Complete Text (Part 5)

When ready to use, add 1 cc. of the bromine to 40 cc. of the XaOH solution. Fill the ureometer tube and add by means of a curved pipet 1 cc. of urine to the upright part of the tube. Read grams off on the scale. Method unsatisfactory. 59 PoUn metJiod. The principle of this method is that crystalized magnesium chloride boils in its own water of crjs- talization at 160 degrees centigrade. If urea he present it is decomposed into ammonia and carbon dioxide. If acid be present the ammonia formed will combine with the acid and can be titrated. Urease method (Marshall). The principle of this method is this : The soy bean extract contains an enzyme capable of splitting urea into ammonium carbonate. Characteristics of the enzyme : 1. It is soluble in water. 2. It reacts quantitatively with urea. It will split off a certain amount in a given time and no more. 3. It does not form NHg from any other source than urea, and will form NH3 in the pres- ence of any other substance, except. (5j. 4. Its optimum activity is 55 deg. C. 5. Its activity is destroyed by acids, heavy metals and alcohol above 20%. 6. It has a self-retarding effect. After a cer- tain amount of (^2:4)2003 is formed its action becomes inhibited. Direct method of performing test: Test tube A. 10 cc. of urine. Test tube B. 10 cc. of urine. 1 — 2 urease tablets dissolved in water. Allow these to stand: 1. At room temperature over night, or 2. 45 to 50 deg. C for 1 hour, or 3. 37 deg. C. for 3 hours. Titrate both tubes for alkalinity, using methyl orange as an indicator. 60 Subtract the no. of cc. used in titrating A from the no. used in titrating B. 1 cc. of N/10 HCl equals .003 grams of urea or .0014 grams X. The disadvantage of the test is that the whole solution becomes clouded and the end reaction is an (obscure one. Indirect method. Test tube A. 5 cc. of urine. Aqueous solution of 1 or 2 urease tablets. A layer of kerosene to prevent foaming. Stopper and allow to stand as before. Test tube B. 25 cc. of X/10 HCl. 2 drops of 1% sol. alizarin, A layer of kerosene. After tube A has stood its proper length of time, connect it up to a suction apparatus in such a way that the indrawn air first passes through a solution of HgSOi. Allow air to be drawn through for about a minute in order to remove any free ammonia that may be present. Now, connect tubes A and B in such a way that the air current passes from A into P. When this has been done, add a teaspoonful of dry potassium carbonate to tube A, quickh^ cork, shake and start suction apparatus. Allow air cur- rent to pass from A into B for about 30 minutes. The ammonia in A is freed and caught in the N/10 HCl in B. Titrate the excess of HCl in B with X/10 XaOH. Subtract the number of cc' s necessary to bring about the end reaction from 25 cc, the amount of X/10 HCl originally added. Repeat this process, using 5 cc of urine to which no urease solution is added. This gives the preformed ammonia. This value subtracted from the former gives the amount of XHo formed from urea. Make the calculation of the amount of urea on 6.1 the basis of: 1 cc. of X/10 HCl equals .003 grams of urea or .0014 grams X. ZJiic acid. Amount : .2 to 2.0 grams per day. Mixed diet .37 gm. or .8% total X. N-free diet .09 gm. or 2.5% total X. Origin. Uric acid is not a product of protein decomposi- tion. Xucleo-protein plus pepsin, trypsin, etc., gives nucleic acid and protein. Xucleic acid plus tetranuclease gives purin and ' pyrimidin dinucleotide. Purin nucleotide plus pliosplio-nuclease and purin nuclease give: the former, adenosin and guanosin ; the latter, adenin and guanin. Adenin plus adenase gives hypoxanthin. Guanine plus guanase gives xanthin. Adenosin plus adenosin deaminase and guanosin plus guanosin deaminase give xanthin and inosin, which when the c-h radical is split off give xanthin and hypoxanthin. Xanthin and hypoxanthin plus xanthin oxi- dase (liver) gives uric acid. Sources of uric acid : 1. Exogenous. The greater portion of nucleic acid comes from the diet : sweet bread, liver, caiTein and theobromin. 2. Endogenous. a. Breaking down of tissue nuclei. b. Purin bases free in the body. 3. Synthesis. There is no evidence that this occurs in man, but it does occur in birds. Uric acid destruction in the body. In birds it is .destroyed by ferments and allan- toin is the excretory product. 