Manual 
 
 inical 
 
 ; col °gy 
 

 CORNELL UNIVERSITY. 
 
 THE 
 
 ilosiuell p. IfTotucr Elbrarg 
 
 THE GIFT OF 
 
 ROSWELL P. FLOWER 
 
 FOR THE USE OF 
 
 THE N. Y. STATE VETERINARY COLLEGE. 
 
 1897 
 
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 3- 
 
 QP 5,9.^™" ""'"""V Library 
 A, 3bora<ory manual of physiological and 
 
 3 1924 000 913 867 
 
A LABORATORY MANUAL 
 
A LABORATORY MANUAL 
 OF PHYSIOLOGICAL AND 
 C 
 
 Al 
 
 Cornell University 
 Library 
 
 By ARTHUR E. AUSTIN, A.B., M.D. 
 
 Professor of Medical Chemistry, Medical Department of 
 Tufts University, Boston 
 
 IS"Ri©GTSinM.ofd([JatX^rris in 
 Laboratory ^sG<arn£llivUffiiMer<ae>hyiyrawdicai 
 
 Department of Tufts University, Boston 
 
 There are no known copyright restrictions in 
 the United States on the use of 
 
 LAMSON, WOLFFE AND COMPANY 
 Boston, New York, London Mdcccxcviii 
 
 http://www.archive.org/details/cu31924000913867 
 
CLu/T* - 
 
 yright, 1898 
 By L.amson, Wolffe and Company 
 
 All rights reserved 
 
 A?3 
 
 
 
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 € 
 
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 PRESS OF 
 
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Table of Contents. 
 
 Page 
 
 Preface ix 
 
 PART I. 
 
 Physiological Chemistry. 
 
 I. Egg Albumen 3 
 
 II. Nucleo-Proteids 6 
 
 Xucleo-Albumen 8 
 
 III. Albuminoides 9 
 
 IV. Blood-Serum 10 
 
 V. Albumoses 12 
 
 VI. Sugars and Starches . . . . 13, 16 
 VII. Fats, Fatty Acids, and Glyc- 
 erine 17," 19, 20 
 
 VIII. Chemistry of Blood 21 
 
 IX. Clinical Analysis of Blood ... 26 
 
 X. Saliva 32 
 
 XI. Chemistry of Gastric Juice . . 33 
 XII. Clinical Analysis of the Stomach 
 
 Contents 37 
 
 Xm. Bile 41 
 
 XrV\ Pancreatic Juice 43 
 
 (v) 
 
vi Contents 
 
 PART n. 
 
 The Urine. 
 
 Page 
 
 I. General Characteristics .... 49 
 II. General Chemistry and Clinical 
 
 Analysis 50 
 
 III. Sediment 60 
 
 part m. 
 
 Toxicology. 
 
 I. Acids : 
 
 Sulphuric 67 
 
 Nitric 67 
 
 Hydrochloric 68 
 
 Oxalic .69 
 
 Carbolic 69 
 
 Hydrocyanic 70 
 
 II. Metallic Poisons : 
 
 Arsenic .71 
 
 Antimony 72 
 
 Mercury 73 
 
 HI. Metallic Poisons (continued) : 
 
 Copper 74 
 
 Lead 75 
 
Contents vii 
 
 Page 
 
 Phosphorus 76 
 
 Nitro-Benzol 76 
 
 IV. Alkaloids: 
 
 Morphine 78 
 
 Strychnine 78 
 
 Veratrine 79 
 
 Atropine 79 
 
 Cocaine 80 
 
 Stas-Otto Method 80 
 
 V. Scheme for the Detection of an Un- 
 known Poison 82 
 
 Appendix. 
 
 Reagents 87 
 
 Standard Solutions 92 
 
 Scheme for Analysis of Blood . . . . 95 
 
 Scheme for Analysis of Gastric Contents . 96 
 
 Scheme for Analysis of Urine . . . . 97 
 
Preface. 
 
 To offer to the medical profession in general 
 and to the medical student in particular a 
 new work embodying laboratory methods of 
 clinical chemistry and toxicology, where so 
 many admirable treatises are in existence, may 
 appear at first sight an unprofitable and 
 useless labor. But all such previous works 
 have either been too voluminous for general 
 use, or too complicated to serve as an ordinary 
 working manual. What the student needs 
 is a work brief enough to give him a clear 
 understanding of the methods of chemical 
 diagnosis, and broad enough so that its pages 
 may comprise all that he will need in the 
 intelligent practice of his profession. While 
 this little work does not pretend to be a com- 
 plete working manual of medical chemistry 
 and toxicology, if it accomplishes the purpose 
 for which it was written, that of leading the 
 fix) 
 
x Preface 
 
 student by simple steps to the more compli- 
 cated methods of clinical chemical diagnosis, 
 the authors will be thankful that their labors 
 have not been in vain. Our thanks are due 
 to Dr. Thorpe for his many useful suggestions 
 and kind assistance while in preparation of 
 this manual. 
 
 A. E. Austin. 
 I. H. Com at. 
 
 Boston, July, 1898. 
 
PART I. 
 PHYSIOLOGICAL CHEMISTRY. 
 
I. 
 
 Egg Albumen. 
 
 Take 20 c.c. egg albumen (white of egg) 
 and shake well in a flask with 150 c.c. of 
 water. Filter, and the filtrate, apart from a 
 slight opalescence, should be wholly clear. 
 With the filtered fluid try the following tests : 
 
 1. Heat a portion in a test-tube to boiling, 
 and there is a white cloudiness, but a separa- 
 tion of coagulated albumen does not take place. 
 This follows by the careful addition of dilute 
 acetic acid (10%), drop by drop. The reac- 
 tion of all albumens must be acid for coagu- 
 lation. 
 
 2. To a portion in a test-tube add one-half 
 the amount of concentrated salt solution and 
 divide into two parts. Heat one part to boil- 
 ing by itself, and there occurs a flocculent pp, 
 or coagulation. To the other part add acetic 
 acid until slightly acid, and it becomes cloudy 
 in the cold, but gives on heating a flocculent 
 pp. The greater amount of salt an albumen 
 solution contains, so much more independent 
 of the acidity is the coagulation by heating. 
 
