REMOTE LAB ACTIVITY
SUBJECT SEMESTER: ________________
TITLE OF LAB: Buffers
Lab format: This lab is a remote lab activity.
Relationship to theory (if appropriate): In this lab you will learn the underlying principles on how
biological systems maintain pH within a narrow range is important to understanding the function of
these systems.
Instructions for Instructors: This protocol is written under an open source CC BY license. You may use
the procedure as is or modify as necessary for your class. Be sure to let your students know if they
should complete optional exercises in this lab procedure as lab technicians will not know if you want
your students to complete optional exercise.
Instructions for Students: Read the complete laboratory procedure before coming to lab. Under the
experimental sections, complete all pre-lab materials before logging on to the remote lab, complete
data collection sections during your on-line period, and answer questions in analysis sections after your
on-line period. Your instructor will let you know if you are required to complete any optional exercises
in this lab.
Remote Resources:
CONTENTS FOR THIS NANSLO LAB ACTIVITY:
Learning Objectives.................................................................................................. 2
Background Information ......................................................................................... 2-6
Equipment ............................................................................................................... 6
Experimental Procedure ......................................................................................... 6
Pre-lab Exercise 1: Adding an Acid to Buffer Solutions .......................................... 7
Exercise 1: Adding an Acid to Buffer Solutions ...................................................... 7
Pre-lab Exercise 2: Adding a Base to Buffer Solutions .......................................... 7-8
Exercise 2: Adding a Base to Buffer Solutions ........................................................ 8-9
Summary Questions: ............................................................................................... 9
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LEARNING OBJECTIVES:
After completing this lab, you should be able to:
1.
2.
3.
4.
5.
Define buffer, acid, and base.
Define pH and describe how the pH of a solution is measured.
Explain the effect of acids and bases on the pH of a buffered liquid.
Distinguish between acidosis, alkalosis, and the etiology of the different types.
Explain the role of the buffering systems inside our body in the prevention of acidosis and
alkalosis.
6. Compare the buffer stabilizing effect of 3 different solutions.
7. Collect and analyze data using a drop counter and digital pH probe.
8. Interpreting the data on a graph to determine the point at which buffer stabilization fails.
BACKGROUND INFORMATION:
Buffer is a solution usually containing an acid and a base, or a salt that tends to maintain a constant
hydrogen ion concentration. Ions are atoms or molecules that have lost or gained one or more
electrons. An example of a common buffer is a solution of acetic acid (CH3COOH) and sodium acetate. In
water solution, sodium acetate is completely dissociated into sodium (Na+) and acetate (CH3COO-) ions.
The hydrogen ion concentration of the buffer solution is given by the expression:
+
[𝐻 ] = 𝐾𝑎
[𝐶𝐻3 𝐶𝑂𝑂𝐻]
𝐶𝐻3 𝐶𝑂𝑂 −
in which Ka is the ionization constant of acetic acid and the expressions in brackets are the
concentrations of the respective substances. The hydrogen ion concentration of the buffer solution is
dependent on the relative amounts of acetic acid and acetate ion (or sodium acetate) present, known as
the buffer ratio. The addition of an acid or a base will cause corresponding changes in the concentration
of acetic acid and acetate ion, but as long as the concentration of the added substances is small
compared to the concentration of the individual buffer components, the new hydrogen ion
concentration will remain close to its original value. By “close to”, we generally mean within a factor of
10 of the original value. So, if the buffer starts with a hydrogen ion concentration of 1 x 10-7 M, then the
buffer is still working as long as the hydrogen ion concentration stays between 1 x 10-6 M and 1 x 10-8 M.
As acid is any substance that in water solution tastes sour, changes the color of certain indicators (e.g.,
reddens blue litmus paper), reacts with some metals (e.g., iron) to liberate hydrogen, reacts with bases
to form salts, and promotes certain chemical reactions (acid catalysis). Examples of acids include the
inorganic substances known as the mineral acids—sulfuric, nitric, hydrochloric, and phosphoric acids—
and the organic compounds belonging to the carboxylic acid, sulfonic acid, and phenol groups. Such
substances contain one or more hydrogen atoms that, in solution, are released as positively charged
hydrogen ions.
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A Base is any substance that in water solution is slippery to the touch, tastes bitter, changes the color of
indicators (e.g., turns red litmus paper blue), reacts with acids to form salts, and promotes certain
chemical reactions (base catalysis). Examples of bases are the hydroxides of the alkali and alkaline earth
metals (sodium, calcium, etc.) and the water solutions of ammonia or its organic derivatives (amines).
Such substances produce hydroxide ions (OH-) in water solutions.
pH is a quantitative measure of the acidity or basicity of aqueous or other liquid solutions. The term,
widely used in chemistry, biology, and agronomy, translates the values of the concentration of the
hydrogen ion—which ordinarily ranges between about 1 and 10-14 gram-equivalents per liter—into
