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Energy and carbohydrate metabolism
NAME
SECTION
DATE .
m
Energy and carbohydrate metabolism E X P E R I M E N T 17A OBJECTIVE:
To perform a test for lactic acid.
EQUIPMENT:
50-ml Erlenmeyer flask test tubes (3)
CHEMICALS:
ferric chloride lactic acid (1%) gastric contents or substitutes (varying concentrations of lactic acid in 0.1 TV hydrochloric acid)
Lactic acid is formed by the action of bacteria on carbohydrates. It is present in unusually large amounts in the stomach contents of persons with cancer of the stomach. Ο II CH3-CH-C-OH I OH lactic acid Pour 30 ml of water into a 50-ml flask. Add 6 drops of 10% ferric chloride ( F e C l 3 ) solution. Mix the contents of the flask. Label three test tubes # 1 to # 3 . Pour 10 ml of the solution into each tube. Tube # 1 is the "control." Add nothing to this tube. Tube # 2 is the "reference." Add 1 ml of a dilute lactic acid solution. Describe the result.
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CHEMISTRY
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Tube # 3 is the "test." Add 1 ml of a specimen of gastric contents (or a substitute). Describe the results.
Does the test sample contain lactic acid?
I M E N T 17B OBJECTIVE:
To perform a quantitative test for glucose using a spectrophotometer.
EQUIPMENT:
test tubes (3, large) cuvets ring stand 500-ml beaker Spectronic 20
CHEMICALS:
glucose (1%) urine dintrosalicylic acid (DNS) (In a 2000-ml flask, place 300 ml 4.5% NaOH, 880 ml 1% aqueous DNS, 225 g potassium sodium tartrate. Mix until dissolved, store in brown, stoppered, glass bottle.)
The tests for glucose that you carried out in previous experiments were qualitative. They provided only the approximate concentration. In this experiment you will perform a quantitative determination, obtaining a more exact answer. Take 3 large test tubes and fill them each with 25 ml of water. Mark each tube with a line to show the water level when the tube is vertical (standing upright). Empty the tubes. Mark them # 1 , # 2 , and # 3 . Obtain a 1-ml urine sample and dilute it 1:10 (1 ml urine + 10 ml water). Place 1 ml of this dilute urine into tube # 1 . To tube # 2 , add 1 ml of a dilute (1%) glucose solution. To tube # 3 , add 1 ml of water.
NAME
SECTION
Figure 17-1
DATE
Spectronic 20. (Courtesy Bausch and Lomb.)
The function of each tube is shown below. Function test (unknown) standard (reference) blank (solvent only)
Tube number 1 2 3
To each of the tubes, add 3 ml of dinitrosalicylic acid solution. Place the tubes in boiling water for exactly 5 minutes. Cool the tubes under running water. Fill each tube with water up to the mark you made so that each solution now has a volume of 25 ml. The color of each solution depends on the concentration of glucose. You can measure the intensity of the color using a spectrophotometer, shown in Figure 17-1. Set the instrument (amplifier control knob on left) to read 0% at 525 nanometer (nm). Place a tube (cuvet) filled with solution # 3 into the sample holder. Adjust the instrument (light control knob on right) to read 100%. Fill the sample tube (cuvet) with solution # 1 . Read the absorbance (A) of the solution. The absorbance measures the decrease in the power of the light beam passing through the sample tube—part of the radiation is absorbed by the colored compound formed from the glucose. ,4(tube#l) =
145
UCKO /EXPERIMENTS
FOR LIVING
CHEMISTRY
/ 17
Fill the sample tube with solution # 2 . Read again. ,4(tube # 2 ) = The concentration of urine (g/100 ml) in tube # 1 is equal to >4(tube#l)= ,4 (tube # 2 )
=
QUESTIONS E X P E R I M E N T 17A
1.
At what point in carbohydrate metabolism is lactic acid produced?
2.
What is the purpose of "blank" and "reference" tubes?
E X P E R I M E N T 17B
146
1.
Why might a quantitative glucose test be useful?
2.
What is the purpose of the spectrophotometer?
3.
What would you do if the reading of your sample went off-scale (too high a reading)?
