DEPARTMENT OF PURE AND APPLIED CHEMISTRY

Visayas State University, Baybay, Leyte

CHEM31a Biochemistry
Laboratory Report

Name
: Mark Ryan R. Tripole
Course/Yr : BS Chemistry II
Group No : 6

Date Performed : 05/25/2015

Date Submitted : 06/01/2015
Score

Experiment No. 9
Chemistry of Urine
OBJECTIVES
Test urine for pH, specific gravity, and the presence of electrolytes and organic
compounds.
Test urine for the presence of abnormally occurring compounds of protein, glucose and
ketone bodies

I. Results

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Shown below is the tabulated data for a sample of urine that has been analyzed at pH 5.5. The data
does not correlate to the results in the experimental procedure and is only used as an example.

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II. Discussion
INTRODUCTION
Simply put, urine is a watery excretion that is composed of waste products that the body
doesnt really need or hasnt had the chance to reabsorb. Some might actually see urine as a sort of
expulsion system for the excess water that is in the body, which is true considering that the majority
of the composition of urine is water (around 95 to 96%). But the main and important consideration is
the fact that urine aids in the dumping of chemicals, dead blood cells and more from the body. So
essentially urine is composed of many components with water serving as the medium that carries the
waste out of the body, and it is these that make up the other 4 to 5%. This laboratory experiment
was intended for the observation of the composition of urine, the interpretation of the amount of
each of the individual components of each sample accordingly, and how the urine samples from each
individual differ from each other depending on certain parameters (illness, etc.). As a guide, urine is
considered to have quite a large amount of urea, followed by the electrolytes like chlorides and
sodium. Urine also contains compounds like uric acid, ketone bodies, proteins and glucose. The
quantitative testing of these compounds can lead to medial diagnosis depending on circumstances,
but this laboratory experiment only goes so far as qualitative analysis. Each part will be discussed
accordingly in the following sections of the laboratory report.
PART A COLOR, pH AND SPECIFIC GRAVITY
Color - The color of urine pretty much varies depending on how hydrated the person is, but this can
also more or less be affected by a whole range of different factors. Normally, urine is a
colorless solution that can range from colorless to an amber hue. But there are other factors
that can cause abnormal coloration of the urine, an example being diseases (melanoma
producing dark or black urine) or merely cosmetic coloration from the consumption of
certain foodstuffs (like beets producing pinkish colored urine).
pH The pH of urine usually falls between the ranges of 4.6 to 8, with 7 being the normal pH
level for urine. The pH variations can occur through many ways, but the two of which are
consumption of certain food as well as intake of medications. Consumption of legumes and
high citrus diets have been known to increase urine pH, while diets that are fixed on meat
tend to lower pH levels of the urine.
Spg By itself, specific gravity is basically a measure of how much more or less dense a particular
liquid is compared to water. Using this parameter in conjunction with urine analysis, it is an
important quantitative variable that is commonly used in the evaluation of kidney function
and help to diagnose renal diseases. The normal specific gravity of urine lies between 1.003
and 1.030. Going over or falling short of these limits are indications of various abnormalities,
with increased specific gravity being attributed to UTIs and decreased specific gravity
being attributed to renal failure.
Looking at the sample of urine that was donated in the laboratory experiment, the color was that of
a pale yellowish color, indicating normal level of hydration. The urine sample showed no turbidity
which would have been another indication of an abnormality. The pH of the urine as tested with pH
paper was found to be within the range of 6-7, indicating normal pH ranges typical of normal urine.
The specific gravity of the urine sample was measured using a refractometer which showed a rating
of 3% brix which converted is approximately 1.?????. This falls within the normal range of 1.003 and
1.030, and so there are no problems with the urine sample.

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PART B - UREA
Urea exists in quite a high concentration in the urine because it is the waste product that is
produced from the breakdown of proteins in the body. So this makes urea one of the components of
urine that will always be found in any sample. Since the laboratory procedure only involved a
qualitative method for analysis, so only the physical characteristic of the testing was observed. The
reaction basically involved reacting a sample of the urine with dilute sodium hydroxide (in the place
of urease which was unavailable at the time) and leaving it to stand for at least one hour before it
was heated. When it was heated, a piece of red litmus paper was placed on the mouth of the test
tube containing the urine, and the conversion from red to blue was an indication of the presence of
gaseous ammonia, and thus the presence of urea as a component of urine. A simple reaction
equation for this reaction is shown below:
2NaOH
+
Sodium hydroxide