numbers between 0 and 14 (see figures 1 and 2).
pH is defined as the decimal logarithm of the reciprocal of the hydrogen ion concentration, [H+], in a
solution:
𝑝𝐻 = −𝑙𝑜𝑔10 ([𝐻+ ])
In pure water, which is neutral (neither acidic nor alkaline), the concentration of the hydrogen ion is 10-7
gram-equivalents per liter, which corresponds to a pH of 7. Because of the logarithmic relationship
between concentration and pH, a buffer that keeps the hydrogen ion concentration within a factor of 10
of the original value will keep the pH within a range of +/- 1 pH unit.
A solution with a pH less than 7 is considered acidic; a solution with a pH greater than 7 is considered
basic, or alkaline. Common tap water is often adjusted to a pH between 7 and 8, because this
reduces the corrosion of metal pipes in the plumbing system.
Figure 1: Relationship between p[OH] and p[H] (red = acid region, blue = basic region
pH Wikipedia http://en.wikipedia.org/wiki/PH_scale (accessed 06/30/2014).
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Figure 2: pH values of some common substances.
pH Wikipedia
http://en.wikipedia.org/wiki/PH_scale (accessed 06/30/2014).
Acidosis is a condition in which there is too much acid in the body fluids.
The kidneys and lungs maintain the balance (proper pH level) of chemicals called acids and bases in the
body. Acidosis occurs when acid builds up or when bicarbonate (a base) is lost. Acidosis is classified as
either respiratory or metabolic acidosis.
Respiratory acidosis develops when there is too much carbon dioxide (an acid) in the body. This type of
acidosis is usually caused when the body is unable to remove enough carbon dioxide through breathing.
Causes of respiratory acidosis include: chest deformities, such as kyphosis, chest injuries, chest muscle
weakness, chronic lung disease and overuse of sedative drugs. Metabolic acidosis develops when too
much acid is produced in the body. It can also occur when the kidneys cannot remove enough acid from
the body. There are several types of metabolic acidosis:
Diabetic acidosis develops when substances called ketone bodies (which are acidic) build up during
uncontrolled diabetes.
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Hyperchloremic acidosis is caused by the loss of too much sodium bicarbonate from the body, which can
happen with severe diarrhea.
Lactic acidosis is a buildup of lactic acid. Different types of acidosis can be caused by cancer, renal
failure, and drinking too much alcohol, exercising vigorously for a very long time, liver failure, low blood
sugar (hypoglycemia), medications, such as salicylates, prolonged lack of oxygen from shock, heart
failure, or severe anemia and seizures.
Alkalosis is a condition in which the body fluids have excess base (alkali).
Decreased carbon dioxide (an acid) or increased bicarbonate (a base) level makes the body too alkaline,
a condition called alkalosis. There are different types of alkalosis.
Respiratory alkalosis is caused by a low carbon dioxide levels in the blood. This can be due to fever,
being at a high altitude, lack of oxygen, liver disease, lung disease, which causes you to breathe faster
(hyperventilate) and salicylate poisoning.
Metabolic alkalosis is caused by too much bicarbonate in the blood. It can also occur due to certain
kidney diseases.
Hypochloremic alkalosis is caused by an extreme lack or loss of chloride, such as from prolonged
vomiting.
Hypokalemic alkalosis is caused by the kidneys' response to an extreme lack or loss of potassium. This
can occur from taking certain water pills (diuretics).
One of the most crucial homeostatic mechanisms inside our body is the regulation of the acid-base
balance. Recall that the homeostasis is the relative constancy of the internal environment.
Any variation of the pH of blood and body fluids will have a major impact on the activity of the enzymes,
proteins, cell shape and regulation, overall metabolism and normal functioning of the cells and the body
as a whole.
Inside our body to avoid having the pathological states described above (acidosis and alkalosis), we have
buffers that resists any pH changes. They are classified in two categories:
The physiological buffer is either the urinary or the respiratory systems that acts in stabilizing our body
pH by controlling the quantity of acids, bases or CO2 we release.
The chemical buffers are substances that have the ability to bind to any excess of hydrogen ion available
in our body or release it into the fluids once its concentration fails. Because we have several substances
acting as chemical buffers and functioning together, we refer to them as the “buffer system” and they
include phosphate, proteins and bicarbonate.
In the lab experiment you will be performing, you will explore the principles of acids and bases by using
digital pH meter to accurately determine the pH of a solution and you will also test the ability of the 2
different buffers (phosphate and albumin solutions) to stabilize the pH. Phosphate ions are part of the
chemical buffer system in our bodies and albumin is a protein that can also provide buffering capability.
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References:
Encyclopædia Britannica Online
http://www.britannica.com/EBchecked/topic/83655/buffer (accessed 06/30/2014)
Encyclopædia Britannica Online
http://www.britannica.com/EBchecked/topic/454823/pH (accessed 06/30/2014)
Brent Wisse, MD, Acidosis
http://www.nlm.nih.gov/medlineplus/ency/article/001181.htm (accessed on 06/30/14)
Brent Wisse, MD, Alkalosis
http://www.nlm.nih.gov/medlineplus/ency/article/001183.htm (accessed on 06/30/14)
pH Wikipedia
http://en.wikipedia.org/wiki/PH_scale (accessed 06/30/2014).
EQUIPMENT:



