Chemistry -Nonprotein Nitrogenous Compounds

What endogenous substance may cause a positive interference in the urease/glutamate dehydrogenase assay? A. Ammonia B. Creatinine C. Glucose D. Cholesterol

A. Adequate specificity is generally obtained when using the urease/glutamate dehydrogenase method. Because urease hydrolyzes urea to ammonia and water, a positive interference form endogenous ammonia will occur with elevated blood levels of ammonia. Such interference may occur from use of aged blood specimens and in certain metabolic diseases

During chemotherapy for leukemia, which of the following analytes would most likely be elevated in the blood? A. Uric acid B. Urea C. Creatinine D. Ammonia

A. An increase in serum uric acid levels may be seen during chemotherapy for leukemia. The cause of this is the accelerated breakdown of cell nuclei in response to the chemotherapy. Other proliferative disorders that may respond similarly are lymphoma, multiple myeloma and polycythemia. It is important that serum uric acid be monitored during chemotherapy to avoid nephrotoxicity.

Which of the following disorders is best characterized by laboratory findings that include increased serum levels of inorganic phosphorus, magnesium, potassium, uric acid, urea and creatinine and decreased serum calcium and eyrthropoietin levels? A. Chronic renal failure B. Renal tubular disease C. Nephrotic syndrome D. Acute glomerulonephritis

A. As renal function continues to be lost over time, chronic renal failure develops. Chronic renal failure is manifested by loss of excretory function, inability to regulate water and electrolyte balance, and increased production of parathyroid hormone, all of which contribute to the abnormal laboratory findings. The decreased production of erythropoietin causes anemia to develop.

When the Jaffe reaction is employed as a kinetic assay to quantify serum creatinine which of the following is used in the analysis? A. Serum sample used directly B. Folin-Wu filtrate C. Somogyi-Nelson filtrate D. Trichloroacetic acid filtrate

A. Because protein will interfere with the Jaffe reaction, serum for a manual creatinine analysis is treated with sodium tungstate and sulfuric acid to precipitate the proteins. The use of tungstic acid to make a protein-free filtrate is known as the Folin-Wu method. The protein-free filtrate, which still contains creatinine and other reducing substances, is then mixed with alkaline picrate reagent to yield the characteristic Jaffe reaction. Automated methods have replaced manual methods. These kinetic methods using the alkaline picrate reagent system have been adapted to use small volumes of serum and have readings taken within a short interval of 25-60 sec following initiation of the reaction. Because of the speed at which the analysis is performed and the small serum sample requirement, serum may be used directly, alleviating the need for a protein-free filtrate.

An increased serum level of which of the following analytes is most commonly associated with decreased glomerular filtration? A. Creatinine B. Uric acid C. Urea D. Ammonia

A. Creatinine is a waste product of muscle metabolism and as such its production is rather constant on a daily basis. Creatinine is freely filtered by the glomerulus, with only a very small amount secreted by the proximal tubule. Thus, measurement of creatinine is a reflection of glomerular filtration. An increase in the serum creatinine level would be indicative of decreased glomerular filtration. Although uricc acid, urea and ammonia levels may be increased with decreased glomerular filtration, increased levels of these analytes are associated with a number of specific metabolic diseases and, therefore, they are not used as indicators of the glomerular filtration rate.

Which of the following reagents is not utilized in a coupled enzymatic reaction method to quantify serum creatinine? A. Picric acid B. Chromogenic dye C. Creatinine amidohydrolase D. Sarcosine oxidase

A. In addition to the endpoint and kinetic methods, which use the Jaffe reaction (picric acid), several methods have been developed that use coupled enzymatic reactions fot the quantification of creatinine. In one such method, creatinine amidohydrolase (creatininase) catalyzes the conversion of creatinine to creatine and subsequently to sarcosine and urea. Sarcosine oxidase catalyzes the oxidation of sarcosine to glycine, formaldehyde and hydrogen peroxide. The hydrogen peroxide reacts with the reduced form of a chromogenic dye in the presence of peroxidase to form an oxidized colored dye product that is read sectrophotometrically.

