Materials Included In Kit

Additional Materials Required

Safety Precautions

Biuret test solution is a corrosive liquid and is especially dangerous to eyes. Silver nitrate solution is moderately toxic by ingestion. It is also irritating to body tissues and will stain skin and clothing. Avoid all body tissue contact. Potassium chloride is slightly toxic by ingestion. Wear chemical splash goggles, chemical-resistant gloves and a chemical-resistant apron. Remind students to wash their hands thoroughly with soap and water before leaving the laboratory. Please review current Safety Data Sheets for additional safety, handling and disposal information.

Disposal

Please consult your current Flinn Scientific Catalog/Reference Manual for general guidelines and specific procedures, and review all federal, state and local regulations that may apply, before proceeding. Neutralize the simulated urine/biuret mixture by adding dilute hydrochloric acid solution and rinse down the drain with excess water according to Flinn Suggested Disposal Method #10. The simulated blood/silver nitrate mixture may be flushed down the drain with copious amounts of water according to Flinn Suggested Disposal Method #26b. The remaining solution as well as the simulated urine/Benedict’s solution mixture may be rinsed of down the drain according to Flinn Suggested Disposal Method #26b.

Lab Hints

• Enough materials are provided in this Super Value Kit for 5 classes of 30 students each, working in pairs (75 total student groups). This laboratory activity may reasonably be completed in one 50-minute class period. The prelaboratory assignment may be completed before coming to lab, and the data compilation and questions may be completed the day after the lab.

Teacher Tips

• This is an excellent activity for students to do upon studying renal physiology or diabetes.
• Flinn carries a diverse selection of forensics kits. See the Forensics section in the Flinn Scientific Catalog/Reference Manual.

Answers to Prelab Questions

1. What type of test would be use to detect the presence of sodium in a blood or urine sample? Describe the test procedure as well as the positive result.

   Sodium would be detected by the use of a flame test. If the flame burned a bright yellow color it would indicate the presence of sodium.

2. Upon adding Benedict’s solution to a urine sample, a red/orange precipitate is formed. What does this indicate about the urine sample?

   The formation of the red/orange precipitate indicates the presence of a reducing sugar in the urine.

Sample Data

Part A. Urinalysis Table

Part B. Blood Analysis Table

Answers to Questions

1. Based on the results of the five tests what is the likely cause of Mr. Smith’s death?

   Elevated levels of proteins, glucose, potassium and chloride ions were found in Mr. Smith’s system. He tested negative for sodium. These results indicate he probably died of kidney failure.

2. If urinalysis was not conducted, what other tests could rule out dehydration as a possible cause of death for him?

   A flame test was conducted to indicate the presence of potassium and absence of sodium. If Mr. Smith had died from dehydration he would have also tested positive for sodium. The presence of phosphate would also confirm his death as kidney failure and not dehydration.

3. Given that no real blood was used in this laboratory activity, what compound may have been added to the simulated blood sample to yield the results found using the chloride test?

   Any chloride compound such as lithium chloride, sodium chloride or potassium chloride could yield a positive result. However, since the flame test resulted in a lavender flame it is likely that the chloride ion introduced to the system was in the form of KCl.

References

Collins, David. Investigating Chemistry in the Laboratory; W. H. Freeman and Company: New York, NY; 2006; pp. 49–57.

Introduction

Mr. Smith was found in his bed deceased. He had no known pre-existing conditions. Perform four tests using his blood and urine to determine a possible cause of death.

Concepts

• Urinalysis
• Blood analysis
• Diseases

Background

The presence and concentrations of chemical species in blood and urine are often used as diagnostic tools for illness and cause of death. Components of blood and urine give insight to the body’s current health. Medical professionals commonly use blood and urine samples to diagnose health conditions. These samples are also valuable to toxicologists to determine a possible cause of death.

The composition of blood and urine varies with changes in health. The presence of proteins, glucose and ketones are often analyzed in urine samples. Healthy individuals exhibit very low concentrations of these chemical compounds. The presence of one or more of these chemical compounds at substantial levels in urine may indicate conditions, such as dehydration, kidney damage or diabetes. Both urine and blood are frequently tested for levels of sodium and potassium. In addition, blood is tested for the presence of ammonium, calcium, chloride and phosphate ions. All of the previously mentioned ions or electrolytes are normally present in urine and/or blood in accepted healthy values. Elevated electrolyte concentrations that vary from the accepted values may be indicative of various diseases or conditions, such as diabetes or heart failure. Explanations of the tests that will be done in this laboratory are included.

Protein Detection
Proteins can be identified using a simple color test based on the reaction of their polypeptide backbones with copper ions in basic solution. When molecules containing two or more peptide linkages react with copper sulfate in the presence of a strong base, a purple complex is formed. This is called the biuret test. The colored product is the result of coordination of peptide nitrogen atoms with copper ions. The amount of product that is formed and thus the intensity of the purple color depend on the nature of the protein and the amount of protein present.

Electrolyte Levels
One method of detecting excess or elevated electrolyte levels in urine or blood is by using a flame test. Most metal ions emit a characteristic wavelength or color of light when heated in a flame. Just as a fingerprint is unique to each person, the wavelength of light emitted after excitation of an element is unique to that element. Only a few elements give off a characteristic color of light in the visible region of the spectrum—the region which is visible to the human eye (400–700 nm). For most elements, the characteristic wavelength is detectable only in the ultraviolet or infrared regions of the spectrum. See Table 1 for a list of a few metal ions that emit characteristic visible colors.

