CN1329005C - Thyroid disease auxiliary diagnosis instrument and its application - Google Patents
Thyroid disease auxiliary diagnosis instrument and its application Download PDFInfo
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- CN1329005C CN1329005C CNB2003101052715A CN200310105271A CN1329005C CN 1329005 C CN1329005 C CN 1329005C CN B2003101052715 A CNB2003101052715 A CN B2003101052715A CN 200310105271 A CN200310105271 A CN 200310105271A CN 1329005 C CN1329005 C CN 1329005C
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Abstract
The present invention relates to a disease diagnosis device, particularly to an auxiliary thyroid disease diagnosis instrument and an application thereof. The auxiliary thyroid disease diagnosis instrument mainly comprises a high-pressure pump A (1), a high-pressure pump B (2), a high-pressure pump C (3), an automatic sampler (4), a six-way valve (5), an octadecy l bonded silica gel column A (6), a mixer (7), an octadecy l bonded silica gel column B (8) and an ultraviolet-visible detector (9), wherein the position (6) of the six-way valve (5) is connected with the high-pressure pump A (1) through the automatic sampler (4); the positions (1) and (4) of the six-way valve (5) are respectively connected with both ends of the octadecy l bonded silica gel column A (6); the position (5) of the six-way valve (5) is connected with a waste liquid discharge pipeline; the position (3) of the six-way valve (5) is connected with an exit of the mixer (7); the high-pressure pump B (2) and the high-pressure pump C (3) are connected with an entrance of the mixer (7); the position (2) of the six-way valve (5) is connected with one end of the octadecy l bonded silica gel column A (8), and the other end of the octadecy l bonded silica gel column A (8) is connected with the waste liquid discharge pipeline through the ultraviolet-visible detector (9). The auxiliary thyroid disease diagnosis instrument has the advantages of simple method, high detection precision, low cost and good application effect.
Description
Technical field
The present invention relates to the medical diagnosis on disease instrument, specifically a kind of thyroid disease auxiliary diagnosis instrument and application thereof.
Background technology
Hyperthyroidism and hypothyroidism are modal two kinds of thyroid diseases, are respectively endocrinopathyes too much owing to intravital thyroid hormone secretion or that wretched insufficiency causes.Hyperthyroidism is comparatively common, and is more common in the women; Have report to claim that the hyperthyroidism prevalence accounts for 2% of female population, annual neopathy rate is 2/1000-3/1000 women's population, and the trend that increases is gradually arranged.Hyperthyroidism increases the body oxygen consumption, is to adapt to the prosperous excessively demand of metabolism on the one hand, has but caused increase [document 1.Videla LA, SirT, the WolffC of oxygen-derived free radicals in the body on the other hand; Free Radic Res Commun, 1988,5:1-10].No matter coenzyme Q10 is as requisite proton transmission medium in the cellular energy ATP generative process, still as the interior free yl scavenger, during hyperthyroidism, its content in body tissue all can significantly reduce [document 2.Krishnamurthy S, Prasanna D; Acta Vitaminol Enzymol, 1984,6:17-21; Document 3.Pandolfi C, Ferrari D, Stanic I, Pellegrini L; Minerva Endocrinol, 1994, Sep:19 (3), 139-42, Italian.; Document 4.Bianchi G, Solaroli E, Zaccheroni V, Grossi G, Bargossi A M, Melchionda N, Marchesini G; Horm Metab Res, 1999,31:620-6242-4].The situation of hypothyroidism just in time in contrast.Just because of the relation that has linear negative correlation between coenzyme Q10 and the thyroxin, therefore can be the index of coenzyme Q10 as thyroid hormone secretion, the secretion situation of coming thyroxin in the detection bodies by the content that detects coenzyme Q10, thereby these two kinds of common thyroid diseases of auxiliary diagnosis hyperthyroidism and hypothyroidism.Secondly, by the recovery that the changes of contents of Q10 in the dynamic monitoring patient body can be observed patient, monitor whole therapeutic process, thereby better therapeutic scheme is selected in the better medication of patient directions.On this point, the content that detects the blood plasma coenzyme Q10 has the important medical meaning.
