CN102692396B - Endotoxin detection system and quantitative detection method thereof - Google Patents
Endotoxin detection system and quantitative detection method thereof Download PDFInfo
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- CN102692396B CN102692396B CN201210196596.8A CN201210196596A CN102692396B CN 102692396 B CN102692396 B CN 102692396B CN 201210196596 A CN201210196596 A CN 201210196596A CN 102692396 B CN102692396 B CN 102692396B
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- 238000001514 detection method Methods 0.000 title claims abstract description 63
- 239000002158 endotoxin Substances 0.000 title claims abstract description 56
- 239000007788 liquid Substances 0.000 claims abstract description 17
- 239000002245 particle Substances 0.000 claims abstract description 15
- 201000009310 astigmatism Diseases 0.000 claims abstract description 6
- 239000003814 drug Substances 0.000 claims abstract description 4
- 238000012545 processing Methods 0.000 claims abstract description 4
- 238000000034 method Methods 0.000 claims description 14
- 239000000243 solution Substances 0.000 claims description 14
- 238000002347 injection Methods 0.000 claims description 10
- 239000007924 injection Substances 0.000 claims description 10
- 238000007689 inspection Methods 0.000 claims description 8
- 238000004519 manufacturing process Methods 0.000 claims description 8
- 238000000149 argon plasma sintering Methods 0.000 claims description 7
- 239000000835 fiber Substances 0.000 claims description 4
- 239000000523 sample Substances 0.000 claims description 4
- 229940079593 drug Drugs 0.000 claims description 3
- 238000001228 spectrum Methods 0.000 claims description 3
- 230000008878 coupling Effects 0.000 claims description 2
- 238000010168 coupling process Methods 0.000 claims description 2
- 238000005859 coupling reaction Methods 0.000 claims description 2
- 238000001914 filtration Methods 0.000 claims description 2
- 239000005304 optical glass Substances 0.000 claims description 2
- 238000012544 monitoring process Methods 0.000 abstract description 9
- 230000035945 sensitivity Effects 0.000 abstract description 5
- 238000006243 chemical reaction Methods 0.000 abstract description 2
- 230000001960 triggered effect Effects 0.000 abstract description 2
- 239000003795 chemical substances by application Substances 0.000 abstract 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 6
- 231100000284 endotoxic Toxicity 0.000 description 5
- 230000002346 endotoxic effect Effects 0.000 description 5
- 240000007164 Salvia officinalis Species 0.000 description 4
- 235000005412 red sage Nutrition 0.000 description 4
- 241000239222 Tachypleus Species 0.000 description 3
- 239000006166 lysate Substances 0.000 description 3
- 238000004848 nephelometry Methods 0.000 description 3
- 239000012086 standard solution Substances 0.000 description 3
- 239000008215 water for injection Substances 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 238000005259 measurement Methods 0.000 description 2
- 239000012528 membrane Substances 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 238000000108 ultra-filtration Methods 0.000 description 2
- 206010067484 Adverse reaction Diseases 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 241000239218 Limulus Species 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 239000008186 active pharmaceutical agent Substances 0.000 description 1
- 230000006838 adverse reaction Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 239000006185 dispersion Substances 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
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- 230000007613 environmental effect Effects 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 230000000977 initiatory effect Effects 0.000 description 1
- 229920006008 lipopolysaccharide Polymers 0.000 description 1
- 239000002502 liposome Substances 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 239000002510 pyrogen Substances 0.000 description 1
- 238000011155 quantitative monitoring Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 239000002699 waste material Substances 0.000 description 1
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Abstract
The invention discloses an endotoxin detection system and a quantitative detection method thereof and belongs to the field of endotoxin monitoring. By the system, a light path detection system is integrated on the basis of the conventional laser particle size detector, by double-laser source irradiation, output light enters a detection pool through a light path, is received by a multi-point scattered light receiver, is subjected to photoelectric conversion and signal amplifying processing and enters a quantitative arithmetic unit; and the quantitative arithmetic unit fits the relativity between the astigmatism and the endotoxin concentration according to the scattering intensity difference of the endotoxin particles at three or more different angles, the endotoxin concentration is calculated and displayed, and when the endotoxin concentration exceeds a limit value, an early warning system is triggered, so that the online monitoring is realized. By the system, the quantitative detection of the endotoxin can be realized, the system is high in detection sensitivity, short in detection time and high in reliability, an agent is not consumed, and pollution to the liquid medicine is avoided.