62 The following facts are known : 1. Uric acid fed can be recovered quantitatively in the urine. 2. Endogenous purine metabolism in persons on an N-free diet is constant. 3. Uric acid can be broken down by other routes than allantoin. a. Glycocoll. b. Oxalic acid-urea. c. Persons fed on purin bases excrete half in the form of urea. Purin bodies can be excreted in forms not going through the uric acid s'tage. 4. When uric acid reaches the system it is ex- creted as such. C/haracteristic properties of uric acid. In the cold it is sparingly soluble in H2O (1 — 40,000). It is fairly soluble in blood serum (1 — 1,000). Its best solvent is a solution of urea. It is insoluble in alcohol, ether, chloroform and acetic acid. It is somewhat soluble in HCl, II2SO4 and weak alkalines. Solutions of uric acid in water are neutral to litmus. Uric acid in sufficient con- centration reduces Fehling's solution, but not Nylander's. Uric acid is kept in solution under the following conditions : 1. With neutral phosphates. 2. When pigments are present. 3. As neutral urates. 4. As monosodium or biurate — Ammonium biurate (urinary type of sedi- ment). Sodium biurate (gout). Excretion of uric acid. 63 Increased : 1. Purin or meat rich diets. 2. Increased protein decomposition. 3. When lencocvtosis is predominant feature in leucaemia, especially when treated with X-ray. From 12 to 15 gms. per day is sometimes excreted. 4. Pernicious anaemia. (In secondary anaemia less than normal.) 5. Articular rheumatism and after medication with salicylates, atophan, colchicum and urotropin. Decreased : 1. Vegetable diet. 2. Certain types of chronic nephritis. 3. Chronic lead poisoning (diagnostic). 4. Quinine and opium. Uric acid in relation to gout : All theories have fared badly. 1. In cases of chronic gout the excretion of uric acid upon a given diet will be the same as in normal individtials on the same diet in most cases. 2. The excretion of uric acid in chronic gouty in- dividuals persists in being lower than normal. 3. Gout}' patients fed with purin bases will show a lower curve of uric acid excretion than normal. 4. On a purin free diet gotity patients will show a lower endogenous excretion than normal. 5. The blood of gouty patients contains more uric acid than normal, 4 to 5 mg. instead of 1 mg. 6. The elimination of uric acid is less than normal from 1 to 3 days preceding an acute gouty attack. 7. With the start of the acute symptoms there is a rise of the excretion above normal which 64 lasts for three or four days, and then is fol- lowed by a much lower output than normal. 8. The relation between an excess of uric acid in the blood and in the urine in acute gouty manifestations is not known. 0. Gouty individuals show utter abnormal condi- tions of metabolism. 10. Probable explanation of gout is that gouty individuals do not possess necessary uric acid elimination and certain tissues seem to pos- sess an aflfinity for uric acid. Factors modifying uric acid excretion. 1. Diet. 2. Tissue metabolism. 3. Indeterminate amount of free purin bases. 4. Amount of conversion of uric acid into urea. 5. Amount of uric acid destruction in the body. 6. How much synthesis occurs. 7. Varyino: degree of blood retention. 8. Kenal capacity of eliminating uric acid. 9. Amount of urinary phosphates. 10. Degree of urinary concentration. 11. Akalinity of urine. Quantitative uric acid determination. Folin method : Urine 300 cc. Folin's reagent To* cc. (gives precititate of phosphates) Folin's reagent : Am. Sulph. 500 gm. Uranium acetate 5 gm. 10% acetic 60 cc. H2O q. s. ad. 1,000 cc. Wait 15 minutes and filter. Place 125 cc. of the filtrate (which equals 100 cc. of urine) in an Ehrle- meyer flask. Add 5 cc. of concentrated XH^OHj let 65 stand 24 hours and filter. Collect the precipitate of ainmoniiTiTi urate, wash it with ammonium sulphate until the filtrate no longer shows a reaction with AgXOo, pierce the filter paper and wash the precipi- tate into a breaker Avitli 100 cc. of water. Add 15 cc. of con. HoSOj, lieat over a fiame, and while still hot titrate witli N/20 potassium xjermanganate (1.5(>7 gn?. in 100 cc. ) to end point, Avhich is a red- dish blush for from 15 to 30 seconds. Calculation : 1 cc. of X/20 potassium permanga- nate equals 3.75 mg. of uric acid. (Uric acid has the proi)erty of reducing i^otassium permanganate.) Correction: ]"or each 100 cc. of urine add 3mg. of uric acid, for a certain amount of urate is soluble and does not change into uric acid. Aiiiinoiiia : De])en<l> upon protein, intake and runs parallel with X excretion. On a mixed diet about .7 gms. are excreted in 24 hours or 4.37% of the total N, and upon an N-free diet about .42 gm. or 11.3% of the total N is excreted. Increased : 1. Decreased oxidation. 