Much report the mensuration of coenzyme Q10 is existing: photometry [document 5.Joanna K, Bozena M, JaninaP J; Journal of Pharmaceutical and Biomedical Analysis, 1998,17,1345-1350; Document 6.Readlieu E, Nilsson I M, Farley T M, Folkers K; AnalBiochem, 1968,23:132], chromatography [document 7.Wang Q, Lee B L, Ong C N; Journal ofChromatography B, 1999,726:297-302; Document 8.Zhiri A, Belichard P; Jourual ofLiquid Chromatograpy, 1994,17 (12), 2633-2640; Document 9.Kommuru T R, Khan MA, Ashraf M, Kattenacker R, Reddy I K; Journal of Pharmaceutical andBiomedical Analysis, 1998,16:1037-1040; Document 10.Grossi G, Bargossi AM, Fiorella P L, Piazzi S, Battino M, Bianchi G P; Jourual ofChromatograpy, 1992,593:217; Document 11.Finckh B, Kontush A, CommentzJ, Hubner Ch, Burdelski M, Kohlschutter A; Anal Biochem, 1995,232:210; Document 12.Kaplan P, Sebastianova N, Turiakova J, Kucera I; Physiol Res, 1996,45:39; Document 13.Vadhanavikit S, Sakamoto N, Ashida N, Kishi T, Folkers K; AnalBiochem, 1984,142:155-158], voltammetry [document 14.Simona C L, Iulia G D, GabrielL R, Hassan Y.A E; Instrumentation science﹠amp; Technology, 2001,29 (2), 109-116], chemoluminescence method [document 15.Battino M, Ferri E, Girotti S, Lenaz G; AnalyticaChimica Acta, 199 1,255:367-371], fluorescence method [document 16.Rokos J A S; AnalBiochem, 1973,56:26], electron magnetic resonance method [document 17.Long Y T, Yu Z H, Chen H Y; Electrochem Commun.1999,1,194] or the like, wherein the most ripe the most frequently used still liquid chromatography.Plasma sample matrix complexity usually needs before the analysis through loaded down with trivial details pretreatment [document 9,10,12,13].People such as Vadhanavikit [document 13] are with the TLC extraction and purify blood sample, and this method precision is relatively poor, and the response rate also has much room for improvement.SPE collection purification and be enriched in one is applicable to the pretreatment of complex sample.Kommuru[document 9] silica gel SPE post is introduced the purge process of plasma sample, removed the oil-soluble impurities of some low poles in the extract well, but the existence of strong polar impurity makes that the solvent front is still very high in the chromatogram.Document [document 10,12] utilizes the SPE post (silicagel column and C18 post) of two kinds of opposed polarities to remove low pole and strong polar impurity in the blood plasma successively, and blood sample is purified significantly, and it is clean that this method goes out the peak, protects effectively and has prolonged column life; But two step Solid-Phase Extraction waste time and energy, and cost is higher, is unsuitable for analyzing in enormous quantities clinically.
Summary of the invention
The purpose of this invention is to provide a kind of thyroid disease auxiliary diagnosis instrument and application thereof, it integrates sample treatment and the higher analytical system of analyzing of automaticity, human body blood plasma coenzyme Q10 content accurately and efficiently is for hyperthyroidism and hypothyroidism patient's diagnosis and treatment provides useful guide.
For achieving the above object, the present invention has utilized column switching technique, set up the liquid-phase chromatographic analysis system that a cover two posts link to each other, this system comprises following assembly: three high-pressure pumps, an automatic sampler, a ultraviolet-visible detector, a system controller, a chromatographic work station, six-way valve, chromatography post A and a chromatography post B; Mainly form by high-pressure pump A (1), high-pressure pump B (2), high-pressure pump C (3), automatic sampler (4), six-way valve (5), octadecyl silane post A (6), blender (7), octadecyl silane post B (8) and ultraviolet-visible detector (9); 6. the position of six-way valve (5) is connected with high-pressure pump A (1) by automatic sampler (4), 1. the position of six-way valve (5) and 4. the position connect affine octadecyl silane post A (6) two ends respectively, 5. the position of six-way valve (5) connects the waste liquid discharge line, 3. the position of six-way valve (5) links to each other with blender (7) outlet, high-pressure pump B (2) links to each other with blender (7) inlet with high-pressure pump C (3), 2. the position of six-way valve (5) links to each other with octadecyl silane post B (8) one ends, and the other end of octadecyl silane post B (8) links to each other with the waste liquid discharge line by ultraviolet-visible detector (9).