To maintain acid-base balance, it is necessary that the blood ammonia level be kept within narrow limits. This is accomplished primarily by which of the following? A. Synthesis of urea from ammonia B. Synthesis of glutamine from ammonia C. Excretion of ammonia in the bile D. Excretion of ammonia in the stools

A. The catabolism of some amino acids involves a transamination reaction in which the alpha-amino group of the amino acid is enzymatically removed. After its removal, the alph-amino group is transferred to an alpha-keto acid (alpha-ketoglutarate) with the formation of L-glutamate. Glutamate, which is the common product formed by most transaminase ractions, then may undergo oxidative deamination in the liver mitochondria with the formation of ammonia. The ammonia thus formed leaves the mitochondria as the amino group of citrlline. Citrulline, in turn, condenses with aspartate, which contains the second amino group needed for urea synthesis, forming argininosuccinate, which ultimately leads to the formation of urea. Therefore, the formation of urea and its excretion in the urineprovide the principal means by which the body is able to free itself of excess ammonia.

Which of the following methods utilizes urease and glutamate dehydrogenase for the quantification of serum urea? A. Berthelot B. Coupled enzymatic C. Conductimetric D. Indicator dye

B. An enzymatic method for quantifying urea employs urease and glutamate dehydrogenase (GLDH) in a coupled enzymatic reaction. Urease catalyzes the production of ammonium carbonate from urea. The ammonium ion produced reacts with 2-oxoglutarate and NADH in the presence of GLDH with the formation of NAD+ and glutamate. The decrease in absorbance, as NADH is oxidized to NAD+ is followed kinetically at 340 nm using a spectrophotometer. In the conductimetric method, the formation of ammonium ions and carbonate ions, from the ammonium carbonate, causes a change in conductivity that is related to the amount of urea present in the sample.

In the urea method, the enzymatic action of urease is inhibited when blood for anaylsis is drawn in a tube containing what anticoagulant? A. Sodium heparin B. Sodium fluoride C. Sodium oxalate D. Ethylenediaminetetra-acetic acid

B. In addition to the fact that sodium fluoride is a weak anticoagulant, it also functions as an antiglycolytic agent and is used as a preservative for glucose in blood specimens. With the urease reagent systems for the quantification of urea, the use of sodium fluoride must be avoided because of its inhibitory effect on this system. Additionally, contamination from the use of ammonium oxalate and ammonium heparin must be avoided, because urease catalyzes the production of ammoniumcarbonate from urea. In several methods, the ammonium ion formed reacts proportionally to the amount of urea originally present in the sample. Anticoagulants containing ammonium would contribute falsely to the urea result.

In the diacetyl method, what does diacetyl react with to form a yellow product? A. Ammonium B. Urea C. Uric acid D. Nitrogen

B. In the diacetyl method, acidic diacetyl reacts directly with urea to form a yellow-diazine derivative. Thiosemicarbazide and ferric ions are reagents used to intensify the color of the reaction. Because urea is quantified directly, the method does not suffer from interferences from ammonia contamination, as do some of the urea methods

What is the end product of purine catabolism in humans? A. Urea B. Uric acid C. Allantoin D. Ammonia

B. Through a sequence of enzymatic reactions, the purine nucleosides, adenosine and guanosine, are catabolized to the waste product uric acid. The catabolism of purines occurs primarily in the liver, with the majority of uric acid being excreted as a urinary waste product. The remaining amount of uric acid is excreted in the biliary, pancreatic and gastrointestinal secretions through the gastrointestinal tract. In the large intestine, uric acid is further degraded by bacteria and excreted in the stool

In the ultraviolet procedure for quantifying uric acid, what does the reaction between uric acid and uricase cause? A. Production of reduced nicotinamide-adenine dinucleotide (NADH) B. The formation of allantoin C. An increase in absorbance D. A reduction of phosphotungstic acid

B. Uric acid absorbs light in the ultraviolet region of 290-293 nm. When uricase is added to a uric acid mixture, uricase destroys uric acid by catalyzing its degradation to allantoin and carbon dioxide. On the basis of these two characteristics, differential spectrophotometry has been applied to the quantification of uric acid. This type of method is used on analyzers that are capable of monitoring the decrease in absorbance as uric acid is destroyed by uricase. The decrease in absorbance is proportional to the concentration of uric acid in the specimen.