I. 
 
 Egg Albumen. 
 
 Take 20 c.c. egg albumen (white of egg) 
 and shake well in a flask with 150 c.c. of 
 water. Filter, and the filtrate, apart from a 
 slight opalescence, should be wholly clear. 
 With the filtered fluid try the following tests : 
 
 1. Heat a portion in a test-tube to boiling, 
 and there is a white cloudiness, but a separa- 
 tion of coagulated albumen does not take place. 
 This follows by the careful addition of dilute 
 acetic acid (10%), drop by drop. The reac- 
 tion of all albumens must be acid for coagu- 
 lation. 
 
 2. To a portion in a test-tube add one-half 
 the amount of concentrated salt solution and 
 divide into two parts. Heat one part to boil- 
 ing by itself, and there occurs a flocculent pp, 
 or coagulation. To the other part add acetic 
 acid until slightly acid, and it becomes cloudy 
 in the cold, but gives on heating a flocculent 
 pp. The greater amount of salt an albumen 
 solution contains, so much more independent 
 of the acidity is the coagulation by heating. 
 
A Laboratory Manual 
 
 3. To a portion in a test-tube add glacial 
 acetic acid, and heat. Acid albumen is formed. 
 Now cool the tube and add NaOH, and a pp. is 
 formed, while the reaction is still slightly- 
 acid; but if NaOH be added in excess, the 
 pp. is dissolved and alkali albumen is formed. 
 
 4. Heat a portion with half the amount of 
 NaOH, and there is formed alkali albumen. 
 Cool and add dilute H 2 S0 4 — the albumen is 
 pp., but again dissolved in an excess of the re- 
 agent. 
 
 5. To a portion in a test-tube add CuS0 4 , 
 and there arises a bluish-white pp. of copper 
 albuminate, which, on the addition of NaOH, 
 is dissolved in a deep-blue fluid. The salts of 
 the other heavy metals in general give pps. 
 
 6. To a portion in a test-tube add HgCl 2 . 
 There arises a thick, white pp. which is insol- 
 uble in excess of the reagent, but soluble in 
 cone, salt solution. 
 
 7. Heat a sample with strong NaOH and 3 
 drops of neutral lead acetate. The solution is 
 blackened, due to the separation of sulphur. 
 Upon acidifying with HC1, there is a separa- 
 tion of large black flakes, while in serum albu- 
 men there is only a cloudy gray fluid. This 
 depends upon the fact that the sulphur in egg 
 albumen is separated in larger quantities, and 
 
Of Physiological Chemistry 5 
 
 the lead albuminate is less soluble in HC1 than 
 that of serum albumen. 
 
 8. Shake a portion with ether, and there is a 
 complete coagulation. 
 
 9. To a small portion add cone. HNO s , and 
 there arises a pp. which is not dissolved even 
 when the amount of acid equals half the amount 
 of albumen solution. To this end it is neces- 
 sary to add much larger quantities or to warm. 
 
 Color Tests for Albumen. 
 
 10. Biuret Test. — To a portion in a test- 
 tube add an equal amount of NaOH, and 
 then a few drops of a very dilute solution 
 CuS0 4 . (One drop CuS0 4 to a test-tube of 
 water.) Purple to amethyst color. 
 
 11. Millon's Test. — To a portion add a few 
 drops of Millon's reagent, and boil. Brick-red 
 pp. 
 
 12. Adamkiewicz Test. — Take one part 
 cone. H 2 S0 4 and two parts of acetic acid. Add • 
 the albumen solution carefully without shaking, 
 so that it shall overlay the acids. Warm 
 slightly, and at the point of contact there 
 develops a beautiful purple-red zone, which 
 grows more intense in standing. 
 
 13. Xantho Proteic Reaction. — To a por- 
 tion add strong UNO-, until the pp. which is 
 
A Laboratory Manual 
 
 formed no longer dissolves. Heat, and the pp. 
 becomes colored yellow. Now cool and add 
 NaOH in excess — color changes to orange. 
 
 Now take 1 part of the albumen solution 
 and nine of water and perform all of these tests 
 again. Note particularly the action with Cu 
 (used in sugar tests) — with Hg (used in urea 
 test) — acetic acid (used in mucus test) — 
 and with Pb (action similar to that of bismuth, 
 also used in sugar test) . 
 
 n. 
 
 Nucleo-Proteids. 
 
 Take £ cake of yeast, place in a mortar, add 
 50 c.c. of water and 3 to 5 drops of a very 
 dilute solution NaOH (until neutral or 
 ■slightly alkaline). Rub well with a pestle, 
 filter, and with the filtrate perform the fol- 
 lowing tests : 
 
 1. Heat a small portion to boiling in a test- 
 tube. It should coagulate. How does it differ 
 from egg albumen? 
 
 2. Add to a portion, in a test-tube, dilute 
 (10%) acetic acid. There arises a pp. which 
 is soluble in excess of the reagent. 
 
 3. Take 25 c.c. of the solution and add 
 AgN0 3 until all the NaCl is precipitated. Filter, 
 
Of Physiological Chemistry 7 
 
 pass H 2 S through the filtrate until all the silver 
 is precipitated as sulphide of silver. Warm 
 and filter again (filter more than once if 
 necessary, in order to get a clear filtrate). 
 
 Take 10 c.c. of this solution and add an equal 
 amount of dilute H 3 S0 4 , and warm for some 
 time (at least 20 minutes) upon the water-bath. 
 Cool, alkalinize with NH 4 OH (there must be 
 an excess), and filter. To the filtrate add 
 silver nitrate — and there arises a white pp. of 
 xanthine bodies. 
 