The application of thyroid disease auxiliary diagnosis instrument, it is by the detection auxiliary diagnosis thyroid disease to coenzyme Q10 content height in the blood plasma, and concrete steps are as follows,
1) gets blood plasma and add the normal propyl alcohol centrifugal degreasing, the sample analysis is directly gone up in the supernatant except that behind the albumen;
2) analytic sample is directly gone up sample by automatic sampler, six-way valve is in " LOAD " position, selects pure methanol drip washing octadecyl silane post A for use, makes that coenzyme Q10 is retained on the post A in the blood plasma;
3) continue with pure washed with methanol post A, some remaining in blood plasma impurity flow into the waste liquid discharge line through post A;
4) six-way valve is in " INJECT " position and makes mobile phase with methanol and isopropyl alcohol, and the coenzyme Q10 component elutes from post A in the blood plasma, arrives the top of octadecyl silane post B through six-way valve, finishes online transmission overall process between post A and post B;
5) six-way valve comes back to " LOAD " position, carries out degree drip washing programs such as methanol and isopropyl alcohol on post B, reaches the purpose of separation determination coenzyme Q10;
6) just can measure accurately according to above-mentioned steps human plasma coenzyme Q10 content, by monitoring to Q10 content in normal person, hyperthyroidism and three groups of people colonies of hypothyroidism, can grasp the approximate horizontal of human plasma content, and by relatively having realized whether suffering from the auxiliary diagnosis accurate diagnosis of hyperthyroidism or hypothyroidism (thereby can assist) of thyroid disease with the normal person; , content is hypothyroidism probably if being significantly higher than normal person's meansigma methods; As significantly being lower than normal person's meansigma methods, then be hyperthyroidism probably; Otherwise then be normal.
Similar above-mentioned principle also is applicable to the judgement of thyroid disease therapeutic effect; According to step 1)～5) to pass through the dynamic monitoring of human plasma coenzyme Q10 content, grasp patient's that can be definite recovery is for selecting and determining that new therapeutic scheme provides auxiliary foundation.
Described post A is a pretreatment column, length 10～30mm, internal diameter 4～4.6mm; Post B is an analytical column, length 100～250mm, internal diameter 4～4.6mm; The flow of pure methanol is 0.3～1ml/min, and the volume ratio of described methanol and isopropyl alcohol is 15: 1～1: 1.
The present invention has following advantage:
1. method is simple.The present invention has simplified the extraction process of coenzyme Q10, select for use normal propyl alcohol can the defat Deproteinization again can the quantified extract coenzyme Q10, avoided with usual employing earlier with the complicated processes of reuse n-hexane extraction after the pure defat.Blood plasma matrix complicated component, if directly the extract sample introduction is analyzed without any processing, often have following problem: solvent front height, impurity particle easily stop up pillar, keep material by force column life is reduced greatly, in addition, impurity often disturbs the mensuration of Q10.Document adopts TLC technology [document 13] or SPE technology [document 9, document 10, document 12] to purify and the enrichment blood sample more.These method complex operations are wasted time and energy, and it is relatively poor to measure precision.Sample pretreatment process of the present invention only needs to slough plasma lipoprotein with normal propyl alcohol, just can directly analyze after the centrifugalize; Adopt liquid chromatograph automatic on-line analytical system to be used for the mensuration of blood plasma coenzyme Q10, automatic pretreatment, auto injection and automatic several big functions of compartment analysis have been realized from sample, simplified The whole analytical process greatly, be suitable for especially analyzing in enormous quantities clinically; Because this method is entirely by computer controlled automatic, guaranteed the in full accord of condition between sample determination, therefore can be directly accurately quantitative with external standard method, removed from seek suitable in the target trouble.
2. accuracy of detection height.The present invention utilizes column switching technique, develop a kind of novel automatic on-line system, this system's compartment analysis blood plasma coenzyme Q10 has been realized the automatization from sample pretreatment to the compartment analysis overall process, avoided the sample pretreatment step that wastes time and energy, also avoid simultaneously the manual operation error that in the sample pretreatment process, produces, improved analytical accuracy and degree of accuracy greatly; The all operations process is by a system controller, and by computer control, automaticity is higher, is suitable for clinical mass detection; In addition, this system has avoided the personal error brought in the experimentation effectively, has guaranteed the accuracy and the degree of accuracy (it is higher to measure precision, and in a few days RSD is 0.4%, and RSD is 3% in the daytime) of the process of measuring.The contents level of the human normal plasma's coenzyme Q10 that records with this method and hyperthyroidism patient's contents level have significant difference (P＜0.05), thereby can assist the diagnosis of hyperthyroidism and hypothyroidism case history clinically.
