure analytic signal S of component to be measured and benzalkonium chloride standard solution ultraviolet spectrum data matrix y are imported computing platform, component dodecyl benzyl dimethyl ammonium chloride to be measured extracts based on pure signal extracting method in the multivariate system of oblique projection by application from benzalkonium chloride potpourri, obtains the stable component pure spectrum signal matrix S ' to be measured of the series concentration of 1 ~ 20 μ g/mL.
Described based on pure signal extracting method in the multivariate system of oblique projection is:
(1) H ', the S that step SB4, C are tried to achieve import computing platform, and ask for pure signal disjunctive model, i.e. oblique projection operator, the expression formula of this operator is:
E
s|H=S (S
tp
h ⊥s)
-1s
tp
h ⊥, wherein P
h ⊥=I-P
h=I-H (H
th)
-1h
t;
In above formula, the transposition of subscript T representing matrix, I is and P
hthe unit matrix that dimension is identical;
(2) y and oblique projection operator E will be obtained in step D
s|Hbe multiplied, namely obtain dodecyl benzyl dimethyl ammonium chloride pure spectrum signal S ' (see Fig. 2) of the series concentration of 1 ~ 20 μ g/mL;
F. the foundation of Component Standard library of spectra to be measured:
Concentration value corresponding with it for the component pure spectrum signal to be measured obtained in step e is carried out least square fitting, obtains the dodecyl benzyl dimethyl ammonium chloride spectroscopic data of unit concentration, stored in standard spectral data storehouse V;
The standard spectrum storehouse V obtained can be used for next step and calculates.
The assay of component to be measured in II, eye drops sample.
G. eye drops sample process:
Accurately pipette appropriate eye drops sample in the volumetric flask of 10mL, dilute suitable multiple with redistilled water, obtained eye drops dilution.
H. the foundation of eye drops sample background spectra database:
SH1. liquid phase-ultraviolet coupling the data of eye drops sample are obtained:
Through efficient liquid phase chromatographic analysis after being filtered by not diluted eye drops sample, gather the multi-wavelength three-dimensional data of eye drops sample through ultraviolet spectrometer (UVS).
The high-efficient liquid phase chromatogram condition used in step SH1 is consistent with the high-efficient liquid phase chromatogram condition used in step SB1.
The UV fiber spectrometer used in the UV fiber spectrometer condition used in step SH1 and step SB1 is consistent.
SH2. conversion data form:
The light intensity data value of the eye drops sample multi-wavelength three-dimensional data gathering gained in step SH1 is changed into absorbance, the every a line of data after conversion represents the chromatographic data under a wavelength, inscribes the spectroscopic data eluting material when each row represents one;
SH3. obtain not containing the bias light modal data of component to be measured:
According to the retention time of the component dodecyl benzyl dimethyl ammonium chloride to be measured that step SB3 determines, from eye drops sample three-dimensional data, deduct the spectroscopic data of component to be measured, all the other spectroscopic datas to be added and as eye drops background spectra database H
1;
SH4. the dimensionality reduction of background spectra database:
Due to the database H that step SH3 obtains
1data volume is large, and calculated amount will be caused very large.Therefore by the background spectra database H of gained in step SH3
1carry out svd, according to system major component number by background spectra database H
1be reconstructed, obtain the background spectra database H that dimension is suitable
1'.
