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RENAL PHYSIOLOGY
URINE FORMATION


LABORATORY:

URINALYSIS
The Urinary System
Organs of the urinary system:
 Kidneys
 Ureters
 Urinary bladder
 Urethra
Kidney function - removes waste products
  from the blood stream and excess water

        Urinary bladder stores urine, and ureters,
        bladder and urethra make up the urinary tract 3
The Urinary System

Urethra                             Aorta

Kidneys                             Liver

Urinary bladder                     Left Ureter

Right Ureter                        Prostate gland




                                                  4
Formation of Urine
Three processes           The nephron:
 of urine                  allows for
 formation:                 reabsorption of
 glomerular filtration
                            water and
 tubular reabsorption
                            electrolytes
 tubular secretion        plays a vital role
                            in maintaining
                            normal fluid
                            balance
                                                 5
Nephron
- functional unit of the kidney where urine is
   formed.


 STAGES IN URINE FORMATION:
 1.Filtration
 2.Reabsorption
 3.Secretion
Secretion
Nephron
                       Reabsorption

          filtration




                          Reabsorption
                          of water
1. filtration


            -blood
                 pressure forces
              small molecules
                 from the
          glomerulus to the capsule



                      Filtrates:
                glucose, amino acids
                    uric acid, urea
2. Tubular Reabsorption

            -return of filtrates
                from blood
          at the proximal tubule
             through diffusion
           and active transport
3. Tubular Secretion


            -movement of molecules
                    from blood
                      into the
             distal convoluted tubule


                  Molecules:
               drugs and toxins
Reabsorption of water

                 -return  of H20
                   via osmosis
                    along the
                loop of Henle and
                  collecting duct
EXCRETION




      excretion of urine formed
PATHWAY OF URINE
Nephron
- functional unit of the kidney where urine is
   formed.


 STAGES IN URINE FORMATION:
 1.Filtration
 2.Reabsorption
 3.Secretion
Urinalysis
Specific Objectives:
            Physical Characteristics
            Normal pH
            Specific Gravity ranges
            Presence of abnormal substances
                in the urine specimens
Physical Composition and Chemical
Properties
Urine
 95% water
 5% waste products
 Other dissolved chemicals




   Urea, uric acid, ammonia, calcium, creatine,
   sodium, chloride, potassium, sulfates,
   phosphates, bicarbonates, hydrogen ions,
   urochrome, urobilinogen
                                                  16
Urinalysis
             • a physical and/or chemical
               examination of the urine.
             • It consists of a battery of chemical and
               microscopic tests to screen for:
                • urinary tract infections
                • renal disease, and
                          disease
                • diseases of other organs that result
                   in the appearance of abnormal
                   metabolites (break-down
                   products) in the urine.
Urinalysis
             • can reveal diseases that have gone unnoticed
               because they do not produce striking signs or
               symptoms.
                • Examples include diabetes mellitus,
                  various forms of glomerulonephritis, and
                  chronic urinary tract infections.
             • In other urinary diseases with symptoms, a
               urinalysis can help to confirm of rule out some
               diseases.
TESTS

• Qualitative Tests for Normal Constituents
   • Urea Test
   • Uric Acid
   • Indican Test (Decompositon of
     Tryptophan)
• Qualitative Tests for Pathological Constituents
   •   Gunning’s Test (Ketone)
   •   Benedict's Test (Glucose)
   •   Exton’s Test (Albumin)
   •   Smith’s Test (Bile pigments)
SPECIMEN COLLECTION
       • A "clean-catch" urine sample is
         performed by collecting the sample
         of urine in midstream.
       • Men or boys should wipe clean the
         head of the penis.
       • Women or girls need to wash the
         area between the lips of the vagina
         with soapy water and rinse well.
       • A small amount of urine should
         initially fall into the toilet bowl before
         it is collected (this clears the urethra
         of contaminants).
       • Then, in a clean container (properly
         labeled with the name of the patient
         and date), catch about 1 to 2
         ounces of urine and remove the
         container from the urine stream.
Specimens Types
Varies in method used and in
   time frame in which to collect
   specimen
Types of specimens:
 Random
 First morning
 Clean catch midstream
 Timed
 24 hour
                                    21
Specimens Types
(cont.)    Random – most common, taken
            anytime of day
           First morning – has a greater
            concentration of substances, taken
            in morning
           Clean catch midstream – genitalia is
            cleaned, urine is tested for
            microorganisms or presence of
            infection
           Timed – specific time of day, always
            discard first specimen before timing
           24 hour – used for quantitative and
            qualitative analysis of substances 22
SPECIMEN COLLECTION
      • For an infant, thoroughly wash the area
        around the urethra.
      • Open a urine collection bag (a plastic bag
        with an adhesive paper on one end), and
        place it on the genital area.
      • For males, the entire penis can be placed
        in the bag and the adhesive attached to
        the skin.
      • For females, the bag is placed over the
        labia majora. Place a diaper, check the
        infant frequently and remove the bag after
        the infant has urinated into it.
      • The urine is then drained into a labeled
        container.
Urinalysis
Evaluation of urine to obtain information
  about body health and disease
Three types of testing:
 Physical
 Chemical
 Microscopic