 4. Take 10 c.c. of the solution, add 2 c.c. 
 of cone. HC1, and cook for some time upon the 
 water-bath, replacing the water as it evaporates. 
 Then cool, neutralize with NaOH, add a few 
 drops of CuSo 4 , and heat again to boiling. 
 There arises a separation of red or yellow sub- 
 oxide of copper, which is best seen upon plung- 
 ing the tube into cold water. This is a 
 reducing body, but it is probably not sugar. 
 
 5. With a small portion try the biuret reac- 
 tion. Purple to amethyst color. 
 
 6. Try Millon's reagent. Brick-red pp. 
 
 7. Add alcohol to a portion and note the 
 effect . Coagulati on ? 
 
 8. Saturate a portion with MgS0 4 in sub- 
 stance, wanning slightly to effect complete 
 solution. Note effect. Precipitation? 
 
8 A Laboratory Manual 
 
 9. Take two yeast-cakes, make a solution of 
 them in the same way as given above, and pre- 
 cipitate the nucleo-proteid by warming and the 
 careful addition of dilute acetic acid. Decant 
 off the fluid, filter, and wash the pp. with water, 
 alcohol, and ether. After thoroughly drying, 
 rub up the pp. with 4 grammes saltpeter 
 mixture, 1 heat to melting in a crucible, dissolve 
 the residue after cooling in nitric acid, and 
 heat until the resulting nitrous acid fumes have 
 been driven off. Take 10 c.c. of ammonium 
 molybdate solution, and add to it the nitric 
 acid solution drop by drop. A yellow pp. 
 shows the presence of phosphorus in the form 
 of the acid. To another portion of the solution 
 add NH 4 OH in excess ; it should remain clear. 
 Now acidify slightly with acetic acid and add 
 uranyl nitrate solution. There is formed a 
 yellowish-white pp. if phosphoric acid is 
 present. 
 
 Nudeo-Albumen. 
 
 Make a solution of casein (2 grammes of 
 casein to 100 c.c. of water) and perform the 
 same tests as for nucleo-proteids. Note 
 whether micleo-albumens contain — 
 [ (1) Xanthine Bodies. 
 
 1 This mixture is composed of 3 parts KST0 3 and 1 part NaijCOj. 
 
Of Physiological Chemistry 
 
 (2) A Keducing Body. 
 
 (3) Phosphorus. 
 
 m. 
 
 Albuminoides. 
 
 Take 5 grammes gelatin, add to it 100 c.c. 
 water, and warm. It dissolves, but upon 
 cooling it becomes semisolid, thus behaving 
 directly opposite to albumen, which coagulates 
 by heat. With the warm solution perform the 
 following tests : 
 
 1. To a portion add a few drops of Millon's 
 reagent, and boil. Brick-red pp. This is due 
 to the gelatin containing albumen as an im- 
 purity, as pure gelatin does not respond to 
 Millon's test. 
 
 2. To another portion add an equal amount 
 of NaOH and a few drops of weak copper 
 sulphate solution. Biuret reaction. Amethyst 
 color. 
 
 3. Add nitric acid, and warm. Yellow color. 
 Cool, and alkalinize with NaOH. Orange 
 color. Xanthoproteic reaction. 
 
 4. Cook a portion with NaOH and a few 
 drops of subacetate of lead. Although gelatin 
 
io A Laboratory Manual 
 
 contains sulphur, there is no blackening, as the 
 sulphur is less easily split off than in egg 
 albumen. 
 
 5. To a portion in 4 test-tubes add respec- 
 tively : . 
 
 (a.) NUtTc acid.. No pp. 
 
 (6.) Potassium ferrocyanide. No pp. 
 
 (c.) Acetic acid. No pp. 
 
 (d.) Copper sulphate. No pp. 
 
 6. To a portion add some mercuric chloride. 
 There is no pp. formed. Now add a few drops 
 of HC1, and there is formed a pp. which is 
 soluble in an excess of the acid. 
 
 7. To a portion add some tannic acid. There 
 is no pp. Now add some NaCl, and there is 
 formed a copious pp. 
 
 8. Try the Adamkiewicz reaction. A nega- 
 tive result with pure gelatin. 
 
 IV. 
 Blood-Serum. 
 
 Blood-serum, as well as serous exudations, 
 contains an albumen soluble in water — the 
 serum albumen; and one which is insoluble 
 in water — the serum globulin. This latter is 
 
Of Physiological Chemistry n 
 
 held in solution by the alkaline reaction and 
 the salts of the serum. 
 
 Rub 20 c.c. of blood-serum in a mortar with 
 an excess of ammonium sulphate (about 15 
 grammes) for a long time, in order that the 
 fluid may be surely saturated with the salt. 
 By this means both albumens are precipitated. 
 Filter through a dry filter, and the filtrate is 
 albumen free. Heated to boiling and treated 
 with acetic acid, it remains clear. Now wash 
 the pp. with a half-saturated solution of ammo- 
 nium sulphate, and the albumen in pait becomes 
 redissolved, while the globulin remaining upon 
 the filter can easily be dissolved by water, with 
 the aid of the salt mixed with it. 
 
 1. Solution of Globulin. — Add a few drops 
 to half a test-tube of water. Precipitate. Solu- 
 bility in water? 
 
 2. Add a mere trace of very dilute NaOH 
 to the previous pp. in the test-tube, and it 
 becomes redissolved. 
 
 3. Neutralize carefully with dilute HC1, and 
 the globulin becomes again precipitated. If 
 in (2) too much NaOH were added in pro- 
 portion to the amount of globulin, it is not 
 reprecipitated, for the NaCl formed holds it in 
 solution. 
 
 To these respective solutions, (a) serum 
 
12 A Laboratory Manual 
 
 albumen and (b) serum globulin, add the 
 reagents employed for egg albumen. Note 
 how serum albumen and serum globulin differ 
 from each other, and also how they both differ 
 from egg albumen. 
 
 Now take 10 c.c. of blood serum, add water 
 to 100 c.c., and perform the following tests : 
 
 1. Heat to boiling — no change. Add a 
 drop or two of HN0 3 and heat again, and there 
 is a separation of coagulated albumen. 
 