I. eye drops sample spectrum measuring:
Adopt fiber spectrometer to carry out spectral scan to eye drops dilution obtained in step G, obtain the ultraviolet spectrum data matrix a of eye drops dilution;
J. dodecyl benzyl dimethyl ammonium chloride assay in eye drops sample:
Respectively by the Component Standard spectra database V to be measured of step F, SH4 and I gained, eye drops background spectra database H
1' and eye drops sample ultraviolet spectrum data matrix a importing computing platform, application space angle criterion reduces the spectrum of component dodecyl benzyl dimethyl ammonium chloride to be measured from eye drops Sample dilution spectrum, draws the content of component dodecyl benzyl dimethyl ammonium chloride to be measured in eye drops dilution;
In described eye drops sample, the concrete steps of dodecyl benzyl dimethyl ammonium chloride assay are as follows:
(1) step delta (the present embodiment is 1000) is reduced according to quantitative accuracy setting;
(2) at formula y
i=a
ix+b
iin bring larger x into
1value, obtains v
1;
Described y
irepresent the absorbance of dodecyl benzyl dimethyl ammonium chloride under i wavelength, a
i, b
ibe constant, x represents the concentration of dodecyl benzyl dimethyl ammonium chloride, v
1represent in concentration to be x
1time dodecyl benzyl dimethyl ammonium chloride multi-wavelength absorbance, v
1for all y
ithe matrix of value composition;
(3) from eye drops dilution sample light modal data a, v is deducted
1(the present embodiment is deduction v to/Δ
1/ 1000) variable, after deduction is designated as da; Background spectra database H
1' merge postscript for contrast space M with variable da, calculate and contrast space M and v
1angle;
(4) from eye drops dilution sample light modal data a, progressively v is deducted
1after, repeat step (3);
(5) after the dodecyl benzyl dimethyl ammonium chloride in eye drops dilution sample is deducted completely, the spectral vector v of comparison space M and dodecyl benzyl dimethyl ammonium chloride
1space angle value there will be maximal value θ
max, record space angle maximal value θ
maxcorresponding reduces step number λ (see Fig. 3, what space angle maximal value was corresponding reduces step number is 843), by the concentration x of dodecyl benzyl dimethyl ammonium chloride
1with the content Y reducing dodecyl benzyl dimethyl ammonium chloride in step number λ estimation eye drops dilution sample
1, calculating formula is Y
1=x
1/ Δ × λ, the Y obtained
1be the content value of dodecyl benzyl dimethyl ammonium chloride in eye drops dilution sample.
If Y
1value and x
1value difference is comparatively large, then bring one and Y into
1be close and be greater than Y
1x
2recalculate.
(6) content of dodecyl benzyl dimethyl ammonium chloride in actual eye drops sample can be tried to achieve according to the extension rate in step G.
In the dimensionality reduction of the local spectra database of step SB4, SH4., described system number of principal components object defining method is:
By matrix H, H
1after carrying out variance criterion, the white noise adding some strength shelters the interference of uneven noise and non-linear factor, judges its independent variable number, obtain system major component number N, N with second order difference value sequence turning point
1.
In the dimensionality reduction of the local spectra database of step SB4, SH4., described by background spectra database H, H
1the method be reconstructed is as follows:
Application svd program [US
1v
1]=svd (H) carries out svd dimensionality reduction to benzalkonium chloride background spectra database H, obtains m rank matrix with orthogonal rows U after decomposition, n rank row orthogonal matrix V
1with the eigenvalue matrix S of matrix H
1, the front N getting U arranges the benzalkonium chloride background spectra database H ' after being dimensionality reduction;
Eye drops sample background spectra database H is obtained with method
1'
In order to verify the accuracy of measurement result of the present invention, have selected 7 kinds of commercially available eye drops samples, after high performance liquid chromatography-Fundamental database is set up in the coupling of UV fiber spectrometer, measure the content in eye drops sample by the inventive method, and with high performance liquid chromatography results contrast (see table 1).
Two kinds of method measurement results of content in table 1 eye drops
The present invention has also carried out recovery experiment, and result is as shown in table 2.
Table 2 recovery is tested
To 1
#, 3
#sample carries out Precision Experiment, and RSD is respectively 2.78%, 0.54%.
The foregoing is only preferred embodiment of the present invention, not in order to limit the present invention, all any amendments done within the spirit and principles in the present invention, equivalent replacement and improvement etc., all should be included within protection scope of the present invention.
Claims (9)
1. the method for quick of benzalkonium chloride in an eye drops, it is characterized in that, in described eye drops, component dodecyl benzyl dimethyl ammonium chloride to be measured is extracted by pure signal extracting method in the multivariate system of oblique projection by the method for quick of benzalkonium chloride from benzalkonium chloride potpourri; The dodecyl benzyl dimethyl ammonium chloride pure signal of extraction and corresponding concentration value are carried out least square fitting Criterion spectra database; Then space angle criterion is adopted to try to achieve the content of component to be measured in eye drops sample.