Paper
Pencil/pen
Computer with Internet access (for the remote laboratory and for data analysis)
PREPARING TO USE THE REMOTE WEB-BASED SCIENCE LAB (RWLS):
For those performing a NANSLO lab for the FIRST TIME, click on this link for information on
how to install software on your computer to access the RWSL interface:
http://www.wiche.edu/nanslo/lab-tutorials.
INTRODUCTION TO THE REMOTE EQUIPMENT AND CONTROL PANEL:
Watch this short video to see how to use the RWSL control panel. (SOON TO BE CREATED)
There are appendices at the end of this document that you can refer to during your lab if you need to
remind yourself how to accomplish some of the tasks using the RWSL control panel.
EXPERIMENTAL PROCEDURE:
Once you have logged on to the remote lab system, you will perform the following laboratory
procedures.
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PRE-LAB EXERCISE 1: Adding an Acid to Buffers Solutions
In this exercise:

You will test the ability of water and buffer solutions to stabilize the pH of a solution.

You will be using 3 flasks containing each 3 different solutions: water, albumin solution, and
phosphate solution. Pure water’s pH is 7. Albumin and phosphate solutions have both a pH
value ranging between 7.00 and 7.20.

You will be adding drops of 0.1 molar hydrochloric acid (HCI) to see its impact on the variation of
the pH for each of the 3 solutions.
Pre-Lab 1 Questions:
1. Using what you know about buffers and water, predict how each solution will perform as a
buffer.
2. Rewrite your answer to question 1 in the form of an If … Then … hypothesis.
PRE-LAB EXERCISE 2: Adding a Base to Buffers Solutions
In this exercise:

You will test the ability of water and buffer solutions to stabilize the pH of a solution.

You will be using 3 flasks containing each 3 different solutions: pure water, albumin solution,
and phosphate solution. Pure water’s pH is 7. Albumin and phosphate solutions have both a pH
value ranging between 7.00 and 7.20.