 2. Add acetic acid and potassium ferro- 
 cyanide. Cloudiness — then a pp. 
 
 3. Acidify with HC1 and then add phospho- 
 tungstic acid. Copious gelatinous pp. 
 
 4. Add tannic acid. Copious pp. 
 
 5. Add mercuric chloride. A pp. — which 
 is soluble in NaCl solution. 
 
 6. Add a few drops of Millon's reagent, and 
 boil. Keddish-brown pp. 
 
 V. 
 Albumoses. 
 
 Use solution provided — 2 grammes to 100 
 c.c. of water. 
 
 1. Heat a portion to boiling ; remains un- 
 changed also after adding acetic acid (a drop) 
 and NaCl. 
 
Of Physiological Chemistry 13 
 
 !. Acidify with acetic acid and add cone. 
 ; solution. The solution becomes cloudy, 
 ; upon heating again becomes clear. Upon 
 iling — cloudy. 
 
 J. Add a few drops HN0 3 , and there is a 
 ndincss to a pp. which is soluble in excess, 
 wanning and NaOH — xanthoproteic re- 
 ion. 
 
 I. Add a few drops acetic acid and the po- 
 sium ferrocyanide. Cloudiness, which dis- 
 Dcars on healing (not always wholly). 
 5. Biuret test — rose to purple color. 
 5. Perform the tests (3) — (4) — (5) — 
 ) — of blood-serum. Insoluble precipitates. 
 
 VI. 
 Sugars. 
 
 1. Take a small portion of milk-sugar and 
 it it to melting upon platinum foil or a 
 see of tin plate. There is developed an odor 
 caramel and a carbonizing, with the final 
 .vino - of a slight amount of ash. 
 Make a solution of 2 grammes milk-sugar to 
 c.c. water, and perform the following 
 its: 
 
14 A Laboratory Manual 
 
 2. Trommer's Test. — Add to a few c.c. of 
 the solution an equal amount of NaOH, and, 
 drop by drop, CuS0 4 , shaking constantly. 
 There arises a deep-blue solution, which upon 
 heating gives a separation of red or yellow 
 suboxide of copper. 
 
 3. Moore's Test.— Add to a portion an equal 
 amount of NaOH and heat to boiling ; yellow 
 to brown color, with the odor of caramel, 
 especially after acidification with dilute H 2 S0 4 . 
 
 4. Nylander's Test. — Saturate a portion 
 with sodium carbonate in substance, and boil. 
 While boiling add a very small portion of bis- 
 muth subnitrate and keep for some time at a 
 boiling point. Gray to black color, with the 
 separation of fine metallic bismuth. 
 
 5. Add some ammonia and a few drops of 
 AgN0 3 , and heat. Separation of metallic 
 silver in the form of a glistening mirror upon 
 the test-tube. The mirror is especially marked 
 if a drop of NaOH be added before boiling. 
 As the AgN0 3 in this test is changed to 
 fulminating silver, care must be taken against 
 its possible explosion by afterwards filling the 
 test-tube with water. 
 
 6. Indigo Test. — Add indigo carmine to the 
 sugar solution until distinctly blue, and then 
 add sodium carbonate solution until alkaline, 
 
Of Physiological Chemistry 15 
 
 id warm. The solution becomes first violet, 
 ien red, yellow, and finally colorless. Pour 
 ilf of the solution in another test-tube and 
 lake with air. It again becomes blue, but 
 pon heating is again decolorized. This con- 
 nues until all the sugar is used up by oxida- 
 on. 
 
 7. Phenylhydrazin Test. — Take 25 c.c. of 
 Le solution in a small beaker, and add 1 
 ramme phenylhydi-azin hydrochlorate and 2 
 rammes sodium acetate. Place in boiling 
 ater for 30 minutes, and then cool by plung- 
 ig the beaker in cold water. A yellow 
 •ystalline deposit is thrown down, which, 
 hen examined under the microscope, is found 
 > consist of wide prisms of phenyl-lactosazon. 
 
 Now make a 2% solution of grape-sugar 
 ad perform the same tests. 
 
 8. Fill two fermentation tubes with solutions 
 f grape and milk sugar respectively. Shake 
 ith a piece of fresh yeast until disintegrated, 
 id place in a brood oven until the next day. 
 ote the difference. Milk-sugar ferments 
 owly and with difficulty. 
 
 9. Grape-sugar can be dissolved in cone. 
 [ 2 SO without color; milk-sugar is colored 
 rown when dissolved in the same acid. 
 
 10. Phenylhydrazin Test. — Bright-yellow 
 
16 A Laboratory Manual 
 
 needles of phenyl-glucosazon, arranged in 
 bundles or sheaves. 
 
 11. In two test-tubes place a few c.c. of 
 grape and milk sugar respectively, and add 
 one or two drops of a 15% alcoholic solution 
 of alpha-naphthol. Then let about 1 c.c. of 
 cone. H 2 S0 4 run carefully down the side of 
 the test-tube so as to underlie the other fluid ; 
 there is formed at the junction of the two fluids 
 a beautiful violet-red ring that increases inten- 
 sity upon slight shaking. 
 
 Make a 2% solution of cane-sugar and try 
 the reducing tests (copper and bismuth) . Cane- 
 sugar will not reduce. 
 
 Try the fermentation test with yeast, the 
 same as for grape and milk sugar. After fer- 
 mentation again try the reducing tests. The 
 cane-sugar is a disaccharide, and, by the action 
 of the invertion of the yeast, has been split up 
 into its two mono-saccharides, glucose and 
 levulose, both of which will reduce. 
 
 Cane-sugar does not form a compound with 
 phenylhydrazin . 
 
 Starches. 
 