SECTION
DATE .
m
Energy and carbohydrate metabolism E X P E R I M E N T 17A OBJECTIVE:
To perform a test for lactic acid.
EQUIPMENT:
50-ml Erlenmeyer flask test tubes (3)
CHEMICALS:
ferric chloride lactic acid (1%) gastric contents or substitutes (varying concentrations of lactic acid in 0.1 TV hydrochloric acid)
Lactic acid is formed by the action of bacteria on carbohydrates. It is present in unusually large amounts in the stomach contents of persons with cancer of the stomach. Ο II CH3-CH-C-OH I OH lactic acid Pour 30 ml of water into a 50-ml flask. Add 6 drops of 10% ferric chloride ( F e C l 3 ) solution. Mix the contents of the flask. Label three test tubes # 1 to # 3 . Pour 10 ml of the solution into each tube. Tube # 1 is the "control." Add nothing to this tube. Tube # 2 is the "reference." Add 1 ml of a dilute lactic acid solution. Describe the result.
143
UCKO/EXPERIMENTS
FOR LIVING
CHEMISTRY
/ 17
Tube # 3 is the "test." Add 1 ml of a specimen of gastric contents (or a substitute). Describe the results.
Does the test sample contain lactic acid?
I M E N T 17B OBJECTIVE:
To perform a quantitative test for glucose using a spectrophotometer.
EQUIPMENT:
test tubes (3, large) cuvets ring stand 500-ml beaker Spectronic 20
CHEMICALS:
glucose (1%) urine dintrosalicylic acid (DNS) (In a 2000-ml flask, place 300 ml 4.5% NaOH, 880 ml 1% aqueous DNS, 225 g potassium sodium tartrate. Mix until dissolved, store in brown, stoppered, glass bottle.)
The tests for glucose that you carried out in previous experiments were qualitative. They provided only the approximate concentration. In this experiment you will perform a quantitative determination, obtaining a more exact answer. Take 3 large test tubes and fill them each with 25 ml of water. Mark each tube with a line to show the water level when the tube is vertical (standing upright). Empty the tubes. Mark them # 1 , # 2 , and # 3 . Obtain a 1-ml urine sample and dilute it 1:10 (1 ml urine + 10 ml water). Place 1 ml of this dilute urine into tube # 1 . To tube # 2 , add 1 ml of a dilute (1%) glucose solution. To tube # 3 , add 1 ml of water.
NAME
SECTION
Figure 17-1
DATE
Spectronic 20. (Courtesy Bausch and Lomb.)
The function of each tube is shown below. Function test (unknown) standard (reference) blank (solvent only)
Tube number 1 2 3
To each of the tubes, add 3 ml of dinitrosalicylic acid solution. Place the tubes in boiling water for exactly 5 minutes. Cool the tubes under running water. Fill each tube with water up to the mark you made so that each solution now has a volume of 25 ml. The color of each solution depends on the concentration of glucose. You can measure the intensity of the color using a spectrophotometer, shown in Figure 17-1. Set the instrument (amplifier control knob on left) to read 0% at 525 nanometer (nm). Place a tube (cuvet) filled with solution # 3 into the sample holder. Adjust the instrument (light control knob on right) to read 100%. Fill the sample tube (cuvet) with solution # 1 . Read the absorbance (A) of the solution. The absorbance measures the decrease in the power of the light beam passing through the sample tube—part of the radiation is absorbed by the colored compound formed from the glucose. ,4(tube#l) =
145
UCKO /EXPERIMENTS
FOR LIVING
CHEMISTRY
/ 17
Fill the sample tube with solution # 2 . Read again. ,4(tube # 2 ) = The concentration of urine (g/100 ml) in tube # 1 is equal to >4(tube#l)= ,4 (tube # 2 )
=
QUESTIONS E X P E R I M E N T 17A
1.
At what point in carbohydrate metabolism is lactic acid produced?
2.
What is the purpose of "blank" and "reference" tubes?
E X P E R I M E N T 17B
146
1.
Why might a quantitative glucose test be useful?
2.
What is the purpose of the spectrophotometer?
3.
What would you do if the reading of your sample went off-scale (too high a reading)?