                                            24
Physical Examination of
Urine
Visual examination of physical
  characteristics
 Color and turbidity
 Volume
 Odor
 Specific gravity
    The refractometer or a reagent
     strip is used to measure
     specific gravity
                                              Refractometer

                             Reagent Strips
                                                              25
MACROSCOPIC URINALYSIS
         • Clearing of the specimen after addition
           of a small amount of acid indicates that
           precipitation of salts is the probable
           cause of turbidity.
         • A red or red-brown (abnormal)
           color could be from a food dye,
           eating fresh beets, a drug, or the
           presence of either hemoglobin or
           myoglobin.
         • If the sample contained many red
           blood cells, it would be cloudy as
           well as red.
MACROSCOPIC
URINALYSIS
              • The first part of a urinalysis is direct
                visual observation. Normal, fresh
                urine is pale to dark yellow or
                amber in color and clear.
              • Normal urine volume is 750 to
                2000 ml/24hr.
              • Turbidity or cloudiness may be
                caused by excessive cellular
                material or protein in the urine or
                may develop from crystallization or
                precipitation of salts upon standing
                at room temperature or in the
                refrigerator.
Normal Values of Urine
 Normal values of various
  elements have been
  established
 Average adult daily urine
  output is 1250 mL/24
  hours
 Intake and output should
  be approximately the
  same
                              28
Examples of appearances of urine




a red, cloudy appearance   red but clear   yellow, but cloudy
• The glomerular filtrate of blood
       plasma is usually acidified by renal
pH     tubules and collecting ducts from a
       pH of 7.4 to about 6 in the final
       urine.
     • However, depending on the acid-
       base status, urinary pH may range
       from as low as 4.5 to as high as
       8.0.
       8.0
       • The change to the acid side of 7.4 is
         accomplished in the distal
         convoluted tubule and the
         collecting duct.
• (which is directly proportional to
             urine osmolality which measures
Specific     solute concentration) measures
             urine density, or the ability of the
gravity      kidney to concentrate or dilute the
             urine over that of plasma.
           • Specific gravity between 1.002 and
             1.035 on a random sample should
             be considered normal if kidney
             function is normal.
             • Since the sp gr of the glomerular filtrate
               in Bowman's space ranges from 1.007
               to 1.010, any measurement below this
                  1.010
               range indicates hydration and any
               measurement above it indicates
               relative dehydration.
                        dehydration
Specific Gravity (sp gr)
         • If sp gr is not > 1.022 after a 12 hour
           period without food or water
            • renal concentrating ability is
              impaired
            • In end-stage renal disease, sp gr
              tends to become 1.007 to 1.010.
         • Any urine having a specific gravity over
           1.035 is
            • either contaminated,
            • contains very high levels of glucose
Apply Your Knowledge
What is the specific gravity shown on this
refractometer screen?




                                             34
-
Apply Your Knowledge Answer
What is the specific gravity shown on
this refractometer screen?