Description
Technical field
The present invention relates to a kind of endotoxin detection technique, relate in particular to a kind of endotoxin detection system and quantitative detecting method thereof detecting based on laser particle, belong to endotoxin monitoring field.
Background technology
Bacterial endotoxin is lipopolysaccharides, is also referred to as liposome, is the composition of Gram-negative bacteria mantle, is extensively present in occurring in nature, and this material enters blood of human body can cause heating, is commonly called as pyrogen reaction.Because this type of material may cause people, produce serious adverse reaction, therefore in drug injection, all need strict control.
At present, water for injection and injection production run also cannot be carried out on-line monitoring by induced by endotoxin, and endotoxic detection also depends on to produce and finishes rear employing limulus reagent test or rabbit method checks.The sensitivity that these methods and process not only detect is low, and once find that the endotoxin in product exceeds standard, entire block will be with waste disposal, this not only Hui Gei enterprise bring economic loss, Ye Huigei enterprise brings huge clinical practice risk simultaneously.Therefore, in order to ensure water for injection and injection quality safety, prevent the burst pollution in production run, the integrality of ultra filtration membrane in monitoring production, the on-line monitoring that realizes induced by endotoxin is significant.
Laser particle detector is not yet applied to endotoxic detection at present, we analyze the higher Ma Erwen Nano ZSZEW3600 type laser diffraction analyzer of sensitivity, it can detect particle in the solution of endotoxin concns more than 10EU/ml, and this cannot meet endotoxic detectable concentration in injection.Laser particle detector is different at the astigmatic signal of different angles according to particle, 2 above multiple spot astigmatism signals is carried out to granularmetric analysis, but can't detect particle concentration, therefore cannot be for endotoxic quantitative detection.
Summary of the invention
The present invention is directed to the defect that prior art exists, and propose endotoxin detection system and the quantitative detecting method thereof that a kind of detection sensitivity is high, can realize online detection.
This endotoxin detection system comprises lasing light emitter, short focus lens, grating, long focus lens, detection cell, scattered light receiver, photoelectric commutator, signal amplifier, signal processor, data acquisition unit, quantitative arithmetical unit, data display equipment and prior-warning device, wherein: the output light of lasing light emitter is successively by short focus lens, after grating and long focus lens, enter detection cell, the light that appears of detection cell receives the laggard photoelectric commutator that enters by scattered light receiver, the output signal of photoelectric commutator is passed through signal amplifier successively, after signal processor and data acquisition unit, enter quantitative arithmetical unit and carry out endotoxin concns calculating, output connection data display and the prior-warning device of quantitative arithmetical unit.
The quantitative detecting method of this endotoxin detection system is: the scattered light signal intensity of endotoxin particle in more than three different angles detection solution, according to light scattering formula I=KCf (D) f (E), by the scattered light signal of any two angles wherein, calculate f (D) and f (E), by light scattering simplification of a formula, be S=fC+g, with the scattered light signal of all the other angles, calculate again the endotoxin concns of solution, realize quantitatively and detecting;
In above formula: I is scattered light signal intensity; K is proportionality constant; C is endotoxin concns; F (D) is particle diameter parametric function; F (E) is scattering spectrum function; S is astigmatism, and S is that the ln linearization data processing by I obtains; Related coefficient f=Kf (D) f (E); G is corrected parameter.
Technique effect:
1, detect and not consume reagent, to liquid without any pollution, economic environmental protection.
2, light path system has been carried out integrated, improved detection sensitivity, can detect endotoxin solution more than 0.5EU/ml.
3, combine the quantitative operational method to 3 above multiple spot astigmatism signals, realized endotoxic quantitative detection.