3. cost is low.This system adopts the automatic on-line device to purify blood sample, and be guaranteed the service life of analytical column, and simultaneously, this system has avoided purifying blood sample with solid phase extraction column, and cost reduces greatly.
4. effect is good.The present invention is based on multidimensional liquid chromatograph and coenzyme Q10, coenzyme Q10 content as hyperthyroidism, the clinical criterion index of hypothyroidism case, can have been remedied the deficiency that FT3 and FT4 make criterion; Thyroid disease auxiliary diagnosis instrument of the present invention, by specific buffer and analytical method, content that can the automatic accurate blood plasma of human body efficiently coenzyme Q10, thereby the diagnosis of auxiliary effectively hyperthyroidism and two kinds of common thyroid diseases of hypothyroidism; By the dynamic monitoring of blood plasma coenzyme Q10 in the patient body, can grasp patient's recovery early, for selecting and determining that new therapeutic scheme provides reliable foundation.
Description of drawings
Fig. 1 is the schematic flow sheet of auxiliary diagnosis instrument of the present invention.
Fig. 2 is in " LOAD " switching position sketch map for six-way valve in the auxiliary diagnosis instrument of the present invention.
Fig. 3 is in " INJECT " switching position sketch map for six-way valve in the auxiliary diagnosis instrument of the present invention.
Fig. 4 is that the liquid chromatograph on-line system of blood plasma coenzyme Q10 separates spectrogram.Peak 1. coenzyme Q10s. operating condition: chromatography post 1 (4.6mm i.d * 20mm), chromatography post 2 (4.6mm i.d * 150mm), detect wavelength: 275nm, sample size: 400ul, all the other conditions see Table 1.
The specific embodiment
1. instrument of the present invention can realize that automatic pretreatment, auto injection and coenzyme Q10 component automatic of blood sample separates and quantitatively; It is mainly finished as follows;
1) gets the fresh plasma sample of volume 0.3ml, after normal propyl alcohol (both volume ratios can be 0.2: 1 to 0.4: the 1 all can) centrifugal degreasing that adds 1ml removes albumen, the sample analysis is directly gone up in the supernatant;
2) be in " LOAD " position (Fig. 2) when six-way valve, with the analytic sample direct injected, select pure methanol drip washing octadecyl silane post A for use, make that coenzyme Q10 is retained on the post A in the blood plasma by automatic sampler;
3) continue with pure washed with methanol post A, in the blood plasma remaining some impurity through in post A inflow waste liquid discharge line waste liquid bottles extremely;
4) when six-way valve is in " INJECT " position (Fig. 3), make mobile phase with methanol and isopropyl alcohol, the coenzyme Q10 component elutes from post A in the blood plasma, through the top that the 3-4-1-2 of six-way valve arrives octadecyl silane post B, finishes online transmission overall process between post A and post B;
5) six-way valve comes back to " LOAD " position, carries out degree drip washing programs such as methanol and isopropyl alcohol on post B, reaches the purpose of separation determination coenzyme Q10; Before carrying out the next round blood sample analysis, reactivate chromatography post 1 with pure methanol.
6) just can measure accurately according to above-mentioned steps human plasma coenzyme Q10 content, by monitoring to Q10 content in normal person, hyperthyroidism and three groups of people colonies of hypothyroidism, the approximate horizontal of human plasma content can be grasped, thereby the accurate diagnosis of hyperthyroidism or hypothyroidism can be assisted; By dynamic monitoring, grasp patient's that can be definite recovery is for selecting and determining that new therapeutic scheme provides reliable foundation.
2. on the auxiliary diagnosis instrument that the present invention makes up, adopt table 1 (wherein, mobile phase A is pure methanol solution, and Mobile phase B is methanol and 9: 1 mixed liquor of isopropyl alcohol volume ratio) and analysis condition shown in Figure 4, the coenzyme Q10 component in the plasma sample is carried out separation determination (Fig. 4).