2. Acute febrile conditions. 3. Liver diseases. 4. Uraemia. 5. Toxic vomiting of pregnancy. 6. Diabetes, characteristicalh' abnormal. 7. Starvation. 8. Chloroform poisoning. Decreased : 1. Xephritis. 2. Most akaline therapy. 3. Conditions with Ioav HCl. XHo elimination is associated with acidosis. In this condition one finds : 1. Increase of acids with formation of neutral salts. 2. Lowered blood and 66 tissue alkalinity. 3. Carrying capacity of the blood for CO, lowered and CO2 accumulates in the tissues (tissue asphyxia). 4. Increase in the H concentra- tion of the blood stimulates the respiratory center and air-hunger results. Occurrence of acidosis : 1. Diseases with clinical symptonis. Diabetic acidosis. Acute nephritis and acute diseases associated A\ith nephritis. In late stages of primary and secondary con- tracted kindey. Food intoxication of children. Atrophic cirrhosis. 2. Diseases without clinical symptoms. Acute febrile conditions. Advanced cachexia. Severe anaemia. After general anaesthesia. Cardiac conditions. Quantitative ammonia determination. Schlosing method : The objection to this method is that the results are too high. Folin's method : 10 to 20 cc. of urine are introduced into an aerat- ing cylinder and a layer of kerosene added to reduce tlie tendency to foam. The apparatus is connected with a flask containing a knoAvn quantity of X/10 HoSO^ (20 cc.) and sufficient water to well cover tlie absorption tube. About one gram of sodium carbonate is added to the urine and the ammonia liberated is aspirated by means of a suction pump into the known acid solution. The current of air should first pass through a dilute £[280^ solution in order to absorb any ammonia that may be x)resent 67 in the indrawn air. Aerate for one hour and titrate the excess of acid Avith X/10 NaOH, using methyl orange or alizarin as indicator.. 1 cc. of N/10 H.SO, equals .0017 grams of XH.. or .0014 grams of X. Ci'catiu'ui : About .50 gm., or 3.0% of the total X, is excreted upon a mixed diet, and about .0 gm., or 17.2% of the total X, is excreted on an X-free diet. It is an anhydride of creatin, but the relationship between tiie two is not clear. Creatin probably comes from arginin, and creatinin is formed by the action of anliATlrating ferments. There is a remark- ably constant excretion of creatin irrespective of the X intake. Folin considers creatinin a valuable index of protein metabolism, while Shaffer thinks it an index of some special i)rocess of normal meta- bolism occurring mostly in muscles. Exogenous elimination depends upon the ingestion of meats. Endogenous elimination has the following char- acteristics : 1. Constant on X-free diet. 2. Varies with different individuals. o. Apparently independent of total X metabolism - J:. Depends upon weight of individual. A normal individual excretes from 7 to 12 mg. per kilo in 21: hours. Source: Largely due to muscle metabolism of the body. It is, however, formed in other than muscle tissue. There is a diminished excretion in anaemia, marasmus, myositis ossificans, chlorosis, phthisis, chr. diffuse nephritis, progressive muscular atrophy, and pseudohypertrophic paralysis. It is said to be increased in the acute stages of some fevers and exophthalmic goiter. The X excretion runs more or less parallel with protein decomposition, but creati- 6S nin does not. Tlie liver is one main source of forma- tion. Qualitative tests for creatinin. JaftVs test: To urine add picric acid and dilute NaOH (10%), which turns a brilliant sliade of red and which be- comes darker on standing. Upon the addition of acetic acid it changes to an ochre color. Weyl's test : To urine add nitroprussid of soda and a few drops of 10% XaOH. A deep red orange or ruby color appears, which, upon the addition of a little glacial acetic acid, turns to green and upon heating to Ber- lin blue. Quantitative determination. Folin method (colorimetric) : A standard solution is made up by the following method : One gram of pure creatinin is dissolved in 1000 cc. of water. 