                     The specific
                      gravity
                      shown
                      here is
                      1.030
                                        35
Chemicals Found in
Urine
 Ketone bodies – may indicate that patient is following
  a low carbohydrate diet or that patient has conditions
  such as starvation
 pH – provides information about metabolic status, diet,
  medication or several conditions
 Blood (hematuria) – may indicate patient is
  menstruating, have urinary tract infection or trauma




                                                            36
Chemicals Found in
Urine (cont.)
      Bilirubin or urobilinogen – first signs
           of liver disease
      Glucose – common in patients with
           diabetes
      Protein – usually indicates renal
           disease
      Nitrite – suggests bacterial infection
      Leukocytes – urinary or renal
           infection
      Phenylketones – indicates PKU
                                                37
Microscopic Examination
of Urine
 Microscopic examination used to view elements not
  visible without microscope
 Centrifuge spins urine to cause heavier substances to
  settle to the bottom


              • Cells        • Casts

              • Crystals     • Yeasts
                             • Parasites
              • Bacteria

                                                          38
Glucose test
       • Benedict’s test
         • Aqueous glucose is mixed with
           Benedict's reagent, a solution of
           copper sulfate, sodium hydroxide, and
           tartaric acid.
         • The mixture is heated.
         • Carbohydrates which react with
           Benedict's reagent to reduce the blue
           copper (II) ion to form a brick red
           precipitate of copper (I) oxide are
           classified as reducing sugars.
Results of Benedict's Test with Glucose,
Sucrose, and Fructose
Glucose

• Less than 0.1% of glucose normally
  filtered by the glomerulus appears in urine
  (< 130 mg/24 hr).
• Glycosuria (excess sugar in urine)

          diabetes
  generally means

  mellitus.
• Exton’s Test
             • There are several acids
Protein        which can be used to
               precipitate proteins -
               sulfosalicylic,
               trichloroacetic, nitric, and
               acetic acids.
          • Sulfosalicylic acid (SSA) is
            the most frequently used acid
            test because it does not
            require the use of heat.
             • Exton’s reagent is 5%
               sulfosalicylic acid in a
To perform the SSA procedure mix equal parts of patient urine and the reagent. Rate the

amount of turbidity according to the following scale:
Interpretation
            • In rough terms, trace positive
              results (which represent a slightly
              hazy appearance in urine) are
              equivalent to 10 mg/100 ml or
              about 150 mg/24 hours (the upper
              limit of normal).
            • 1+ corresponds to about 200-500
              mg/24 hours
            • a 2+ to 0.5-1.5 gm/24 hours
            • a 3+ to 2-5 gm/24 hours
            • a 4+ represents 7 gm/24 hours or
              greater.
• Normally, only small plasma
Interpretation   proteins filtered at the glomerulus
                 are reabsorbed by the renal
                 tubule.
               • Normal total protein excretion
                 does not usually exceed 150
                 mg/24 hours or 10 mg/100 ml in
                 any single specimen.
               • More than 150 mg/day is
                 defined as proteinuria.
               • Proteinuria > 3.5 gm/24 hours
                 is severe and known as
                 nephrotic syndrome.
Apply Your Knowledge
    A urine analysis has
    detected that a patient has
    protein in his urine. Why is
    this important?




                                   46
-
Apply Your Knowledge Answer

A urine analysis has detected that a
patient has protein in his urine. Why
is this important?

Protein in the urine usually indicates
renal disease


                                         47
Ketones
Gunning’s Test
• Ketones (acetone, aceotacetic acid, beta-
  hydroxybutyric acid) resulting from either
  diabetic ketosis or some other form of
  calorie deprivation (starvation)
• Shows positive test for:
acetaldehyde and methyl ketones
Ketones
Gunning’s Test
• Detection of COOH group is possible
  through its transformation to an ester in
  reaction with alcohol.
• The esters have characteristic smell of
  flowers or fruits
• It should appear the yellow precipitate of
  iodoform with characteristic smell
• Ketones Normal:There are no ketones in
  your blood or urine.
• Abnormal:Ketones are present in your
  blood or urine.
• . Indications of a positive test:
   The formation of a yellow precipitate or suspension of
   iodoform is a positive test




          a negative test (left) and a positive test (right)
Smith’s Test (Bile pigments)
emerald green at the point of contact between urine sample
and alcoholic iodine mixture