Table 1 valve switching time and flow rate of mobile phase change
| Time t (min) | 0-4 | 4-6.5 | ＞6.5 |
| Mobile phase A (ml/min) | 0.5 | 0 | 0.5 |
| Mobile phase B (ml/min) | 1.5 | 1.5 | 1.5 |
Coenzyme Q10 among 6 hypothyroidism patients, 39 hyperthyroidism patients and 76 human normal plasmas is measured, the intravital coenzyme Q10 content of normal person is 0.67 ± 0.25mg/l (mean ± SD), the hyperthyroidism disease people is that (mean ± SD), hypothyroidism are 0.89 ± 0.17mg/l (mean ± SD) to 0.42 ± 0.17mg/l.Hyperthyroidism patient and human normal plasma's coenzyme Q10 content are carried out statistical disposition, the two contain measurer significant difference (P＜0.05).
3. contrast: thyroxin in vivo the metabolism time longer, so it can not be as the sensitive index of hyperthyroidism disease.The hyperthyroidism symptom has appearred in some patient, FT3 in the body, and FT4 value still keeps normally, such as: a routine patient is arranged, occurred the hyperthyroidism symptom, through medical science detection FT3=6.1, FT4=22, TSH=0.596 singly organizes data from this and can't conclude his whether hyperthyroidism.By detecting blood plasma coenzyme Q10 content, its value is 0.33mg/l, is starkly lower than the normal person, thereby can assist and be diagnosed as hyperthyroidism.
4. the content that detects the blood plasma coenzyme Q10 can be understood patient's recovery as soon as possible.One routine patient is arranged, its when hyperthyroidism and the detection case of treatment after a period of time see Table 2, from this patient's FT3, FT4, TSH is difficult to judge whether he recovers, but just has been perfectly clear from the changes of contents of coenzyme Q10, there has not been any discomfort symptom when in fact, this patient checks.
Physiological data before and after table 2 patient's hyperthyroidism relatively
Claims (1)
1. a thyroid disease auxiliary diagnosis instrument is characterized in that: mainly be made up of high-pressure pump A (1), high-pressure pump B (2), high-pressure pump C (3), automatic sampler (4), six-way valve (5), octadecyl silane post A (6), blender (7), octadecyl silane post B (8) and ultraviolet-visible detector (9); 6. the position of six-way valve (5) is connected with high-pressure pump A (1) by automatic sampler (4), 1. the position of six-way valve (5) and 4. the position connect affine octadecyl silane post A (6) two ends respectively, 5. the position of six-way valve (5) connects the waste liquid discharge line, 3. the position of six-way valve (5) links to each other with blender (7) outlet, high-pressure pump B (2) links to each other with blender (7) inlet with high-pressure pump C (3), 2. the position of six-way valve (5) links to each other with octadecyl silane post B (8) one ends, and the other end of octadecyl silane post B (8) links to each other with the waste liquid discharge line by ultraviolet-visible detector (9).
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| CNB2003101052715A CN1329005C (en) | 2003-12-05 | 2003-12-05 | Thyroid disease auxiliary diagnosis instrument and its application |
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| CNB2003101052715A CN1329005C (en) | 2003-12-05 | 2003-12-05 | Thyroid disease auxiliary diagnosis instrument and its application |
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| CN1329005C true CN1329005C (en) | 2007-08-01 |
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN2063400U (en) * | 1989-09-20 | 1990-10-10 | 中国人民解放军第一四八医院 | Hyper thyroidism index diagnosing instrument |
| CN1065529A (en) * | 1991-04-01 | 1992-10-21 | 山东省化学研究所 | The method of capillary supercritical fluid chromatography desolventizing and desolventizing split sampling system thereof |
| CN2479915Y (en) * | 2001-07-03 | 2002-03-06 | 陈启 | Tyroid size measurer |
| CN2541844Y (en) * | 2002-06-03 | 2003-03-26 | 复旦大学 | Heat expansion microflow high pressure gradienjt pump |
-
2003
- 2003-12-05 CN CNB2003101052715A patent/CN1329005C/en not_active Expired - Fee Related
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN2063400U (en) * | 1989-09-20 | 1990-10-10 | 中国人民解放军第一四八医院 | Hyper thyroidism index diagnosing instrument |
| CN1065529A (en) * | 1991-04-01 | 1992-10-21 | 山东省化学研究所 | The method of capillary supercritical fluid chromatography desolventizing and desolventizing split sampling system thereof |
| CN2479915Y (en) * | 2001-07-03 | 2002-03-06 | 陈启 | Tyroid size measurer |
| CN2541844Y (en) * | 2002-06-03 | 2003-03-26 | 复旦大学 | Heat expansion microflow high pressure gradienjt pump |
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