 Measure out 100 c.c. water, and with a 
 portion of the same rub up 1 gramme of starch 
 in a mortar. Pour into an evaporating dish 
 
Of Physiological Chemistry 17 
 
 and with the rest of the water wash all the 
 starch into the dish. Now heat to boiling 
 under constant stirring. Allow to cool and 
 perform the following tests : 
 
 1. Add to a portion in a test-tube 3 or 4 
 drops of a solution of iodine and potassium 
 iodide. Blue color. 
 
 2. Boil a portion for 20 minutes with dilute 
 HC1. Cool, and to a portion add iodine 
 solution again. Mahogany-red color due to 
 erythrodextrin. The starch has been converted 
 to erythrodextrin and glucose (grape-sugar). 
 Alkalinize another portion fully, and test for 
 glucose with reducing tests. 
 
 3. Take 10 c.c. of the starch solution, add 
 1 c.c. saliva, and place in a brood oven until 
 the next day. Test for erythrodextrin and 
 maltose. 
 
 vn. 
 
 Fats. 
 
 1. Rub a small portion of lard or oil upon 
 paper (not filter-paper) , and it becomes trans- 
 lucent. 
 
 2. Dissolve a piece of lard the size of a pea 
 in a few c.c. of alcohol with the aid of a little 
 
1 8 A Laboratory Manual 
 
 ether, and add a couple drops rosolic acid, 
 which gives the solution a yellow color. Now 
 add from a burette a ^ Normal sol. NaOH, 
 and one or two drops changes the color to red. 
 
 3. Rub up a small portion in a mortar with 
 bisulphate of potash, and heat the mixture in a 
 dry test-tube. Visible fumes and the pungent 
 odor of acrolein show a neutral fat. 
 
 4. Warm a small portion in a test-tube with 
 sodium carbonate solution. The fat is dis- 
 tributed in the solution, but the latter is not 
 clear ; saponification does not take place. 
 
 5. Dissolve 5 grammes KOH in 35 c.c. alco- 
 hol, by warming over the water bath. Then 
 melt 5 grammes of lard over the water bath, 
 and add to it while stirring the KOH solution. 
 Cook for 20 to 30 minutes, or until a small 
 portion in a test-tube of water shows no oil 
 drops. There is complete saponification. 
 
 6. Add to 65 c.c. water 20 c.c. dil. H 3 S0 4 
 (20%) and warm somewhat, but not to boil- 
 ing. Now add the soap solution, while still 
 hot, carefully to the dilute acid (there is a 
 liability to snap). Allow to cool, and the 
 fatty acids arise to the surface of the fluid as a 
 layer which can be broken through, and the 
 watery solution which contains the glycerine 
 can be poured off. Hot water should be added 
 
Of Physiological Chemistry 19 
 
 this fat, and the process repeated until the 
 sh water is no longer acid. Dry the fatty 
 is between filter-paper. 
 
 Fatty Acids. 
 
 L. Behavior to paper same as neutral fat. 
 5. Make a solution with alcohol and ether, 
 I add NaOH as above (2), using rosolic 
 I as an indicator. It takes 2 or more c.c. 
 change the color to red. They are acid. 
 5. No acrolein by heating with bisulphate of 
 ;ash. 
 
 t. Add to a portion a half-saturated solution 
 sodium carbonate, and heat. The fatty 
 is dissolve with the development of C0 2 
 1 the formation of a sodium soap. Dip the 
 t-tube in water, and the soap solidifies. 
 }. Dissolve 2 grammes of fatty acids in 100 
 . of water, with the addition of the smallest 
 isible quantity of NaOH needed to effect solu- 
 1. With this solution perform the following 
 ts: 
 
 [a.) Add HC1. Separation of fatty acids. 
 b.) Addition of chloride of calcium. In- 
 nble chalk soap. The fluid no longer foams 
 )n shaking. 
 'c.) Add lead acetate, and warm. A white 
 
20 A Laboratory Manual 
 
 pp. which upon warming becomes tough and 
 crumbly — ; " Lead Plaster." 
 
 (d.) Take a little of the solution, add a few 
 drops of oil, and shake. There is formed a fine 
 white emulsion. 
 
 (e.) Try the same thing with sodium car- 
 bonate solution and oil. Negative result. An 
 emulsion can only be formed when some free 
 fatty acids are present, as when a fat becomes 
 rancid. 
 
 (/.) Place some of the fat in a test-tube 
 and an equal amount of the fatty acids in 
 another. Place both tubes in a beaker of 
 water, and heat upon a piece of wire gauze, 
 occasionally stirring the water so that the 
 temperature shall be the same everywhere. 
 It will be noticed that the fat melts much 
 sooner than the fatty acids obtained from the 
 same fat. 
 
 Glycerine. 
 
 Take some of the watery solution from Fats 
 (6), and with it perform — 
 
 1. Trommer's Test. — Blue solution, but no 
 reduction of copper. 
 
 2. Mix some glycerine with a little borax 
 and heat a small portion in the flame upon 
 platinum foil. The flame is temporarily col- 
 ored green. 
 
Of Physiological Chemistry 
 
 21 
 
 Try the acrolein test. 
 Make an emulsion with oil and soap solu- 
 and examine under the microscope. 
 
 vm. 
 
 Chemistry of Blood. 
 
 . Take a piece of lacmoid paper, wet with 
 :. salt solution, and place upon it with a 
 s rod a drop of blood. A blue spot shows 
 alkalinity of blood. 
 
 . Add to about 10 c.c. water a few drops 
 >lood, shaking thoroughly. Then add some 
 h tincture guaiac (made by dissolving a 
 knife-pointful of the powder with some 
 thol in a test-tube) until the solution be- 
 tes milky. Now add some old oil of tur- 
 tine, and upon shaking the solution becomes 
 )red an intense blue. 
 
 ;. Add to a few c.c. blood several times its 
 Lime of peroxide of hydrogen. There arises 
 trong foaming and development of oxygen, 
 h gradual decomposition of the blood-color- 
 substance. This catalytic action of the 
 od is lost in the presence of hydrocyanic 
 1, but the acid must be added to the blood 
 
22 A Laboratory Manual 
 
 directly. In the blood of an animal killed by 
 hydrocyanic acid, this catalytic action persists. 
 