You will be adding drops of the sodium hydroxide (NaOH) to see its impact on the variation of
the pH for each of the 3 solutions.
Pre-Lab 1 Questions:
1. Using what you know about buffers and water, predict how each solution will perform as a
buffer.
2. Rewrite your answer to question 1 in the form of an If … Then … hypothesis.
EXERCISE 1: Adding an Acid to Buffers Solutions
DATA COLLECTION:
1. In the flask containing water: Using the digital pH meter, measure the initial pH and record
your data in a table.
2. Add 1 drop of the hydrochloric acid (HCI) solution and measure the new pH of the solution.
Record your data in a table.
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3. Repeat step 2 nine additional times.
Rinse the pH meter probe.
4. In the flask containing albumin solution: Using the digital pH meter, measure the initial pH and
record your data in table 2 below.
5. Add 1 drop of the hydrochloric acid (HCL) solution and measure the new pH of the solution.
Record your data in a table.
6. Repeat step 5 nine additional times.
Rinse the pH meter probe.
7. In the flask containing phosphate buffer solution: Using the digital pH meter, measure the
initial pH and record your data in Table 3 below.
8. Add 1 drop of the hydrochloric acid (HCL) solution and measure the new pH of the solution.
Record your data in a table.
9. Repeat step 8 nine additional times.
Rinse the pH meter probe.
ANALYSIS:
10.
11.
12.
13.
14.
Compare the final pH of each solution. Which one has “absorbed” the HCI most efficiently?
Which solution is the most effective buffer?
Which solution is the least effective buffer?
Rewrite your hypothesis in light of your new information collected in this exercise.
Graph the variation of the pH vs the drops of HCI and estimate the point at which buffer
stabilization fails.
EXERCISE 2: Adding a Base to Buffers Solutions
DATA COLLECTION:
1. In the flask containing water: Using the digital pH meter, measure the initial pH and record
your data in a table.
2. Add 1 drop of the sodium hydroxide (NaOH) solution and measure the new pH of the solution.
Record your data in a table.
3. Repeat step 2 nine additional times.
Rinse the pH meter probe.
4. In the flask containing albumin solution: Using the digital pH meter, measure the initial pH and
record your data in a table.
5. Add 1 drop of the sodium hydroxide (NaOH) solution and measure the new pH of the solution.
Record your data in table 5 below.
6. Repeat step 5 nine additional times.
Rinse the pH meter probe.
7. In the flask containing phosphate buffer solution: Using the digital pH meter, measure the
initial pH and record your data in a table 6 below.
8. Add 1 drop of the sodium hydroxide (NaOH) solution and measure the new pH of the solution.
Record your data in a table.
9. Repeat step 8 nine additional times.
Rinse the pH meter probe.
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ANALYSIS:
10.
11.
12.
13.
14.
Compare the final pH of each solution. Which one has “absorbed” the NaOH most effectively?
Which solution is the most effective buffer?
Which solution is the least effective buffer?
Rewrite your hypothesis in light of your new information collected in this exercise.
Graph the variation of the pH versus the drops of NaOH and estimate the point at which buffer
stabilization fails.
SUMMARY QUESTIONS
1. Based on the experiment you just completed above, how do you explain the difference in the
buffering capacities between the solutions you tested?
2. Based on your understanding of the pH scale, calculate the hydrogen ion concentration of a
solution:
a.
with a pH 7.0?
b. with a pH 9.0
c. with a pH of 4.0?
3. The pH of blood in the human body is between 7.35 and 7.45. Discuss the benefits of healthy eating
and exercises in maintaining homeostasis.
4. You are working in an independent laboratory and you want to investigate the effectiveness of
different commercial antacids such as Tums, Maalox, Alka-Seltzer in neutralizing the hydrochloric
acid produced by the stomach. How will you conduct your investigation?
5. Explain the role of the respiratory system in maintaining normal blood pH?
6. Research the buffering activity of hemoglobin. Describe what you find to the best of your ability.
For more information about NANSLO, visit www.wiche.edu/nanslo.
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