From what precursor is creatinine formed? A. Urea B. Glucose C. Creatine D. Uric acid

C. Creatine is synthesized from the amino acids arginine, glycine and methionine. In tissues that include the kidneys, small intestinal mucosa, pancreas and liver, arginine and glycine form guanidoacetate through a transmtheylase reaction to form creatine. Creatine is transported in the blood to muscle tissue. Creatine in the form of phosphocreatine is a high-energy storage compound that provides the phosphate needed to produce adenosine triphosphate (ATP) for muscle metabolism. When ATP is formed from phosphocreatine, free creartine is also released. Creatine, through a spontaneous and irreversible reaction, forms creatinine. Creatinine serves no functional metabolic role. It is excreted in the urine as a waste product of creatine.

When it is not possible to perform a creatinine assay on a fresh urine specimen, to what pH level should the urine be adjusted? A. 3.0 B. 5.0 C. 7.0 D. 9.0

C. Creatinine assays are preferably performed on fresh urine specimens. If an acid urine specimen is kept for a time, any creatine in the urine will be converted to creatinine. In alkaline urine, an equilibrium situation will occur between the creatine and creatinine present in the specimen. To avoid either of these situations, it is recommended that the urine be adjusted to pH 7.0 and that the specimen be frozen. It is thought that at a neutral pH, the integrity of the urine specimen will be maintained because it will require days or even weeks for equilibrium to occur between the two compounds.

An endogenous substance assayed to assess the glomerular filtration rate may be described as being filtered by the glomeruli, not reabsorbed by the tubules, and only secreted by the tubules when plasma levels become elevated. What is this frequently assayed substance? A. Inulin B. Uric acid C. Creatinine D. Urea

C. Creatinine is an endogenous substance that is filtered by the glomeruli and normally is neither reabsorbed nor secreted by the tubules. When plasma levels of creatinine rise, some secretion of creatinine by the tubules will occur. The filtration properties of creatinine and the fact that it is a substance normally present in blood make the creatinine clearance test the method of choice for assessing the glomerular filtration rate.

Which of the following does not need to be done when collecting, handling and using a specimen for ammonia analysis? A. Avoid using a hemolyzed specimen B. Collect blood in EDTA or heparin evacuated tubes C. Place specimen in a 37 degree Celsius water bath immediatley D. Advise patient not to smoke for 8 hours before blood collection

C. Plasma is the specimen of choice for ammonia analysis. EDTA and heparin (not the ammonium salt) are acceptable anticoagulants. Because exposure of blood to air is contrindicated, the evacuated blood collection tube should be filled completely. The blood specimen should be placed on ice immediately and centrifuged as soon as possible to inhibit deamination of amino acids. Because the concentration of ammonia in red blood cells is approximately three times greater than in plasma, the analysis should be performed on a nonhemolyzed specimen. Because of the false increase in ammonia levels caused by smoking, patients should be instructed to refrain from smoking for 8 hours before blood collection.

In the Berthelot reaction, what contaminant will cause the urea level to be falsely elevated? A. Sodium fluoride B. Protein C. Ammonia D. Bacteria

C. The Berthelot reaction is based on the production of a blue-indophenol compound when ammonia reacts in an alkaline medium with phenol and sodium hypochlorite. This basic colorimetric reaction can be used to quantify both urea and blood ammonia levels. Therefore, any ammonia contamination (i.e in the distilled water used to make reagents for the urea procedure and on glassware) must be avoided so that falsely elevated urea values will not be obtained.