4, detecting pattern is divided into two kinds of online detection and offline inspection, and applying flexible, practicality is good, detection time is short, can meet the on-line monitoring requirement of water for injection, injection and integrity of ultrafiltration membrane.
5, set up early warning system, in the time of can exceeding standard at the endotoxin concns of solution, triggered early warning, realized on-line monitoring, reliability is high.
Accompanying drawing explanation
Fig. 1 is the composition structural representation of system of the present invention, number in the figure title: 1, lasing light emitter; 2, short focus lens; 3, grating; 4, long focus lens; 5, detection cell; 6, scattered light receiver; 7, photoelectric commutator; 8, signal amplifier; 9, signal processor; 10, data acquisition unit; 11, quantitative arithmetical unit; 12, data display equipment; 13, prior-warning device.
Fig. 2 is quantitative measurement of endotoxin Computing Principle schematic diagram.
Fig. 3 is the detection early warning process flow diagram of system of the present invention.
Embodiment
Below the invention will be further described.
The structure of endotoxin detection system of the present invention as shown in Figure 1, comprise lasing light emitter 1, short focus lens 2, grating 3, long focus lens 4, detection cell 5, scattered light receiver 6, photoelectric commutator 7, signal amplifier 8, signal processor 9, data acquisition unit 10, quantitative arithmetical unit 11, data display equipment 12 and prior-warning device 13, wherein: the output light of lasing light emitter 1 is successively by short focus lens 2, after grating 3 and long focus lens 4, enter detection cell 5, the light that appears of detection cell 5 receives the laggard photoelectric commutator 7 that enters by scattered light receiver 6, the output signal of photoelectric commutator 7 is successively by signal amplifier 8, after signal processor 9 and data acquisition unit 10, enter quantitative arithmetical unit 11 and carry out endotoxin concns calculating, output connection data display 12 and the prior-warning device 13 of quantitative arithmetical unit 11.
The present invention carries out integrated on the basis of conventional laser particle diameter detector to light path system, the scattering light path that is preferably 90 ° of left and right detects.Lasing light emitter 1 is double light beam laser source, adopts the FLS-2600B lasing light emitter of Canadian EXFO company; Short focus lens 2 adopts Extra-low Dispersion lens; Grating 3 adopts coupling fiber type fiber grating; Long focus lens 4 adopts low consumption NV-202m lens.Detection cell 5 is made by optical glass, comprise two kinds, online detection cell and offline inspection pond: the two ends up and down of online detection cell are provided with liquid outlet and inlet, online detection cell is connected in injection production line pipeline by liquid outlet and inlet and forms loop, to realize the continuous detecting of liquid, on liquid outlet and inlet, be equipped with electromagnetic valve, at inlet place, be also provided with ultrasonic probe, for getting rid of bubble, disturb; Offline inspection pond is the detachable detection cell of semitight, and this detection cell top is provided with filling opening, for manually changing solution.Scattered light receiver 6 adopts the Picometrix LLC high speed multiple spot wide angle scattered light receiver module of API company, is multi-point and arranges every logarithm; Photoelectric commutator 7 adopts the HESMC photoelectric commutator of U.S. Heng Qi Electronics Co., Ltd.; Signal amplifier 8 is comprised of amplifying circuit and filtering circuit, and wherein amplifying circuit is connected into differential amplifier circuit form by three ICL7650 amplifiers, and wave filter is conventional RC network; Signal processor 9 adopts the CONVERTER-VLC.602 programmable signal processor of Italian VAL.CO; Data acquisition unit 10 adopts the A/D converter Max132 of Maxim.Quantitatively arithmetical unit 11Wei our unit independent research, quantitatively the Computing Principle of arithmetical unit as shown in Figure 2, according to light scattering formula I=KbCf (D) f (E) f (Δ λ), in formula: I is scattered light signal intensity, K is proportionality constant, b is sample path length to be measured, C is endotoxin concns (unit: EU/ml), f (D) is particle diameter parametric function, f (E) is scattering spectrum function, f (Δ λ) is function of wavelength, sample path length to be measured (colorimetric pool) is fixing, wavelength is fixed, so b and f (Δ λ) are constant, light scattering formula can be reduced to I=KCf (D) f (E).With the endotoxin standard solution detection of variable concentrations, can draw constant K value, the scattered light signal intensity of system endotoxin particle in more than three different angles detection solution, according to I=KCf (D) f (E), by the scattered light signal of any two angles wherein, calculate f (D) and f (E), by light scattering simplified formula, be S=fC+g, in formula: S is astigmatism, S is that the ln linearization data processing by I obtains, related coefficient f=Kf (D) f (E), g is corrected parameter, with the scattered light signal of all the other angles, calculate endotoxin content in solution again, realize quantitatively and detecting.Data display equipment 12 adopts 128 * 64 dot matrix display terminals of the golden company of initiating, for showing detected endotoxin concns value; Prior-warning device 13 adopts integrated achievement electronics CMS7000-500 alarm monitoring software and piezoelectric type early warning system, the detection early warning flow process of system as shown in Figure 3, in early warning software, can preset endotoxin concns limit value, when in the liquid detecting, endotoxin concns surpasses limit value, attention device is reported to the police.