1 cc. of this plus 20 cc. picric add plus 1.5 cc. of 10% NaOH plus enough- water" to make 100 cc. gives the standard color. One cubic centimeter is treated in the same man- ner, and the resulting color is compared with the standard solution by means of a colorimeter. Undetermined Nitrogen. Mixed diet .7 grams or 4:.31% X-free diet .29 grams or 7.3 % This is determined by subtracting the sum of the other forms of N from the total N. Source : 1. Amino acids which are known to be excreted. Tyrosin, leucin, giycocoll (1^-2 grams per day). In cystinuria there is a perverted metabolism to amino acid; 181 such cases have been reported. These patients do not have the power to reduce the amino acids 69 further than cvstiii. Gystin calculi fre- quently form. A marked hereditary ten- dency prevails in this disease in which males are more frequently effected than females. In diaminouria putrescin and cadaverine are excreted. 2. Hippuric acid. 0.1 to 1 gram per day. Ben- zoic acid foods, such as fruit, berries, prunes, etc., combine with giycocoll. 3. Oxyproteic acid 1 These are responsible for Alloxyproteic acid J Ehrlich's diazo reaction. Their excretion is probably increased in cancer. They contain an S mollecule. 4. Allantoin. This is normally present only in traces. It exists normally and abnormally in conditions little understood. REDUCING BODIES OF THE URINE. Subjects to be considered : 1. General review of carbohydrates. 2. Qualitative determination of glucose. 3. Sugars and reducing bodies other than glucose. 4. Quantitative determination of sugar. 5. Scheme for running down reducing sub- stances in urine. 6. Acid bodies. 7. Diabetes. Carbohydrates. The available carbohydrates of the body exist as glycogen, which is stored in the liver and muscle tissues until called upon b}^ the tissues for energy, heat and tissue formation. Sugar is constantly being converted by the liver into glycogen and back again into sugar in accordance to the body needs. The balance between these two in health is such that the percentage in the blood is from .1 to .15%. 70 Origin of carbohydrates : 1. Sugars and starches. 2. Proteins and amine acid bodies. ?>. Fats. Function of carbohydrates : 1. Energy. 2. Heat. "^ B. Protecting agency for protein. 4. When in excess forms fats. 5. Actual synthesis of tissues (nucleic acid). Kegulation : 1. Supply (glycogenetic function). 2. Storage ( glycogenesis ) . 3. Tissue consumption (glycolysis). When any one of these breaks down glycosuria may result, also in cases of renal permeability for carbohydrates (renal diabetes and phloridzin dia- betes). Tlie adrenal and pancreas are antagonistic in func- tion. In health the two are in perfect equilibrium, but in diabetes the restraint is removed from the liver. The i)ancreas also seems to give off some- thing to the tissues T\iiich enables them to burn carboliydrates to COg and HgO. In diabetes this falls short and sugar is eliminated through the kidneys, due to its accumulation in the blood. A'^ormal urine contains a small amount of glucose (animal gum or isomaltose). It amounts to about .3 to .G grams in 24 hours. Glycuronic acid occurs in amounts of about .004 grams in 100 cc. Traces of pentose occur occasionally after the ingestion of fruits. Lactose and galactose occur frequently in sucklings. 71 Assimulation limit for carbohyrates (two kinds) : .1. Individuals who show sugar after the inges- tion of large amounts of starch (glyco- suria ex amylo). 2. Individuals excreting sugar following sugar intake (glycosuria e saccharo). A normal individual can take as much as 150 to 200 grams of glucose on an empty stomach without showing glucose in the urine. Per- sons with nephritis have a lower sugar threshold. The liver has the greatest toler- ance for glucose and least tolerance for lactose and galactose. Maltose, levulose and saccharose come in the intermediate group. Hamman's method of making threshold test : From 150 to 200 grams of dextrose are dissolved in 150 cc. of watei^ and flavored with lemon or orange juice. The mixture is made ice-cold with cracked ice and the volume made up to 300 cc. This is taken slowly on