2. the method for quick of benzalkonium chloride in eye drops as claimed in claim 1, is characterized in that, specifically comprise the following steps:
The first step, Benza is prepared:
Pipette the benzalkonium chloride of 12.02mg/mL, use redistilled water constant volume, preparation series concentration is the benzalkonium chloride standard solution of 1 ~ 20 μ g/mL;
Second step, the foundation of benzalkonium chloride background spectra database:
Through efficient liquid phase chromatographic analysis after being filtered by the benzalkonium chloride standard solution of preparation, gather the multi-wavelength three-dimensional data of serial benzalkonium chloride standard solution through ultraviolet spectrometer (UVS); The light intensity data value gathering the multi-wavelength three-dimensional data of the benzalkonium chloride of gained changes into absorbance; The retention time of component dodecyl benzyl dimethyl ammonium chloride to be measured is determined according to the substance characteristics of dodecyl benzyl dimethyl ammonium chloride, myristyl benzyl dimethyl ammonium chloride in benzalkonium chloride potpourri, according to benzalkonium chloride chromatogram, from benzalkonium chloride standard solution three-dimensional data, deduct the spectroscopic data of component to be measured, all the other spectroscopic datas to be added and as benzalkonium chloride background spectra database H; The background spectra database H of gained is carried out svd, according to system major component number, background spectra database H is reconstructed, obtain the background spectra database H ' that dimension is suitable;
3rd step, according to the time point that the chromatogram of benzalkonium chloride finds the highest response of component to be measured corresponding, deducts component spectra signal to be measured as component pure signal S to be measured;
4th step, the benzalkonium chloride standard solution adopting fiber spectrometer obtained to the first step carries out spectral scan, obtains the ultraviolet spectrum data matrix y of benzalkonium chloride standard solution;
5th step, the benzalkonium chloride background spectra database H ' of gained, the pure analytic signal S of component to be measured and benzalkonium chloride standard solution ultraviolet spectrum data matrix y are imported computing platform, component dodecyl benzyl dimethyl ammonium chloride to be measured extracts based on pure signal extracting method in the multivariate system of oblique projection by application from benzalkonium chloride potpourri, obtains the stable component pure spectrum signal matrix S ' to be measured of the series concentration of 1 ~ 20 μ g/mL;
6th step, carries out least square fitting by concentration value corresponding with it for the component pure spectrum signal to be measured obtained, obtains the dodecyl benzyl dimethyl ammonium chloride spectroscopic data of unit concentration, stored in standard spectral data storehouse V; The standard spectrum storehouse V obtained calculates for next step.
3. the method for quick of benzalkonium chloride in eye drops as claimed in claim 1, is characterized in that, describedly specifically comprises based on pure signal extracting method in the multivariate system of oblique projection:
Step one, the H ', the S that try to achieve are imported computing platform, and ask for pure signal disjunctive model, i.e. oblique projection operator, expression formula is:
E
s|H=S (S
tp
h ⊥s)
-1s
tp
h ⊥, wherein P
h ⊥=I-P
h=I-H (H
th)
-1h
t;
In above formula, the transposition of subscript T representing matrix, I is and P
hthe unit matrix that dimension is identical;
Step 2, by the y that obtains and oblique projection operator E
s|Hbe multiplied, namely obtain the dodecyl benzyl dimethyl ammonium chloride pure spectrum signal S ' of the series concentration of 1 ~ 20 μ g/mL.
4. the method for quick of benzalkonium chloride in eye drops as claimed in claim 1, is characterized in that, need to build eye drops sample background spectra database by liquid phase-ultraviolet coupling after described Criterion spectra database; Adopt svd to choose system number of principal components to Fundamental database dimensionality reduction in conjunction with second order difference method, obtain the background spectra database that data volume is less; And obtain eye drops sample ultraviolet spectrum data.
5. the method for quick of benzalkonium chloride in eye drops as claimed in claim 4, is characterized in that, further comprising the steps:
Step one, pipettes eye drops sample in the volumetric flask of 10mL, dilutes suitable multiple with redistilled water, obtained eye drops dilution;
Step 2, the foundation of eye drops sample background spectra database:
Step 3, adopts fiber spectrometer to carry out spectral scan to obtained eye drops dilution, obtains the ultraviolet spectrum data matrix a of eye drops dilution.
