JPH0658364B2 - Catecholamine final metabolite analyzer - Google Patents

Description

Detailed Description of the Invention a. TECHNICAL FIELD The present invention relates to a liquid chromatograph device for quantitatively analyzing catechol final metabolites in urine.

B. BACKGROUND ART Measurement of catechol final metabolites such as vanillyl mandelic acid (hereinafter referred to as VMA) and homovanillic acid (hereinafter referred to as HVA) contained in urine is performed by a liquid chromatograph for the purpose of screening for neuroblastoma. However, there is a problem in that the extraction operation with an organic solvent such as ethyl acetate is required to remove the impurities contained in the urine, and the analysis work is troublesome.

C. In view of such problems, an object of the present invention is to provide a liquid chromatograph apparatus for catecholamine final metabolite analysis, which enables rapid analysis without extraction work.

D. Configuration of the Invention That is, the feature of the present invention is that a mixed solution of an acidic buffer solution and acetonitrile is used as a mobile phase, and a silica gel carrier chemically bound with a cyanopropyl group is used as a stationary phase.

E. Examples Therefore, details of the present invention will be described below based on illustrated examples.

FIG. 1 shows an embodiment of the present invention, in which reference numeral 1 is a column packed with silica gel particles chemically bonded to a cyanopropyl group, one end of which is a mobile phase to be described later via a pump 2. The container 4 containing the liquid 3 is connected to the electrochemical detector 5 on the other side. 3 is the above-mentioned mobile phase solution, which is prepared by mixing 5% by volume of acetonitrile with an acidic buffer solution such as tartaric acid, acetic acid or phosphoric acid. Reference numeral 6 in the figure indicates a sample injection port.

In this example, when the collected urine is injected into the column 1 while the mobile phase liquid 3 is flowing in the column 1, the contaminant components contained in the urine are separated in an extremely short time and flow into the detector 5. To do. On the other hand, the target components VMA and HVA are detected by the electrochemical detector as peaks separated from each other without being affected by the contaminant components.

[Example] As a mobile phase liquid, 5 mmol of tartaric acid (95% by volume) and acetonitrile (5% by volume) were mixed, and as a column, a packing in which a cyanopyryl group was chemically bonded to a spherical silica carrier of 5 μm was used. Using a liquid chromatograph equipped with a tube with an inner diameter of 6 mm and a length of 15 cm, a standard sample prepared by mixing and dissolving standard VMA, VLA and HVA in distilled water was analyzed, and shown in FIG. As described above, the peaks of VMA, VLA and HVA could be detected separately.

Next, urine of a normal child and a child with neuroblastoma was diluted about 10 times with a diluent having the same composition as the mobile phase solution, and analyzed by the above-mentioned apparatus. As shown in Fig. 3 (a) and (b), the peaks of VMA and HVA can be separated and detected without being affected by impurities contained in urine, and It was possible to detect the peak heights of VMA and HVA while distinguishing them significantly.

From this, it was found that VMA and HVA contained in urine can be quantified with high accuracy without being affected by contaminants.

Further, when the concentration of acetonitrile was changed and measured in the range of 3 to 8% by volume, the time required for the target component to elute increased as the concentration of acetonitrile decreased, but there was no problem in practical use.

In addition, when a so-called gradient elution method, in which the volume ratio of acetonitrile to the acidic buffer solution is changed, was applied and analysis was performed, the elution time of the target component could be shortened.

When a fluorescence detector was used as the detector, only the target component could be selectively detected without being affected by the interfering components as in the electrochemical detector.

In the above-mentioned examples, VMA, VLA, and HVA useful for the screening of patients with neuroblastoma have been described, but other end products of catecholamine collection could also be detected. Further, when other acidic buffer solutions such as acetic acid and phosphoric acid were used instead of tartaric acid, the detection accuracy was not affected at all.

F. Effects As described above, according to the present invention, a mixture of acetonitrile and an acidic buffer solution is used as a mobile phase, and a silica gel carrier chemically bound with a cyanopropyl group is used as a stationary phase. The catecholamine final metabolite can be quantitatively analyzed with high accuracy and in a short time only by performing the treatment, and an extremely effective device can be realized for rapid processing of a sample in mass screening for neuroblastoma.

[Brief description of drawings]

FIG. 1 is a block diagram of an apparatus showing an embodiment of the present invention, and FIG.
FIG. 3 and FIG. 3 are chromatogram diagrams showing an example of the measurement results by the same device. 1 ... column, 3 ... mobile phase liquid

Claims (1)

[Claims] 1. A catecholamine final metabolite analyzer comprising an acidic buffer solution containing acetonitrile as a mobile phase and a silica gel carrier chemically bound with a cyanopropyl group as a stationary phase.