Use flow process when the present invention detects is online: according to need of production, quantitatively in arithmetical unit, presetting endotoxin concns limit value and coefficient f and parameter g, open production line, make solution enter online detection cell, according to S=fC+g, calculate endotoxin concns, when this concentration is less than limit value, system keeps operation, when this concentration during higher than limit value early warning system report to the police.
Operation steps during offline inspection of the present invention is: quantitatively in arithmetical unit, presetting coefficient f and parameter g, take out offline inspection pond, with pure water, clean up, with solution to be measured, swing and wash after 2～3 times, add solution to be measured, detection cell is placed in to native system, according to S=fC+g, calculates endotoxin concns.
The quantitative detection example of endotoxin off-line of red sage roo drip liquid is provided below:
Change offline inspection pond, get working stamndard endotoxin, use and check that water is mixed with the standard solution of concentration 100EU/ml, re-use and check that water progressively dilutes the standard solution for series concentration, the series concentration being mixed with is respectively 0.5,1,5,10,50EU/ml, solution is placed in to detection cell and detects, and by quantitative arithmetical unit, carries out data regression calculating, linear equation is: S=0.056C+0.165, R
2=0.997, result of calculation is in Table 1.
Table 1
(Shanghai Worldbest & Anhui Jinhui Pharmaceutical Co., Ltd. provides, lot number: 05110209), adopt respectively native system detection method and tachypleus amebocyte lysate nephelometry to calculate endotoxin content, the results are shown in Table 2 to get red sage roo drip liquid intermediate.
Table 2
Result demonstration, the endotoxin content that native system records and tachypleus amebocyte lysate nephelometry are comparatively approaching.Tachypleus amebocyte lysate nephelometry is for Chinese Pharmacopoeia records method at present, and measurement result is comparatively accurate, and the result that native system records is close with it, and this has also illustrated the reliability of native system.
The online quantitatively detection example of endotoxin of red sage roo drip liquid is provided below:
Get two parts of red sage roo drip liquids, a for normal liquid, (Shanghai Worldbest & Anhui Jinhui Pharmaceutical Co., Ltd. provides, lot number: 10011605), another part, for the liquid after polluting, carries out respectively on-line monitoring to two parts of liquids.Detected parameters is set: f is that 0.056, g is 0.165, and endotoxin limit value is 1EU/mL, opens production line and detect, by regression equation S=0.056C+0.165, calculate endotoxin concns, the results are shown in Table 3.
Table 3
Result shows, when contaminated with endotoxins surpasses limit value, native system can be reported to the police, and can meet the quantitative monitoring of the contaminated with endotoxins situation of injection in producing.