        Bilirubin – green
        Bilicyanine – blue
        Choletelin - yellow
• Smith’s Test should be conducted to
  determine whether bilirubin is present in the
  urine
  (i.e., bilirubinuria).
• Urobilinogen is formed by intestinal bacteria
  from the breakdown of conjugated bilirubin,
  and it is usually excreted in feces.
• However a small amount may be
  reabsorbed and excreted in urine.
• A positive test response indicates normal
  enterohepatic circulation of biliary
  pigments. High concentrations of biliary
  pigments may occur in hemolytic crisis, or
  cases of hepatic or intestinal dysfunction.
56
EXPERIMENT 27….
FIRST PHASE:

• Collect urine sample from a donor from
  each group

SECOND PHASE:

 • Unknown sample will be tested
Record all results and analysis in
your lab work sheet.
• Post lab group discussion upon
  submission of all reports will be conducted.

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Kidney urine

  • 2.
  • 3. The Urinary System Organs of the urinary system:  Kidneys  Ureters  Urinary bladder  Urethra Kidney function - removes waste products from the blood stream and excess water Urinary bladder stores urine, and ureters, bladder and urethra make up the urinary tract 3
  • 4. The Urinary System Urethra Aorta Kidneys Liver Urinary bladder Left Ureter Right Ureter Prostate gland 4
  • 5. Formation of Urine Three processes The nephron: of urine  allows for formation: reabsorption of  glomerular filtration water and  tubular reabsorption electrolytes  tubular secretion  plays a vital role in maintaining normal fluid balance 5
  • 6. Nephron - functional unit of the kidney where urine is formed. STAGES IN URINE FORMATION: 1.Filtration 2.Reabsorption 3.Secretion
  • 7. Secretion Nephron Reabsorption filtration Reabsorption of water
  • 8. 1. filtration -blood pressure forces small molecules from the glomerulus to the capsule Filtrates: glucose, amino acids uric acid, urea
  • 9. 2. Tubular Reabsorption -return of filtrates from blood at the proximal tubule through diffusion and active transport
  • 10. 3. Tubular Secretion -movement of molecules from blood into the distal convoluted tubule Molecules: drugs and toxins
  • 11. Reabsorption of water -return of H20 via osmosis along the loop of Henle and collecting duct
  • 12. EXCRETION excretion of urine formed
  • 14. Nephron - functional unit of the kidney where urine is formed. STAGES IN URINE FORMATION: 1.Filtration 2.Reabsorption 3.Secretion
  • 15. Urinalysis Specific Objectives:  Physical Characteristics  Normal pH  Specific Gravity ranges  Presence of abnormal substances in the urine specimens
  • 16. Physical Composition and Chemical Properties Urine  95% water  5% waste products  Other dissolved chemicals Urea, uric acid, ammonia, calcium, creatine, sodium, chloride, potassium, sulfates, phosphates, bicarbonates, hydrogen ions, urochrome, urobilinogen 16
  • 17. Urinalysis • a physical and/or chemical examination of the urine. • It consists of a battery of chemical and microscopic tests to screen for: • urinary tract infections • renal disease, and disease • diseases of other organs that result in the appearance of abnormal metabolites (break-down products) in the urine.
  • 18. Urinalysis • can reveal diseases that have gone unnoticed because they do not produce striking signs or symptoms. • Examples include diabetes mellitus, various forms of glomerulonephritis, and chronic urinary tract infections. • In other urinary diseases with symptoms, a urinalysis can help to confirm of rule out some diseases.
  • 19. TESTS • Qualitative Tests for Normal Constituents • Urea Test • Uric Acid • Indican Test (Decompositon of Tryptophan) • Qualitative Tests for Pathological Constituents • Gunning’s Test (Ketone) • Benedict's Test (Glucose) • Exton’s Test (Albumin) • Smith’s Test (Bile pigments)
  • 20. SPECIMEN COLLECTION • A "clean-catch" urine sample is performed by collecting the sample of urine in midstream. • Men or boys should wipe clean the head of the penis. • Women or girls need to wash the area between the lips of the vagina with soapy water and rinse well. • A small amount of urine should initially fall into the toilet bowl before it is collected (this clears the urethra of contaminants). • Then, in a clean container (properly labeled with the name of the patient and date), catch about 1 to 2 ounces of urine and remove the container from the urine stream.