Exxpress 30 mg/dL of urea nitrogen as urea. A. 14 mg/dL B. 20 mg/dL C. 50 mg/dL D. 64 mg/dL

D. Because the substances classified as nonprotein-nitrogen (NPN) compounds were quantified by assaying for their nitrogen content, it became customary to express urea as urea nitrogen. When urea was expressed as urea nitrogen, a comparison could be made between the concentration of urea and the concentration of other NPN compounds. When it is necessary to convert urea nitrogen values to urea, the concentration may be calculated easily by multiplying the urea nitrogen value by 2.14. This factor is derived from the molecular mass of urea (60 daltons) and the molecular weight of its two nitrogen atoms (28): 60/28=2.14

What is the compound that comprises the majority of the nonprotein-nitrogen fractions in serum? A. Uric acid B. Creatinine C. Ammonia D. Urea

D. Constituents in the plasma that contain the element nitrogen are characterized as being protein- or nonprotein-nitrogen compounds. The principal substances included among the nonprotein-nitrogen compounds are urea, amino acids, uric acid, creatinine, creatine and ammonia. Of these compounds, urea is present in the plasma in the greatest concentration, comprising approximately 45% of the nonprotein-nitrogen fraction.

What compound normally found in urine may be used to assess the completeness of a 24-hour urine collection? A. Urea B. Uric acid C. Creatine D. Creatinine

D. Creatine is predominantly found in muscle cells, where the quantity of creatine is proportional to muscle mass. As muscle metabolism proceeds, creatine is freed from its high-energy phosphate form, and the creatine, thus liberated, forms the anhydride creatinine. The quantity of creatinine formed daily is a relatively constant amount because it is related to muscle mass. Therefore, it has been customary to quantify the creatinine present in a 24-hour urine specimen as an index of the completeness of the collection.

In gout, what analyte deposits in joints and other body tissues? A. Calcium B. Creatinine C. Urea D. Uric acid

D. Gout is a pathological condition that may be caused by a malfunction in the renal excretion of uric acid. Two of the major characteristics of gout are hyperuricemia and a deposition of uric acid as monosodium urate crystals in joints, periarticular cartilage, bone, bursae and subcutaneous tissue. Such a deposition of urate crystals causes inflammation of the affected area and precipitates an arthritic attack.

Which of the following statements can be associated with the enzymatic assay of ammonia? A. Increase in absorbance monitored at 340 nm B. Nicotinamide-adenine dinucleotide (NAD+) required as a cofactor C. Ammonium ion isolated from specimen before the enzymatic step D. Reaction catalyzed by glutamate dehydrogenase

D. Ion-exchange, ion-selective electrode and enzymatic methods have been employed for the analysis of ammonia in plasma specimens. Because the enzymatic method is a direct assay, prior separation of ammonium ions is not required. The enzymatic reaction catalyzed by glutamate dehydrogenase follows: 2-Oxoglutarate + NH4+ + NADPH <--> Glutamate + NADP+ + H2O The rate of oxidation of NADPH to NADP+ is followed as a decreasing change in absorbance at 340 nm.

When a blood ammonia determination is performed, the blood specimen must be treated in a manner that will ensure that A. The deamination process continues in vitro B. Glutamine formation in vitro is avoided C. The transamination process continues in vitro D. Ammonia formation in vitro is avoided

D. It is necessary that certain precautions in specimen handling be exercised because the enzymatic process of deamination of amides continues at room temperature after a blood sample is drawn. When blood is drawn for ammonia analysis, it is critical that any in vitro ammonia formation be prevented. It is recommended that the tube containing the blood specimen be placed in an ice bath immediately after the blood is drawn, because the cold environment will help retard metabolic processes. It is also important that the chemical analysis of the specimen be started within 20 minutes of drawing the specimen.