Claims (2)
1. for an endotoxin detection system for drug injection, it is characterized in that: comprise lasing light emitter (1), short focus lens (2), grating (3), long focus lens (4), detection cell (5), scattered light receiver (6), photoelectric commutator (7), signal amplifier (8), signal processor (9), data acquisition unit (10), quantitative arithmetical unit (11), data display equipment (12) and prior-warning device (13), wherein: the output light of lasing light emitter (1) is successively by short focus lens (2), after grating (3) and long focus lens (4), enter detection cell (5), the light that appears of detection cell (5) receives the laggard photoelectric commutator (7) that enters by scattered light receiver (6), and the output signal of photoelectric commutator (7) is successively by signal amplifier (8), after signal processor (9) and data acquisition unit (10), enter quantitative arithmetical unit (11) and carry out endotoxin concns calculating, quantitatively output connection data display (12) and the prior-warning device (13) of arithmetical unit (11), described lasing light emitter (1) adopts double light beam laser source, described detection cell (5) is made by optical glass, and detection cell (5) comprises online detection cell and offline inspection pond, described online detection cell is provided with liquid outlet and inlet, online detection cell is connected in injection production line pipeline by this liquid outlet and inlet, on described liquid outlet and inlet, be equipped with electromagnetic valve, at inlet place, be also provided with the ultrasonic probe disturbing for getting rid of bubble, described offline inspection pond is the detachable detection cell of semitight, and filling opening is located at detection cell top, described scattered light receiver (6) adopts 3 above multiple spot wide angle scattered light receivers, described grating (3) is coupling fiber type fiber grating, described signal amplifier (8) is comprised of amplifying circuit and filtering circuit, described endotoxin detection system is used the scattering light path of 90 ° to detect.
2. the quantitative detecting method based on the endotoxin detection system for drug injection claimed in claim 1, is characterized in that:
The method comprises the following steps: the scattered light signal intensity of endotoxin particle in more than three different angles detection solution, according to light scattering formula I=KCf (D) f (E), by the scattered light signal of any two angles wherein, calculate f (D) and f (E), by light scattering simplification of a formula, be S=fC+g, with the scattered light signal of all the other angles, calculate again the endotoxin concns of solution, realize quantitatively and detecting;
In above formula: I is scattered light signal intensity; K is proportionality constant; C is endotoxin concns; F (D) is particle diameter parametric function; F (E) is scattering spectrum function; S is astigmatism, and S is that the ln linearization data processing by I obtains; Related coefficient f=Kf (D) f (E); G is corrected parameter.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201210196596.8A CN102692396B (en) | 2012-06-14 | 2012-06-14 | Endotoxin detection system and quantitative detection method thereof |
| PCT/CN2013/077191 WO2013185621A1 (en) | 2012-06-14 | 2013-06-13 | Endotoxin detection systems and detection methods thereof |
| US14/408,016 US9958431B2 (en) | 2012-05-14 | 2013-06-13 | Endotoxin detection systems and detection methods thereof |
| EP13803561.3A EP2863206A4 (en) | 2012-06-14 | 2013-06-13 | Endotoxin detection systems and detection methods thereof |
| JP2015516431A JP6093854B2 (en) | 2012-06-14 | 2013-06-13 | Detection method based on endotoxin detection system |
| US15/877,627 US20180224426A1 (en) | 2012-06-14 | 2018-01-23 | Endotoxin detection systems and detection methods thereof |
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| CN103278476B (en) * | 2013-06-08 | 2015-06-03 | 南京拓鉒医药科技有限公司 | Endotoxin detection system and limit detection method thereof |
| US9958431B2 (en) * | 2012-05-14 | 2018-05-01 | Nanjing Tuozhu Pharmaceutical & Tech Co., Ltd. | Endotoxin detection systems and detection methods thereof |
| WO2015085066A1 (en) * | 2013-12-06 | 2015-06-11 | BacterioScan Inc. | Optical measurement cuvette having sample chambers |
| CN112504922B (en) * | 2020-10-20 | 2022-09-02 | 华南师范大学 | Online measurement system and method for particle size distribution of atmospheric particulates |
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| US4188121A (en) * | 1977-02-01 | 1980-02-12 | Hirleman Edwin D Jr | Multiple ratio single particle counter |
| CN101520414A (en) * | 2008-02-29 | 2009-09-02 | 怀亚特技术公司 | Method for determining average properties of molecules in solution by injection into a flowing solvent |
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