PRACTICUM II URINE CHEMICAL EXAMINATION KETONES 2.3 Ketones Practice : Urine Ketone Examination Objective : To determine the presence of ketone bodies in the urine Ketone substances or ketone objects in urine are acetone, aceto-acetic acid and betahydroxybutyric acid. Because acetone, the most important substance among ketones, is volatile, the urine examined must be fresh; If the urine is left, the aceto-acetic acid turns into acetone, as does the beta-hydroxybutyric acid which first becomes aceto-acetic acid, so that these substances also disappear from the urine. 2.3.1 Rothera Method Objective :To detect ketones in urine, especially aceto acid acetate or acetone. Working Principle: The presence of ketones (acetoacetic acid or acetone) in the urine will reacts with nitroprusside to form a purple ring. Tools and materials : 1. Urine sample 2. Test tube 3. Rothera reagent 4. NH4OH 28% 5. Measuring cup Ways of working : 1. Put 5 ml of urine into a test tube. 2. Add 1 gram of rotera powder to the tube and shake until it dissolves. 3. While tilting and being careful, flow or drip 1 – 2 ml of 28% concentrated ammonium hydroxide through the wall onto the urine. The 28% concentrated ammonium hydroxide must form the top layer of the liquid in the tube. 4. Place the tube into the tube rack and read the results in 3 minutes. 5. The reaction results are observed 6. The reddish purple color at the border of the two liquid layers indicates the presence of ketone substances. The faster the color occurs and the darker the color, the greater the amount of ketones. The color brown is given a negative meaning. Because this test cannot be given a regular and definite semiquantitative assessment, state the results as positive (+) and negative (-) only. 2.3.2 Method: Lange Tools and materials: - Test Tubes & Tube Racks - Scale & Bulb Pipette - Pasteur pipette - Glacial Acetic Acid Sodium - 5% Sodium Nitroprusside Lard - Lar. Ammonia 10% Ways of working : 1. Put 2 ml of urine into a test tube 2. Added to urine, 0.1 mlGlacial Acetic Acid & Lar. Na-Nitroprusside 5% 1-2 drops then shake until homogeneous 3. Through the tube wall, slowly flow the liquid. 2 ml of 10% ammonia 4. After that, the tube is placed on the tube rack in an upright position for 3 minutes. 5. Observe the reaction results Interpretation of Results: Negative (-): no purple ring occurs at the border of the two liquid layers. Positive (+): a purple ring occurs at the border of the two liquid layers.