A serum creatinine was found to be 6.0 mg/dL. Which of the following urea nitrogen serum results would support the same pathological condition? A. 6 mg/dL B. 20 mg/dL C. 35 mg/dL D. 70 mg/dL

D. Serum urea nitrogen and creatinine levels are frequently reuested together so that their ratio can be evaulated. The normal ratio of serum urea nitrogen to creatinine ranges between 10:1 and 20:1. Abnormal values obtained when kidney function tests are performed may be the result of a prerenal, renal or postrenal malfunction. The ratio of urea nitrogen to creatinine is sometimes used as an index in the assessment of kidney function and as a means of differentiating the source of the malfunction.

What analyte is measured using the Jaffe reaction? A. Urea B. Uric acid C. Ammonia D. Creatinine

D. The Jaffe reaction, which was described in 1886, is still used for creatinine analysis. The Jaffe reaction employs the use of an alkaline picrate solution that reacts with creatinine to form a bright orange-red complex. A drawback to this procedure is its lack of specificity for creatinine, because noncreatinine chromogens, glucose and proteins are also able to react with alkaline picrate.

The creatinine clearance tests is routinely used to assess the glomerular filtration rate. Given the following information for an average-size adult, calculate a creatinine clearance. Urine creatinine-120 mg/dL Plasma creatinine-1.2 mg/dL Urine volume for 24 hours-1520 mL A. 11 mL/min B. 63 mL/min C. 95 mL/min D. 106 mL/min

D. The creatinine clearance test is used to assess the glomerular filtration rate. An accurately timed 24 hour urine speecimen and a blood sample, drawn in the middle of the 24-hour urine collection, are required. The creatinine concentrations of the urine specimen and the plasma are determined and these values, along with the urine volume, are used to determine the creatinine clearance. The body surface area will not be used in the calculation because the clearance is being done on an average size adult. The following general mathematical formula is used to calculate creatinine clearance: (U/P) X V= Creatinine clearance (mL/min) where U= urine creatinine concentration in milligrams per deciliter, P=plasma creatinine concentration in milligrams per deciliter and V= volume of urine per minute, with volume expressed in milliliters and 24 hours expressed as 1440 minutes. Applying this formula to the problem presented in the question: (120mg/dL / 1.2 mg/dL) X (1520 mL/24 hr /1440 min/24 hr)= 106 mL/min It should be noted that both the size of the kidney and the body surface area of an individual influence the creatinine clearance rate. Because normal values for creatinine clearance are based on the average adult body surface area, it is necessary that the clearance rate be adjusted when the body surface area of the individual being tested differs significantly from the average adult area. This type of adjustment is especially critical if the individual is an infant, a young child, or an adolescent. The body surface area may be calculated from an individual's height and weight, or it may be determined from a nomogram. The average body surface area is accepted as being 1.73 m^2. The mathematical formula used to calculate a creatinine clearance when the body surface area of the individual is required follows: (U/P) X V X (1.73/A) = Creatinine clearance (mL/min/standard surface area) where 1.73= standard adult surface area in square meters and A=body surface area of the individual in square meters.

Which of the following disorders is not associated with an elevated blood ammonia level? A. Reye syndrome B. Renal failure C. Chronic liver failure D. Diabetes mellitus

D. The gastrointestinal tract is the primary source of blood ammonia. With normal liver function, ammonia is metabolized to urea for urinary excretion. When blood ammonia levels become elevated, toxicity of the central nervous system occurs. Diseases associated with elevated blood ammonia levels include Reye syndrome, renal failure, chronic liver failure, cirrhosis and hepatic encephalopathy.

When mixed with phosphotungstic acid, what compound causes the reduction of the former to a tungsten blue complex? A. Urea B. Ammonia C. Creatinine D. Uric acid

D. Uric acid may be quantified by reacting it with phosphotungstic acid reagent in alkaline solution. In this reaction, uric acid is oxidized to allantoin and the phosphotungstic acid is reduced, forming a tungsten blue complex. The intensity of the tungsten blue complex is proportional to the concentration of uric acid in the specimen.