  • 21. Specimens Types Varies in method used and in time frame in which to collect specimen Types of specimens:  Random  First morning  Clean catch midstream  Timed  24 hour 21
  • 22. Specimens Types (cont.)  Random – most common, taken anytime of day  First morning – has a greater concentration of substances, taken in morning  Clean catch midstream – genitalia is cleaned, urine is tested for microorganisms or presence of infection  Timed – specific time of day, always discard first specimen before timing  24 hour – used for quantitative and qualitative analysis of substances 22
  • 23. SPECIMEN COLLECTION • For an infant, thoroughly wash the area around the urethra. • Open a urine collection bag (a plastic bag with an adhesive paper on one end), and place it on the genital area. • For males, the entire penis can be placed in the bag and the adhesive attached to the skin. • For females, the bag is placed over the labia majora. Place a diaper, check the infant frequently and remove the bag after the infant has urinated into it. • The urine is then drained into a labeled container.
  • 24. Urinalysis Evaluation of urine to obtain information about body health and disease Three types of testing:  Physical  Chemical  Microscopic 24
  • 25. Physical Examination of Urine Visual examination of physical characteristics  Color and turbidity  Volume  Odor  Specific gravity  The refractometer or a reagent strip is used to measure specific gravity Refractometer Reagent Strips 25
  • 26. MACROSCOPIC URINALYSIS • Clearing of the specimen after addition of a small amount of acid indicates that precipitation of salts is the probable cause of turbidity. • A red or red-brown (abnormal) color could be from a food dye, eating fresh beets, a drug, or the presence of either hemoglobin or myoglobin. • If the sample contained many red blood cells, it would be cloudy as well as red.
  • 27. MACROSCOPIC URINALYSIS • The first part of a urinalysis is direct visual observation. Normal, fresh urine is pale to dark yellow or amber in color and clear. • Normal urine volume is 750 to 2000 ml/24hr. • Turbidity or cloudiness may be caused by excessive cellular material or protein in the urine or may develop from crystallization or precipitation of salts upon standing at room temperature or in the refrigerator.
  • 28. Normal Values of Urine  Normal values of various elements have been established  Average adult daily urine output is 1250 mL/24 hours  Intake and output should be approximately the same 28
  • 29.
  • 30. Examples of appearances of urine a red, cloudy appearance red but clear yellow, but cloudy
  • 31. • The glomerular filtrate of blood plasma is usually acidified by renal pH tubules and collecting ducts from a pH of 7.4 to about 6 in the final urine. • However, depending on the acid- base status, urinary pH may range from as low as 4.5 to as high as 8.0. 8.0 • The change to the acid side of 7.4 is accomplished in the distal convoluted tubule and the collecting duct.
  • 32. • (which is directly proportional to urine osmolality which measures Specific solute concentration) measures urine density, or the ability of the gravity kidney to concentrate or dilute the urine over that of plasma. • Specific gravity between 1.002 and 1.035 on a random sample should be considered normal if kidney function is normal. • Since the sp gr of the glomerular filtrate in Bowman's space ranges from 1.007 to 1.010, any measurement below this 1.010 range indicates hydration and any measurement above it indicates relative dehydration. dehydration
  • 33. Specific Gravity (sp gr) • If sp gr is not > 1.022 after a 12 hour period without food or water • renal concentrating ability is impaired • In end-stage renal disease, sp gr tends to become 1.007 to 1.010. • Any urine having a specific gravity over 1.035 is • either contaminated, • contains very high levels of glucose
  • 34. Apply Your Knowledge What is the specific gravity shown on this refractometer screen? 34
  • 35. - Apply Your Knowledge Answer What is the specific gravity shown on this refractometer screen? The specific gravity shown here is 1.030 35
  • 36. Chemicals Found in Urine  Ketone bodies – may indicate that patient is following a low carbohydrate diet or that patient has conditions such as starvation  pH – provides information about metabolic status, diet, medication or several conditions  Blood (hematuria) – may indicate patient is menstruating, have urinary tract infection or trauma 36
