Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving glucose or galactose
Patent
1993-09-23
1997-11-04
Gitomer, Ralph
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving glucose or galactose
435 4, C12Q 154, C12Q 100
Patent
active
056838891
DESCRIPTION:
BRIEF SUMMARY
TECHNICAL FIELD
The present invention relates to a reagent for measurement of the content of magnesium ions in various samples, which is useful for the fields of medical sciences, food analysis and environmental preservation.
BACKGROUND ART
The measurement of magnesium ions in biological fluids (e.g. blood, etc.), foods, the environment, such as rivers, and the like has widely been made, and is considered to be important. Methods for measuring magnesium ions are classified into a an instrumental analysis method (e.g. atomic absorption spectrophotometry, etc.) and by a chemical method (e.g. xylidyl blue method, etc.). According to the former method, comparatively accurate results can be obtained. However, expensive instruments are especially required and operation is complicated, whereby, great skill is required. Further, the latter method is a simple method, however, it has a problem that results including large error factors are obtained because it lacks specificity to magnesium ions and the influence of coexisting substances in the sample can not be avoided.
In order to solve these problems, there has been proposed various reagents for the measurement of magnesium ions which utilize an enzyme reaction. These reagents are those which utilize the fact that various kinases enzymes such as hexokinase (hereinafter abbreviated as HK) act on a complex of a magnesium ion with adenosine 5'-triphosphate (hereinafter abbreviated as ATP) as a substrate to manifest their activities. Examples thereof include reagents utilizing a coupling reaction of HK with glucose-6-phosphate dehydrogenase (hereinafter abbreviated as G6PDH) Pat. No. 4,657,854!; reagents utilizing a coupling reaction of glucokinase (hereinafter abbreviated as GlcK) having high specificity to glucose, the 25th annual meeting of Japan Society of Clinical Chemistry, page 75, 1985: U.S. Pat. No. 4,778,754!; reagents utilizing a coupling reaction of three enzymes, glycerol kinase (hereinafter abbreviated as Glyk), glycerol-3-phosphate dehydrogenase (hereinafter abbreviated as G3PDH) and pages 629-632, 1986! and the like. In WO88/08137, there is described an adenylate kinase or HK which is sensitive to a magnesium ion which can be used, however, there is no disclosure about the accuracy of such measurements.
The composition for measurement utilizing these enzyme reactions is provided in light of the chemical method and accuracy of the measurement. However, ATP can form a complex with divalent metal ions (e.g. manganese, calcium, zinc, etc.) in addition to magnesium and the complex can also be a substrate of an enzyme and, therefore, there is a risk that error will occur upon determination of magnesium ions or when a large amount of these metal ions are also present in a sample.
On the contrary, there has also been proposed that these error factors are avoided by masking a divalent metal ion other than a magnesium ion with a chelating agent (see Japanese Patent Kokai No. 1-51100).
Among various enzymatic measuring methods, a method utilizing a coupling reaction of GlcK with G6PDH is most practical from the viewpoints of stability of the reagent, accuracy of the measurement and the like. However, in this method, measuring range is comparatively narrow, particularly when a sample containing a magnesium ion in high concentration is used, which requires dilution of a sample whenever the measurement is made making the measurement complicated. Therefore, regarding a sample containing a magnesium ion in high concentration such as urea and the like, measurement errors are caused by the complicated operation.
DISCLOSURE OF THE INVENTION
The object of the present invention is to provide a reagent for measurement of magnesium ions, which has a measuring range wherein the determination can be made without dilution even in a sample containing high concentration of magensium ions.
The present inventors have studied intensively for the purpose of providing a composition for measurement of a magnesium ion which satisfies the above requirements. As a result, it has be
REFERENCES:
patent: 4623625 (1986-11-01), Scopes
patent: 4778754 (1988-10-01), Murachi
Scopes, et al., The Biochemical Journal, "Simultaneous purification and characterization of glucokinase, fructokinase and glucose-6-phosphate dehydrogenase from Zymomonas mobillis", vol. 228, No. 3, pp. 627-634, Jun. 15, 1985.
Kawahara Kazue
Kondou Hitoshi
Nagata Kazuhiko
Gitomer Ralph
Iatron Laboratories Inc.
Unitika Ltd.
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