 4. Add to a little blood in a test-tube some 
 ether and water, and shake thoroughly. The 
 coloring matter of the corpuscles becomes dis- 
 solved, and the blood becomes " lakey." The 
 biliary acids have the same action upon blood. 
 
 5. Dilute 10 c.c. blood to 100 c.c. with 
 water and perform the following tests : 
 
 (a.) Take a half test-tube of blood, place 
 in the spectroscope, and continue to add water 
 until the two lines of oxyhemoglobin in green 
 and- yellow can be seen. Now add a few drops 
 of ammonium sulphide, and allow to stand for a 
 few minutes. There is a change of color from 
 red to green, and the appearance of only one 
 line in the spectroscope, that of reduced htemo- 
 globin, occupying a position midway between 
 that of the other two. 
 
 (b. ) Take another portion of the blood solu- 
 tion and add a few drops of freshly prepared 
 cone, solution of potassium ferricyanide. The 
 solution becomes brown and shows the charac- 
 teristic four-line spectrum of methsemoglobin. 
 Now add a few drops of ammonium sulphide, 
 allow to stand for a short time, shake strongly, 
 and again examine with a spectroscope. The 
 spectrum of oxyhemoglobin will be foundi 
 
Of Physiological Chemistry 23 
 
 thtenioglobin can be reduced to haemoglobin, 
 1 again oxydized to oxyhemoglobin. 
 [c.) Pass hydrogen sulphide through some 
 the dilute blood and examine with a spectro- 
 ipe — the spectrum of sulph-htemoglobin. It 
 embles that of methsemoglobin except that 
 : line in red is farther to the left, and that it 
 inot be reduced. Try the reduction test, 
 ae as in (6). 
 
 (d.) Add to 10 c.c. of the dilute blood 
 c.c. NaOH, and heat. The solution be- 
 lies brownish-green. Examination with the 
 metroscope shows one broad band of absorp- 
 n extending to red — the spectrum of hsema- 
 
 in alkaline solution. Now add a few drops 
 immonium sulphide, allow to stand for a short 
 le, and again examine with a spectroscope, 
 e one absorption band has disappeared, and 
 
 its place will be found the two lines of 
 luced haematin. 
 5. Pass ordinary illuminating gas through 
 
 c.c. blood until it becomes a cherry-red. 
 amine with the spectroscope. The spectrum 
 embles that of oxyhemoglobin, except that 
 ;annot be reduced. Try the reducing tests. 
 [a.) Add to this blood and to genuine 
 >od equal amounts (about \ its volume) of 
 OH. Xote the difference ; place a drop of 
 
24 A Laboratory Manual 
 
 each in an evaporating dish, and note difference 
 in color. 
 
 (b.) Add to 10 c.c. carbon monoxide 
 blood 15 c.c. of a 20% solution potassium 
 ferricyanide and 2 c.c. acetic acid — and to the 
 same amount of unmodified blood add the same 
 reagents. Allow to stand a few moments, and 
 note the difference. 
 
 (c.) To 5 c.c. of carbon monoxide blood 
 add four times as much water. To a measured 
 portion of this add three times its volume of a 
 1% tannic solution. Do the same with un- 
 modified blood, and note the difference. 
 
 7. Dry a drop or two of blood upon a micro- 
 scope slide, and rub up the dried blood with 
 the smallest possible quantity df sodium chlo- 
 ride. Place a cover glass upofij it, and add a 
 couple drops of glacial acetic aciWl to the edge 
 of the cover glass, allowin>*Jdne a«id to flow 
 under the latter. Nov warm upon the water 
 bath until evaporated,^ >dd a drop or two more 
 of the acid, and alio?' to cool. Examine 
 with a micro'scopew'fl" or J" objective). 
 Small dark-brown, rhombic crystals of hsemin 
 chloride show the presence of blood. Stains 
 upon linen or wood can be soaked out in a 
 normal salt solution and tested in the same 
 way. These crystals can be obtained from 
 
Of Physiological Chemistry 25 
 
 e traces of blood, and their presence is eon- 
 ive of the presence of blood in medico-legal 
 ni nations. 
 
 . Take 'M) c.c. fresh blood, add 8 times as 
 :h water, and heat to boiling, with constant 
 ring. While boiling add carefully dilute 
 f ( ) acetic acid until slightly acid, and 
 r. The nitrate must be clear. Evaporate 
 filtrate to a small volume, and divide into 
 
 halves. With one, try Trommer's Test 
 sugar. The other half is to be still further 
 centrated, and a drop allowed to evaporate 
 n an object glass, and examined with a 
 roscope. Crystals of NaC'l. 
 'he rest is to be evaporated to dryness in 
 evaporating dish, heated to redness, cooled, 
 
 dissolved in a little water. Examine for 
 ium chloride with AgN0 3 — for phosphates 
 h ammonium molybdate. 
 f, in the above, after heating to redness, a 
 3k residue remains, add a little saltpetre 
 dure, and heat to redness again. After 
 ling, dissolve the residue, which should be 
 arless, in nitric acid, evaporate oft' the 
 ess of the acid, and test for phosphates 
 L chlorides as above. 
 
 >. Dry the pp. formed in (H) , take 1 gramme 
 ;he dried pp. and rub thoroughly in a mortar 
 
26 A Laboratory Manual 
 
 with 4 grammes saltpeter mixture. Heat to 
 melting in a crucible for about 20 minutes, and 
 after cooling dissolve the residue in dilute sul- 
 phuric acid (8 parts by weight cone. H 2 S0 4 -|- 3 
 parts by weight of water). Filter, and to a 
 portion of the filtrate, which should be clear, 
 add some potassium sulphocyanide. A blood- 
 red color shows iron. 
 
 IX. 
 
 Clinical Analysis of Blood. 
 