  • 37. Chemicals Found in Urine (cont.) Bilirubin or urobilinogen – first signs of liver disease Glucose – common in patients with diabetes Protein – usually indicates renal disease Nitrite – suggests bacterial infection Leukocytes – urinary or renal infection Phenylketones – indicates PKU 37
  • 38. Microscopic Examination of Urine  Microscopic examination used to view elements not visible without microscope  Centrifuge spins urine to cause heavier substances to settle to the bottom • Cells • Casts • Crystals • Yeasts • Parasites • Bacteria 38
  • 39. Glucose test • Benedict’s test • Aqueous glucose is mixed with Benedict's reagent, a solution of copper sulfate, sodium hydroxide, and tartaric acid. • The mixture is heated. • Carbohydrates which react with Benedict's reagent to reduce the blue copper (II) ion to form a brick red precipitate of copper (I) oxide are classified as reducing sugars.
  • 40. Results of Benedict's Test with Glucose, Sucrose, and Fructose
  • 41. Glucose • Less than 0.1% of glucose normally filtered by the glomerulus appears in urine (< 130 mg/24 hr). • Glycosuria (excess sugar in urine) diabetes generally means mellitus.
  • 42. • Exton’s Test • There are several acids Protein which can be used to precipitate proteins - sulfosalicylic, trichloroacetic, nitric, and acetic acids. • Sulfosalicylic acid (SSA) is the most frequently used acid test because it does not require the use of heat. • Exton’s reagent is 5% sulfosalicylic acid in a
  • 43. To perform the SSA procedure mix equal parts of patient urine and the reagent. Rate the amount of turbidity according to the following scale:
  • 44. Interpretation • In rough terms, trace positive results (which represent a slightly hazy appearance in urine) are equivalent to 10 mg/100 ml or about 150 mg/24 hours (the upper limit of normal). • 1+ corresponds to about 200-500 mg/24 hours • a 2+ to 0.5-1.5 gm/24 hours • a 3+ to 2-5 gm/24 hours • a 4+ represents 7 gm/24 hours or greater.
  • 45. • Normally, only small plasma Interpretation proteins filtered at the glomerulus are reabsorbed by the renal tubule. • Normal total protein excretion does not usually exceed 150 mg/24 hours or 10 mg/100 ml in any single specimen. • More than 150 mg/day is defined as proteinuria. • Proteinuria > 3.5 gm/24 hours is severe and known as nephrotic syndrome.
  • 46. Apply Your Knowledge A urine analysis has detected that a patient has protein in his urine. Why is this important? 46
  • 47. - Apply Your Knowledge Answer A urine analysis has detected that a patient has protein in his urine. Why is this important? Protein in the urine usually indicates renal disease 47
  • 48. Ketones Gunning’s Test • Ketones (acetone, aceotacetic acid, beta- hydroxybutyric acid) resulting from either diabetic ketosis or some other form of calorie deprivation (starvation) • Shows positive test for: acetaldehyde and methyl ketones
  • 49. Ketones Gunning’s Test • Detection of COOH group is possible through its transformation to an ester in reaction with alcohol. • The esters have characteristic smell of flowers or fruits • It should appear the yellow precipitate of iodoform with characteristic smell • Ketones Normal:There are no ketones in your blood or urine. • Abnormal:Ketones are present in your blood or urine.
  • 50. • . Indications of a positive test: The formation of a yellow precipitate or suspension of iodoform is a positive test a negative test (left) and a positive test (right)
  • 51. Smith’s Test (Bile pigments)
  • 52.
  • 53. emerald green at the point of contact between urine sample and alcoholic iodine mixture Bilirubin – green Bilicyanine – blue Choletelin - yellow
  • 54. • Smith’s Test should be conducted to determine whether bilirubin is present in the urine (i.e., bilirubinuria). • Urobilinogen is formed by intestinal bacteria from the breakdown of conjugated bilirubin, and it is usually excreted in feces. • However a small amount may be reabsorbed and excreted in urine.
  • 55. • A positive test response indicates normal enterohepatic circulation of biliary pigments. High concentrations of biliary pigments may occur in hemolytic crisis, or cases of hepatic or intestinal dysfunction.
  • 56. 56
  • 58. FIRST PHASE: • Collect urine sample from a donor from each group SECOND PHASE: • Unknown sample will be tested
  • 59. Record all results and analysis in your lab work sheet. • Post lab group discussion upon submission of all reports will be conducted.