(NH2)2CO
+ urea

2NH3
+
Na2CO3
ammonia (g) + sodium carbonate

By quantitative standards, the standard amount of urea in urine usually ranges between 12 to
20 grams within a 24 hour period. Abnormal values for these are attributed to diseases, with low
levels being an indication of kidney problems, and higher levels indicating a substantial increase of
protein breakdown in the body.
PART C URIC ACID
Uric is basically one of those rare waste products that are excreted in the urine but in only a
very small amount, at least by normal standards. It is actually considered as one of the penultimate
products of the tissue waste in the human body, being the waste product from the breaking down of
purine nucleotides. In simpler terms, uric acid is produced from the natural breakdown of the bodys
cells as well as the food consumed. Most of the uric acid as is is removed from the body in urine, and
a small amount of it passes along in the stool. But certain physical ailments can actually hinder the
removal of uric acid from the blood which can lead to symptoms such as gout, etc. The test
performed for this part of the experiment was simply taking a sample of the urine and adding to it
some concentrated hydrochloric acid. The beaker with the sample and HCl mixture was then left
covered with a plastic wrap for a day and then observed for the formation of any crystals of uric acid.
This basically works on the principle that uric acid is not soluble at extremely acidic pH levels, which
in this case would be bought about by the addition of the concentrated hydrochloric acid.
The groups sample showed no formation of any crystals of uric acid, indicating a low or close
to no amount of uric acid present in the urine. This might be an indication of kidney problems,
meaning that the kidneys arent able to get rid of the uric acid well enough. But this cant be
properly observed with only qualitative testing, since the actual quantification of uric acid over a 24
hour period and proper parameters is required for actual diagnosis.
PART D - ELECTROLYTES
In basic terms, an electrolyte is any substance that contains free ions that behaves as an
electrically conductive medium. All higher life forms cannot exist without electrolytes, and that
includes human beings. But then these only exist in a constant concentration throughout the body
and are constantly replaced by the consumption of food or supplements. Any of the excess
electrolytes are then filtered out by the kidneys and particularly out through the urine. To keep it
simple, electrolytes are important substances because they aid in the regulation of nerve and muscle
function, the hydration of the body, blood pH levels, rebuilding of damaged tissue and blood
pressure. For the experimental procedure, some of the more common electrolytes were tested for,
and they shall be discussed below. Also in reference to the data obtained, the urine sample used
tested positive for the presence of the ions to be discussed.
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1. Chlorides
Considered as one of the important electrolytes in blood by helping in the regulation of
concentration of fluid inside and outside of the cells. The bulk of the chlorides come from the
intake of salt (sodium chloride) and are mostly absorbed by the intestines. The excess leaves the
body through the urine. The test involved the addition of a solution of silver nitrate to the
sample of urine that has been acidified with nitric acid that would ultimately result in the
formation of a white precipitate. The acidification serves the purpose of preventing false
positives, because silver pairs up with carbonates to form a precipitate that is also white in color.
A simple equation of this reaction is shown below:
Cl- + AgNO3

AgCl (s) + NO3-

2. Sulfates
Sulfates are essentially to the body because all cells require inorganic sulfate for normal
function. Sulfates are considered to be among the most important macronutrients in cells, being
the major source of sulfur in many living organisms. Despite the fact that sulfate itself doesnt
possess a catalytic function or a role in human energy metabolism, there is much evidence to
suggest that sulfates are not metabolically inert molecules and play an essential role in life. The
qualitative test for this in urine is also through the use of the solubility rules, by adding in a
solution of barium chloride to acidified urine. The equation for this reaction is shown below:
SO42- + BaCl2

BaSO4 (s) + 2Cl-

3. Phosphates
Phosphates are important because they aid in building and repairing bones and teeth, aid in
muscle contraction and helps nerve function. Much like the other electrolytes, the phosphates
are regulated by the kidneys and any excess amounts are excreted through the urine. This was
tested in the experimental procedure through reaction of the acidified urine sample with
ammonium molybdate, which produced a yellow precipitate indicating the presence of
phosphates. The ammonium molybdate ((NH4)2MoO4) forms a precipitate of ammonium
phosphomolybdate ((NH4)3PO4.12MoO4) which is a bright yellow compound that is extremely
insoluble even in dilute nitric acid.
4. Sodium and Potassium
These two cations are basically considered to be the power generators inside the cells of the
body, and they are invaluable in the process of neural transmission. The presence of the two
electrolytes in urine can be tested for qualitatively through the use of a simple flame test. A
flame test wire was dipped into some hydrochloric acid and heated to red hot in the flame of a
Bunsen burner. This was done to prevent any form of false positives. The wire was then placed
into the urine solution and then heated. The formation of a yellow flame indicated the presence
of sodium and the formation of a discernable red flame as seen through a cobalt glass square
indicated the presence of potassium in the urine sample.
5. Calcium
This particular electrolyte plays a vital role in signal transduction pathways (activation of
receptors by an external signaling molecule that can alter the cells metabolism, etc.). Calcium
also plays an important role in the contraction of all muscle cell types and in some other
important processes such as fertilization. With regard to the experimental procedure, this
electrolyte was test for on the basis of the solubility rules, where a urine sample was acidified
with acetic acid and then reacted with ammonium oxalate. The formation of a white precipitate
of calcium oxalate indicated a positive result for the urine sample. Shown below is a simple
equation for the reaction:
Ca2+ + (NH4)2C2O4