 Specific Gravity (Roy's method) . — A num- 
 ber of test-tubes are two-thirds filled with a 
 mixture of glycerine and water in different 
 proportions, so that the specific gravity in the 
 tubes shall vary between 1030 and 1070. 
 Blood is then drawn from the tip of the -finger 
 or the lobe of the ear into a capillary tube con- 
 nected with an ordinary hypodermic syringe. 
 A drop of blood is placed in each tube, in 
 which it will sink as long as the specific gravity 
 of the glycerine mixture is lower than that of 
 the blood, while it will remain suspended in a 
 mixture the specific gravity of which is equiv- 
 alent to its own. 
 
 Normal specific gravity — 1.046 — 1.067. 
 
Of Physiological Chemistry 27 
 
 Alkaliuity (Lowy's method). — Five c.c. of 
 fibrinated blood, accurately measured, are 
 ixed in a small flask with 45 c.c. water; if 
 t defibrinated, the blood is mixed with the 
 me amount of a 0.25% solution ammonium 
 alate. Titrate the solution with a 1/25 nor- 
 il solution of tartaric acid, until a drop 
 aced upon a piece of lacmoid paper (which has 
 en previously soaked in a cone, solution of 
 ignesium sulphate) gives rise to a blue tinge. 
 Calculation — 1 c.c. ^ normal solution 
 rtaric acid = .0016 grammes NaOH. 
 Xormal alkalinity in terms of sodium hydrate 
 182 to 218 milligrammes for every 100 c.c. 
 blood. 
 
 Quantitative Estimation of Haemoglobin 
 ? leisehl's method) . — The instrument used is 
 e hasmometer of Fleischl, and the principle 
 ' the method depends upon the comparison of 
 e color of the blood, diluted with water, with 
 at of a glass wedge stained with a red pig- 
 ent. The principal part of the instrument 
 insists of two semicircular compartments, 
 le receiving light from a plaster-of-Paris 
 Sector through the glass wedge and called 
 e red chamber, the other receiving light 
 rectly from the reflector and called the white 
 lamber. 
 
28 A Laboratory Manual 
 
 To use the instrument, the two compart- 
 ments are partially filled with water, and the 
 required amount of blood is obtained by placing 
 one end of the capillary pipette in contact with 
 a drop of blood obtained from the tip of the 
 finger or the lobe of the ear. The pipette is 
 immersed in the white chamber and rotated 
 between the fingers. By this method the blood 
 passes into the water, and any traces of blood 
 remaining in the pipette are carefully washed 
 out. The two compartments are then com- 
 pletely filled with water, and the glass wedge 
 so adjusted, by means of the screw, that the 
 color in the two chambers shall be the same. 
 The number facing the notch in the scale aper- 
 ture of the platform will then indicate the per- 
 centage of haemoglobin, that of a healthy 
 individual corresponding to 100. As the nor- 
 mal amount of haemoglobin contained in 100 
 grammes of blood is 13.7 grammes, the number 
 100 on the scale of Fleischl's instrument corre- 
 sponds to 13.7%. 
 
 Calculate % according to equation : 
 100: 13.7 : : reading on the scale : X 
 
 The Enumeration of the Red Blood Cor- 
 puscles (Thoma-Zeiss method) . — The instru- 
 ment used is the Thoma-Zeiss blood-counting 
 apparatus, 
 
Of Physiological Chemistry 29 
 
 Blood is drawn from the tip of the finger or 
 lobe of the ear into the capillary pipette, to the 
 mark 1 or 0.5, according to the degree of dilu- 
 tion desired. The blood is then dilated with 
 Pacini's fluid, which is drawn into the pipette, 
 past the bulb, until the mark 101 is reached. 
 If the blood had previously been drawn to the 
 mark 0.5, it now has been diluted 200 times ; 
 if to the mark 1, it has been diluted 100 
 times. The contents of the bulb are now 
 thoroughly mixed by shaking. A drop of the 
 mixture is placed in the counting-chamber, so 
 that it shall completely fill it, and the cover- 
 slip is adjusted. The slide is placed under the 
 microscope, a \" or |" objective being used. 
 Now focus until the small squares come dis- 
 tinctly into view. Each one of these small 
 squares is fa m.m. long, fa m.m. wide, and 
 fa m.m. deep ; its cubic contents are therefore 
 Wffff cubic m.m. Count the corpuscles in at 
 least 16 small squares and take the average, 
 which will give the number of corpuscles in one 
 small square, or in ^fa-^ cubic m.m. One cubic 
 m.m. of this diluted blood will then contain 
 4,000 times the number contained in one small 
 square, and one cubic m.m. of the undiluted 
 blood will contain the product of this figure 
 multiplied by the degree of the original dilution. 
 
30 A Laboratory Manual 
 
 The Enumeration of the Red Blood Cor- 
 puscles by the Hasmatokrit. — The graduated 
 glass tube is completely filled, by suction, with 
 blood from the finger or ear. The blunt point 
 of the tube is then quickly covered with the 
 finger, and the tube fixed in the frame of the 
 centrifuge. This is rotated for two to three 
 minutes, and the number of corpuscles which 
 have been thrown to the bottom of the tube 
 are read off. Multiply the reading on the 
 tube by 100,000, and the product is the num- 
 ber of red corpuscles in one cubic rn.m. 
 
 The Enumeration of the White Blood Cor- 
 puscles (Thoma-Zeiss method) . — This enu- 
 meration is done with the same apparatus as 
 for the red corpuscles, except that a special 
 diluting pipette is used. The diluting fluid 
 is a 0.3% solution acetic acid colored with 
 methyl-violet, which destroys the red cor- 
 puscles and brings out the white very dis- 
 tinctly. The pipette is filled with blood to the 
 mark 0.5 if we wish to dilute 20 times, or to 
 1 if a dilution of 10 times is desired. Then 
 fill the pipette to the mark 11 with the acetic 
 acid solution. Place a drop in the Thoma- 
 Zeiss cell, use a \" or \" objective, and focus 
 as above. As the number of white corpuscles 
 is smaller than the red, it is necessary to count 
 