6. the method for quick of benzalkonium chloride in eye drops as claimed in claim 5, it is characterized in that, the foundation of described eye drops sample background spectra database specifically comprises:
The first step, obtains the liquid phase-ultraviolet coupling data of eye drops sample, through efficient liquid phase chromatographic analysis after being filtered by not diluted eye drops sample, gathers the multi-wavelength three-dimensional data of eye drops sample through ultraviolet spectrometer (UVS);
Second step, the light intensity data value of the eye drops sample multi-wavelength three-dimensional data gathering gained is changed into absorbance, the every a line of data after conversion represents the chromatographic data under a wavelength, inscribes the spectroscopic data eluting material when each row represents one;
3rd step, obtain not containing the bias light modal data of component to be measured, according to the retention time of the component dodecyl benzyl dimethyl ammonium chloride to be measured determined, from eye drops sample three-dimensional data, deduct the spectroscopic data of component to be measured, all the other spectroscopic datas to be added and as eye drops background spectra database H
1;
4th step, the dimensionality reduction of background spectra database, by the background spectra database H of gained
1carry out svd, according to system major component number by background spectra database H
1be reconstructed, obtain the background spectra database H that dimension is suitable
1'.
7. the method for quick of benzalkonium chloride in eye drops as claimed in claim 6, it is characterized in that, in the dimensionality reduction of described local spectra database, system number of principal components object defining method is: by matrix H, H
1after carrying out variance criterion, add the interference that white noise shelters uneven noise and non-linear factor, judge its independent variable number with second order difference value sequence turning point, obtain system major component number N, N
1;
By background spectra database H, H
1the method be reconstructed following (for H):
Application svd program [US
1v
1]=svd (H) carries out svd dimensionality reduction to benzalkonium chloride background spectra database H, obtains m rank matrix with orthogonal rows U after decomposition, n rank row orthogonal matrix V
1with the eigenvalue matrix S of matrix H
1, the front N getting U arranges the benzalkonium chloride background spectra database H ' after being dimensionality reduction.
8. the method for quick of benzalkonium chloride in eye drops as claimed in claim 1, it is characterized in that, need dodecyl benzyl dimethyl ammonium chloride assay in eye drops sample after described acquisition eye drops sample ultraviolet spectrum data, respectively by the Component Standard spectra database V to be measured of gained, eye drops background spectra database H
1' and eye drops sample ultraviolet spectrum data matrix a importing computing platform, application space angle criterion algorithm reduces the spectrum of component dodecyl benzyl dimethyl ammonium chloride to be measured from eye drops Sample dilution spectrum, draws the content of component dodecyl benzyl dimethyl ammonium chloride to be measured in eye drops dilution.
9. the method for quick of benzalkonium chloride in eye drops as claimed in claim 8, is characterized in that, further comprising the steps:
Step one, reduces step delta according to quantitative accuracy setting;
Step 2, at formula y
i=a
ix+b
iin bring larger x into
1value, obtains v
1;
Y
irepresent the absorbance of dodecyl benzyl dimethyl ammonium chloride under i wavelength, a
i, b
ibe constant, x represents the concentration of dodecyl benzyl dimethyl ammonium chloride, v
1represent in concentration to be x
1time dodecyl benzyl dimethyl ammonium chloride multi-wavelength absorbance, v
1for all y
ithe matrix of value composition;
Step 3, deducts v from eye drops dilution sample light modal data a
1/ Δ, the variable after deduction is designated as da; Background spectra database H
1' merge postscript for contrast space M with variable da, calculate and contrast space M and v
1angle;
Step 4, progressively deducts v from eye drops dilution sample light modal data a
1after, repeat step 3;
Step 5, after the dodecyl benzyl dimethyl ammonium chloride in eye drops dilution sample is deducted completely, the spectral vector v of comparison space M and dodecyl benzyl dimethyl ammonium chloride
1space angle value there will be maximal value θ
max, record space angle maximal value θ
maxcorresponding reduces step number λ, by the concentration x of dodecyl benzyl dimethyl ammonium chloride
1with the content Y reducing dodecyl benzyl dimethyl ammonium chloride in step number λ estimation eye drops dilution sample
1, calculating formula is Y
1=x
1/ Δ × λ, the Y obtained
1be the content value of dodecyl benzyl dimethyl ammonium chloride in eye drops dilution sample; If Y
1value and x
1value difference is comparatively large, then bring one and Y into
1be close and be greater than Y
1x
2recalculate;
Step 6, namely tries to achieve the content of dodecyl benzyl dimethyl ammonium chloride in actual eye drops sample according to extension rate.
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