CaC2O4 + 2NH4+

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PART E - GLUCOSE
Glucose is a compound that usually not found in urine because of the normal capability of the
kidneys to reclaim all of the filtered glucose back into the blood stream. The presence of glucose in
the urine is basically condition that is known as glucosuria, which is a condition caused by elevated
blood glucose levels that are brought about by untreated diabetes milletus. A simple and functional
test for this would be the Benedicts test for reducing sugars. Based on the laboratory results, the
urine sample used didnt change much from the original blue color it had when mixed with the
Benedicts reagent, which indicates a low level or close to none depending of the amount of cupric
oxide formed as necessary. A simple illustration of this reaction (taking into consideration a urine
sample that has a high amount of glucose) is shown below:

PART F KETONE BODIES

Ketone bodies is the collective name used for three different compounds that are produced
by the liver from fatty acids during low periods of food intake (fasting) (which are acetone,
acetoacetic acid and -hydroxy butyric acid. Simply put, these are substances that are made when
the body breaks down fat for energy. The body normally gets energy from the carbohydrates
obtained in the diet, but if there is not enough supply of glucose in the body, the fats are used
instead, and these are excreted in the urine. In normal urine, ketone bodies will not be present. In
the experimental procedure, the urine sample tested positive for ketone bodies, which could be an
indication of a medical condition known as ketonuria. The test for this particular compound in the
urine is known as the Rotheras Test which is a qualitative test to test for ketone bodies. The urine
sample was saturated with ammonium sulfate in order to concentrate the ketone bodies to the
center of the solution and also has an added function of the ammonium ions being able to precipitate
any proteins that might be present (false positives). Nitroprusside was then added to this mixture,
followed by the gradual addition of ammonium hydroxide or ammonia solution. The formation of a
purple ring at the junction between the urine and ammonia/ammonium hydroxide solution. The
purple ring is a product of the complex between the ketone and the nitroprusside in the presence of
ammonia.
PART G - PROTEINS
In terms of the presence of proteins in urine, healthy individuals usually dont have any
protein in their urine, which is facilitated through the regulation of the proteins by the kidneys,
which basically ensure that they remain in the blood stream. A small amount of proteins is usually
not much for concern, but high amounts of protein in urine are indications of kidney failure, diabetes
or urinary tract infection. For the laboratory experiment, the upper portion of the urine sample was
heated, and with the initial formation of turbidity 5 drops of acetic acid was added (these are false
positives). The heating of the same upper portion a second time showed the evolution of a faint
turbidity which is an indication of a small and trace amount of protein in the sample. This can be
bought about by exercise and other normal activities.

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III. Conclusion

IV. References

"Uric Acid"
http://www.mayomedicallaboratories.com/articles/hottopics/transcripts/2010/2010-2akidney-stones/2a-23.html
"Chlorides in Urine"
http://www.webmd.com/a-to-z-guides/chloride-cl
"Qualitative Analysis of Urine"
http://physrev.physiology.org/content/81/4/1499
"Urine Specific Gravity"
http://en.wikipedia.org/wiki/Urine_specific_gravity
"Hydrolysis of Urea"
http://www.sciencemadness.org/talk/viewthread.php?tid=19575
"What happens when you add sodium hydroxide to urea"
http://www.scienceforums.net/topic/38367
"Urea Nitrogen Urine Test"
http://www.nlm.nih.gov/medlineplus/ency/article/003605.htm
"Uric Acid in Urine"
http://www.webmd.com/a-to-z-guides/uric-acid-in-urine
"Benedict's Test Urine"
http://www.doctorslounge.com/endocrinology/forums/backup/topic-2562
"Ketone Bodies"
http://en.wikipedia.org/wiki/Ketone_bodies
"What a Urine Glucose Test Means"
http://www.nlm.nih.gov/medlineplus/ency/article/003581.htm
"Urine Tests"
http://edusanjalbiochemist.blogspot.com/2013/01/urinalysis-chemical
"Urine"
http://en.wikipedia.org/wiki/Urine
"Protein Urine Test"
http://www.healthline.com/health/protein-electrophoresis-serum#Overview1
"Urinalysis"
library.med.utah.edu/WebPath/TUTORIAL/URINE/URINE.html

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