CS228836B1 - Composition for photometric kinetic determination of creatinine for suppressing billirubin interference - Google Patents

Abstract

The reagent of the invention contains per mole of mole picric acid given a number of moles copper salts, preferably sulphate or copper (I) chloride, given the amount of moles soda-sodium tartrate and moles of dibutylnaphthalenesulfonate sodium. Reagent on contains some moles of hydroxide sodium. Serum creatinine determination and urine is suitable for laboratory use diagnostic method for monitoring n kidney disease.

Description

( ⁵⁴ ) Reagent for photometric kinetic determination of creatinine to suppress bilirubin interference

The agent according to the invention contains, per 1 mol of picric acid, a given amount of moles of copper salt, preferably copper sulfate or chloride, a given amount of moles of sodium tartrate and moles of sodium dibutylnaphthalenesulfonate. The reagent further comprises a certain amount of moles of sodium hydroxide.

The determination of creatinine in serum and urine is suitable for laboratory diagnostic methods for monitoring n kidney disease.

228,836, 228,838

The present invention provides a reagent for the photometric kinetic determination of creatinine to suppress bilirubin interference.

Determination of serum creatinine and urine is a serious laboratory diagnostic method for the monitoring of kidney disease, which is a common analysis carried out in all basic types of healthcare facilities.

Photometric methods for determining the creatinine content of a biological material are mostly based on the Jaffa reaction of creatinine with an alkaline picric acid solution to form an orange-colored reaction product. This reaction is of little specificity and, in addition to creatinine, a number of other serum components react, such as glucose, some keto-compounds and reducing agents, which are collectively included under the so-called pseudochromogens.

More complicated procedures for photometric determination of creatinine in biological material often follow previous separation of interfering substances, or separation of creatinine from the sample. They are laborious and time-consuming and therefore hardly used in practice. Simple older routine methods of photometric determination of serum creatinine are mostly based on photometric determination after previous deproteinization of serum, thus eliminating especially adverse effects of proteins and bilirubin. _{228 83e}

Kinetic methods for the determination of serum creatinine are significantly faster and simpler, no deproteinization is necessary. The omission of serum protein deproteinization is replaced in kinetic procedures by the addition of surfactants, usually sodium lauryl sulfate, which partially eliminates the effect of proteins. The interference of other pseudochromogens in the kinetic assay is suppressed due to their different reaction rates * as compared to the reaction rate of creatinine with picric acid. For example, the assay in this case does not interfere with acetacetate, uric acid and ascorbic acid. However, the interference of bilirubin, whose presence causes a negative error proportional to its concentration in the analyzed sample, is not suppressed. Even at normal serum bilirubin concentrations of 20 [mu] mol / l, a systemic negative error of up to 25 [uraol creatinine / 1] can be achieved in a kinetic assay with hitherto known agents. Falsely lower serum creatinine due to bilirubin interference may result in misdiagnosis and patient injury, especially in the initial condition of renal impairment. Thus, it is desirable to provide a photoipetric kinetic assay for creatinine in biological fluids to provide a tensile agent that would eliminate the effect of serum bilirubin.

In order to eliminate the effect of bilirubin in the photometric kinetic determination of creatinine, the agent according to the invention is characterized in that it contains from 0.01 to 0.06 moles of copper salt, preferably copper sulphate or chloride, per mol of picric acid. 2 moles of sodium potassium tartrate and 0.5 to 0.9 moles of sodium dibutylnaphthalenesulfonate. The agent according to the invention may further comprise 12 to 16 moles of sodium hydroxide, based on 1 mol of picric acid. _{228 83g}

The agent of the invention may be prepared separately in the form of two solutions. The first solution may preferably comprise picric acid, a copper salt, sodium potassium tartrate and sodium dibutyl naphthalenesulfonate, the second solution may be sodium hydroxide alone. Splitting the agent of the invention into two solutions is preferred for two reasons. In the kinetic determination of creatinine by authoritative analyzers, the sample is usually mixed with one reagent solution and the other is used as a starter for the kinetic reaction. Separation of the hydroxide solution from the picric acid solution also increases the stability of the reagent. If a single reagent is used for analysis, it is convenient to prepare them by mixing the two solutions prior to the analysis.

When analyzing sera containing 100 µmol of creatinine / l to which bilirubin was added in an amount of 20 to 200 µmol / l, the correct creatinine content was always found with the agent of the invention and the effect of the added bilirubin was not manifested. In contrast to the agents known to date, the correct creatinine content can also be determined with the agent of the invention in samples containing normal or even significantly increased amounts of bilirubin, which has a major impact on early recognition of renal impairment, allows proper diagnosis, treatment and control.

Example

A solution is prepared containing 25.6 g picric acid, 32.0 g sodium dibutylnaphthalenesulfonate, 2.6 g sodium potassium tartrate and 0.87 g of copper (II) pentahydrate in 6600 ml of water. When <

228 838

A solution containing 66 g of sodium hydroxide in 220 ml of water is added.

Before use, 3 parts by volume of the first solution are mixed with 1 part by volume of the second solution.

Example 2

Prepare a reagent containing in 1000 ml of an aqueous solution of 126 mmol of picric acid, 0.126 mmol of cupric nitrate, 0.126 mmol of sodium potassium tartrate, 100 mmol of sodium dibutylnaphthalisulfonate and 151 mmol of sodium hydroxide. The reagent is used directly.

Example 3

Prepare a reagent containing in 100 ml of an aqueous solution of 12.6 mmol of picric acid, 0.76 mmol of cupric chloride, 2.5 mmol of sodium potassium tartrate, 8 mmol of sodium dibutylnaphthalenesulfonate and 201 mmol of sodium hydroxide. The reagent is used directly ·

Example 4

To a 0.5 ml sample of serum or urine diluted 1 to 49 with water previously, 1.6 ml of the reagent prepared according to Examples 1 to 3 is added and the change in absorbance measured between 30 and 120 s after mixing the sample with the reagent at constant temperature. in a cuvette of 1 cm at 510 nm.

- 228 838

Κ 0.5 ml of serum or diluted urine sample is added with 1.2 ml of the first solution according to Example 1 and the reaction is started with 0.4 ml of sodium hydroxide solution. The procedure is the same as in the previous case.

Claims (2)

SUBJECT OF THE INVENTION 228 836 A creatinine photometric kinetic assay reagent for suppressing bilirubin interference, characterized by: 1 mol of picartaric acid contains 0.01 to 0.06 moles of copper salt, preferably copper sulphate or chloride, 0.01 to 0.2 moles of sodium tartrate and 0.5 to 0.9 moles of sodium dibutylnaphthalenesulfonate. 2. Reagent according to claim 1, characterized in that it contains 12 to 16 moles of sodium hydroxide, based on 1 mol of picric acid.