TEST FOR GLUCOSE IN URINE

TEST FOR GLUCOSE

A TEST FOR GLUCOSE- Glucose In urine (glycosuria), glucose test may suggest if the patient has symptoms of diabetes or glycosuria patent is being treated with drugs of diabetes.

mainly two types of test have done in the medical laboratory. for glucose estimation in urine tests are performed in a medical laboratory to detect glucose in the urine  these are

• Benedict's Test (Qualitative)
• Benedict's Quantitative Glucose Test

The qualitative test detects the presence or absence of glucose in urine and reports as positive and negative.

The qualitative test detected the quantity of glucose concentration in human urine it is express in the quantity that you will learn below.

GLUCOSE - Glucose In normal urine, glucose may be present only in trace amounts. In the kidneys, glucose is filtered by the glomeruli and reabsorbed by the tubules. However, when the blood glucose concentration rises above 10.0 mmol/L (renal threshold), the tubules cannot reabsorb all the glucose. 

• The surplus glucose appears in the urine and this is glycosuria. In pathological conditions, glucose is about the only carbohydrate found in urine. 
• Diabetes mellitus is the most important of these conditions when as much as 280 mmol/L of glucose can be found in urine. 
• Glycosuria can also occur in severe liver disease and in endocrine hyperactivity.

Galactose-  Galactose in urine (galactosuria) may be harmless following heavy consumption of galactose or lactose. But galactosuria in infants can be due to a deficiency of an enzyme necessary for the conversion of lactose to glucose. This is a pathological condition that may result in mental retardation.

Sucrose- Sucroseappears in the urine (sucrosuria) of patients suffering from pancreatitis. It is also found in the urine of some infants suffering from gastroenteritis. Although it is a reducing substance, it gives a negative result with Benedict's solution.

• There are some tests which specifically detect glucose and others are designed to detect reducing substances. 
• The reducing substances are also commonly known as "sugars".

Benedict's Test (Qualitative)

Principle- The test is based on the reduction of alkaline blue cupric sulphate to the yellow or red cuprous oxide by the aldehyde or ketone group of  carbohydrate or ketone group of stances in urine, the colour of the reagent remains blue-green.

Benedict's qualitative reagent:-

Solution A  173 g sodium citrate and 100 g anhydrous sodium carbonate are dissolved with the aid of heat in about 600 ml of distilled water.

Solution B  17.3 g copper sulphate is dissolved in 100 ml distilled water.

Add solution B, slowly with constant stirring, to solution A. After cooling, transfer to a one litre volumetric flask and make up the volume to the mark with distilled water.

Benedict's qualitative Method- Place about 0.5 ml of urine in a large test tube. Add 5 ml of Benedict's reagent to the urine. Mix well and place in a boiling water bath for 5 minutes. Allow to cool and observe for colour change due to precipitation of cuprous oxide.

Reporting of results

Reporting of results
Green colour Negative	-
Yellow-green precipitate	+/-
< 0.5 g/dl) Greenish-yellow precipitate	+
(0.5 to 1 g/dl) Yellow precipitate	++
(1 to 2 g/dl) Orange to brick-red precipitate	+++
>2 g/dl)	++++

Note-White precipitate due to phosphates or development of any other colour should be ignored.

Benedict's Quantitative Glucose Test-

Benedict's Quantitative Principle

Glucose reduces cupric ions to cuprous ions. These react with potassium thiocyanate to produce a white precipitate of cuprous thiocyanate. The blue colour disappears as the white precipitate is formed. This indicates a complete reduction of cupric ions which is the endpoint.

Benedict's quantitative reagent 

• Dissolve 200 g of sodium citrate
• 75 g of anhydrous sodium carbonate  
• 125 g of potassium thiocyanate in 600 ml of distilled water. 
• Heat to dissolve completely, cool to room temperature and filter with ordinary filter paper. 
• To the mixture, add 18 g of copper sulphate dissolved in about 100 ml of distilled water, slowly with constant stirring. 
• Add 5 ml of 5% potassium ferrocyanide. 
• Mix well and make final volume to I litre with distilled water. 
• The reagent is stable at room temperature.

Benedict's quantitative Method­:- 

• 25 ml of Benedict's quantitative solution in a 250 ml beaker, add about 10 g of anhydrous sodium carbonate.
• Bring the mixture to the boil, and slowly run in the urine sample from a pipette, while the mixture is still boiling. 
• When the blue colour of the reagent begins to fade, add the urine drop by drop until all
• the blue colour has disappeared. (All the cupric ions in the reagent are now reduced.) Note the volume of urine used.

Benedict's quantitative Calculation:- 25 ml of Benedict's quantitative reagent are reduced by 0.05 g of glucose.

Therefore,

 

Note- The test can also be performed by using 5.0 ml of Benedict's quantitative reagent and 2.0 g of anhydrous sodium carbonate.

	The approximate R1 value of urinary
S. NO.	SUGAR	CHARACTERISTICS of COLOUR	Approx R1
1.	Lactose	Gray	0.14
2.	GALACTOSE	Gray	0.34
3.	FRUCTOSE	PINK	0.39
4.	GLUCOSE	GREY	0.42
5.	PENTOSES	GRAY BROWN	>0.5

  

Commercial Strips for Glucose

There are various brands of commercial test strips available.
The most popular ones include,

1. Clinistix (Ames), 
2. BM test (BCL) 
3. Glucostrips (Hi-Tech)
4. Diabur (Boehringer). 

They are all based on similar principles.

Commercial Strips for Glucose

Clinistix

Principle of Clinistix

• The test is based on the principle that glucose is oxidised by atmospheric oxygen in the presence of glucose oxidase to gluconic acid and hydrogen peroxide. 
• The hydrogen peroxide, in the presence of peroxidase, oxidises the chromogen to various shades of purple.

Method

• Dip the reagent end of the strip into fresh urine and remove immediately. 
• Compare the test colour with the colour chart within 10 seconds. 
• Read carefully in a good light. 
• The test is no longer valid after 10 seconds.

interpretation of the results-

•  Purple within 10 seconds = Presence of glucose.
•  Cream after 10 seconds = Absence of glucose.

The sensitivity of the test is as high as 0.01 g/100ml.Some test strips such as Diabur test 500. measure glucose semi-quantitatively.

Note

False-negative or false positive test results do occur with reagent strips. example,

(i)                Oxidation of certain substances such as ascorbic acid or some drug products, in preference to the chromogen may give a false-negative result.

(ii)              A false-negative result may also be obtained due to a high concentration of catalase, as in E.coli infection, when hydrogen peroxide destroyed.

    
Test For Fructose-Seliwanoff's Test
  
Test For Lactose Fearon's Methylamine Test
  
Test For Pentose-Bial's test
  
Chromatography of Urine Sugars