Sugar Detection
Benedict’s qualitative solution is used to test for the presence of reducing sugars in urine. A reducing sugar contains an aldehyde or another group which is capable of reducing copper ions in Benedict’s solution. In a reaction with copper(II), an aldehyde group in glucose, for example, is oxidized to a carboxylic acid (see Equation 1). All monosaccharides and some disaccharides are reducing sugars. Examples of reducing sugars include glucose, fructose, galactose and lactose. Notably, sucrose, which is table sugar, is not a reducing sugar. Initially, the copper(II) ions in the Benedict’s solution impart a characteristic blue color to the solution. However, when Benedict’s solution is added to a solution containing a reducing sugar, the blue copper(II) ions are reduced to copper(I) ions by the reducing sugar to form red copper(I) oxide, Cu₂O, which precipitates out of solution. Therefore, the formation of a red precipitate indicates a positive test for reducing sugars.

Chloride Detection
Elevated concentrations of chloride ions may be detected using a chemical test with silver ions in solution. Blood contains many dissolved ions from the dissociations of salts, such as sodium or potassium chloride.

An ionic salt compound is composed of two parts—cations (positively charged ions) and anions (negatively charged ions). When an ionic salt is dissolved in water, the salt crystal dissociates or separates into its corresponding cations and anions. For example, potassium cloride (KCl) dissociates into potassium cations (K⁺) and chloride anions (Cl^–) according to Equation 2. Similarly the ionic salt silver nitrate, AgNO₃, dissociates into silver cations (Ag⁺) and nitrate anions (NO₃^–) according to Equation 3. When two ionic salts are mixed together in water, two new combinations of cations and anions are possible. In some cases the cation from one salt and the anion from another salt may combine to form an insoluble solid product, called a precipitate. For example, if solutions of potassium chloride and silver nitrate are mixed together, a solid precipitate of silver chloride (AgCl) forms as shown by Equation 4. Notice the potassium cations (K) and nitrate anions (NO₃^–) remain dissolved in solution. They do not combine to form a precipitate and thus do not participate in the reaction. They are therefore referred to as spectator ions. A net ionic equation is one that includes only the ions participating in the reaction. Thus Equation 4 can be reduced to Equation 5. The reaction shown in Equation 5 is used as the basis of a test to detect excess chloride ions in blood samples.

Experiment Overview

In order to determine Mr. Smith’s possible cause of death samples of both urine and blood will be tested. A sample of simulated urine will be analyzed for the presence of excess proteins and glucose. A sample of simulated blood will be tested for elevated levels of sodium, potassium and chloride ions.

Materials

Prelab Questions

1. What type of test would be use to detect the presence of sodium in a blood or urine sample? Describe the test procedure as well as the positive result.
2. Upon adding Benedict’s solution to a urine sample, a red/orange precipitate is formed. What does this indicate about the urine sample?

Safety Precautions

Biuret test solution is a corrosive liquid and is especially dangerous to eyes. Benedict’s qualitative solution is a skin and eye irritant. Silver nitrate solution is moderately toxic by ingestion. It is also irritating to body tissues and will stain skin and clothing. Avoid all body tissue contact. Wear chemical splash goggles, chemical-resistant gloves and a chemical-resistant apron. Wash hands thoroughly with soap and water before leaving the laboratory. Please follow all laboratory safety guidelines.

Procedure

Part A. Urinalysis

Protein

1. Place 100 mL of tap water in a 200-mL beaker.
2. Heat the beaker of water on a hot plate to prepare a boiling water bath for step 11. Proceed to step 3 while the water heats to a boil.
3. Using a clean graduated cylinder, measure 5 mL of simulated urine.
4. Add the simulated urine to a clean 13 x 100 mm test tube.
5. Using a graduated pipet, measure 2.5 mL of biuret solution. Hint: Use the graduations on the side of the pipet to do this.
6. Add the biuret solution to the test tube containing the 5 mL of simulated urine.
7. After 2–3 minutes, record observations in the Urinalysis Table of The Coroner’s Report Worksheet.

Glucose

8. Using a clean 10-mL graduated cylinder, measure 5 mL of Benedict’s qualitative solution.
9. Add the Benedict’s qualitative solution to a clean 13 x 100 mm test tube.
10. Using a graduated pipet add 8 drops of simulated urine to the test tube containing the Benedict’s qualitative solution.
11. Place the test tube in the boiling water bath from step 2.
12. Allow the test tube to sit in the boiling water for 3–5 minutes and record observations in the Urinalysis Table of the worksheet.
13. Turn off the hot plate and allow the boiling water to cool before discarding.

Part B. Blood Analysis

Sodium and Potassium

14. Using a clean graduated cylinder, measure 5 mL of simulated blood.
15. Add 2–3 mL of simulated blood to a clean 13 x 100 mm test tube. Note: The remaining simulated blood will be used in the Chloride Test.
16. Light a Bunsen burner and obtain an inoculating loop.
17. Dip the inoculating loop in the simulated blood solution and hold the tip of the loop over the flame.
18. Record observations in Blood Analysis Table of the worksheet.

Chloride

1. Using a clean graduated pipet, add 1 mL of simulated blood to a clean 13 x 100 mm test tube.
2. Using a clean graduated pipet, add 3–5 drops of silver nitrate to the test tube containing the simulated blood.
3. Wait 3–5 minutes. Record observations in the Blood Analysis Table of the worksheet.
4. After all test have been completed, obtain the Pathology Evidence Chart from the instructor.

Part C. Pathology Evidence Chart

23. Use the following information to analyze the results obtained in the urine and blood test and to identify the possible cause of death.
    {12022_Procedure_Table_2_Pathology evidence chart}

    X = Positive test results

24. Please consult your instructor for appropriate disposal procedures.

Student Worksheet PDF

12022_Student1.pdf