Of Physiological Chemistry 31 
 
 number of corpuscles in a whole field ; 
 t is, in the area of one microscopic circle, 
 jn determine the cubic contents of this mi- 
 scopic circle, and from this calculate the 
 ire number in one cubic m.m. 
 Dhe cubic contents are determined as follows : 
 int the number of cross vertical lines in the 
 i of the microscope, remembering that 
 
 distance between each two lines is fa m.m. 
 is gives the diameter of the circle. Divide 
 
 diameter by 2, which gives the radius ; 
 :are the radius, and multiply the square of 
 
 radius by 3.1416. The product is the area 
 the circle, and this area, multiplied by 0.1, 
 ich is the depth of the cell in a fraction of a 
 ilimeter, gives the cubic contents of the 
 jroscopic circle. The number of white cor- 
 acles counted in this single field is there- 
 e contained in the fraction of one cubic m.m. 
 iresented by the cubic contents of the field 
 mted. From this calculate the number of 
 ite blood corpuscles in one cubic m.m. of 
 iited blood, and this, multiplied by the de- 
 se of dilution (10 or 20), will give the entire 
 tnber in one cubic m.m. 
 Formula for the determination of the cubic 
 itents of the circle : 
 Radius 2 x 3.1416 x fa — cub io contents of 
 
32 A Laboratory Manual 
 
 X. 
 Saliva. 
 
 1. Add to some saliva in a test-tube a few 
 drops of dilute (10%) acetic acid. Cloudi- 
 ness due to mucin — not soluble in excess of 
 the acid. 
 
 2. Add to saliva a few drops HN0 3 . A 
 cloudiness which becomes yellow upon heat- 
 ing. Cool and alkalinize with NaOH — orange 
 color. This xanthoproteic reaction is due to 
 mucin or an albuminoid body. 
 
 3. To a small portion of saliva add an equal 
 amount of water and a few drops Millon's 
 reagent. White pp., which becomes red upon 
 warming. Due to mucin. 
 
 4. Add NaOH and a few drops of a very 
 dilute CuS0 4 solution. Violet color due to 
 mucin. 
 
 5. Add to a portion one drop of HC1 
 and a few drops of a very dilute solution of 
 ferric chloride, and shake thoroughly. Red 
 color from the formation of ferric sulphocyan- 
 ide. Saliva contains potassium sulphocyanide 
 (KCNS), its presence being due to the gland 
 containing adenin (C 5 H 6 "NT 5 ), which is a poly- 
 mere of hydrocyanic acid (CHN) . 
 
Of Physiological Chemistry 33 
 
 6. Make a starch clyster ( 1 gramme starch to 
 100 c.c. water), and, after cooling, take lOc.c. 
 of this solution, add 1 c.c. of saliva, and digest 
 at 40° C. for an hour. Then put into a fermen- 
 tation tube and allow to stand until the next 
 day. It will be found that the resulting sub- 
 stance (maltose) is fermentable. 
 
 7. Perform the same test as in (6), but add 
 to the starch and saliva mixture 1 c.c. digestive 
 HC1. The action of the ptyalin is annulled. 
 Prove by Trommer's test and the fermentation 
 tube. 
 
 8. Determine the reaction of saliva. It is 
 normally alkaline, but in pathological condi- 
 tions, as in fevers or diabetes, it is acid. 
 
 9. Examine some saliva under a micro- 
 scope, and it will be found to contain certain 
 morphological elements, such as salivary cor- 
 puscles, pavement epithelial cells, and various 
 fungi and bacteria. 
 
 XL 
 Chemistry of Gastric Juice. 
 
 Hydrochloric Acid. — 1. (a.) Add to some 
 digestive hydrochloric acid in a test-tube a few 
 drops of methyl-violet solution. Steel-blue 
 color. 
 
34 A Laboratory Manual 
 
 (b.) Make the same test with water. Violet 
 color. 
 
 (c.) Make the same test with lactic acid. 
 Violet color, with slight shade of blue. 
 
 2. Make the same test with HC1 solution, 
 diluted 1 to 5. 
 
 3. Take a portion of the HC1 solution in 
 2 test-tubes. To one add an equal amount of 
 water, to the other an equal amount of albu- 
 mose-peptone solution ; then add methyl-vio- 
 let to both and compare the color. 
 
 4. Make the same tests with the HC1 
 solution diluted 1 to 5. The presence of large 
 amounts of peptone makes methyl-violet use- 
 less as a reagent. 
 
 5. Add to a portion of the lactic acid solution 
 a few drops of methyl-violet, and divide into 
 three parts. To No. 1 add an equal amount of 
 water; to No. 2 add an equal amount 3% salt 
 solution ; to No. 3 add an equal amount cone, 
 salt solution. Nos. 1 and 2 change color but 
 slightly. No. 3, however, becomes a distinct 
 steel-blue. Sodium chloride disturbs the reac- 
 tion in the presence of lactic acid only when 
 strongly concentrated, due to the separation 
 of the HC1 by the lactic acid. 
 
Of Physiological Chemistry 35 
 
 6. Evaporate a few drops of the mixtures 
 formed under 2, 3, 4, and 5, in a small evapo- 
 rating dish, to dryness over a free flame, 
 avoiding too strong a heat. Intense blue resi- 
 due. Peptones disturb the reaction only when 
 present in large quantities. 
 
 7. To a few drops of the digestive HC1 in 
 an evaporating dish add a few drops of tro- 
 psolin 00 solution, and warm gently. Lilac- 
 blue color. 
 
 8. Add to a few drops of digestive HC1 
 in an evaporating dish a drop of Gunzburg's 
 reagent, and evaporate to dryness over a free 
 flame, avoiding too strong a heat. Beautiful 
 purple-red residue. Make the same tests with 
 2, 3, 4, and 5. Peptone does not disturb the 
 reaction and lactic acid does not give it. 
 
 9. To a little HC1 in a test-tube add a 
 drop of dimethyl-amido-azo-benzol. Brilliant 
 red color. 
 
 10. Take two test-tubes, and in No. 1 place 
 a few c.c. digestive HC1 diluted 10 times ; in 
 the other place an equal amount of water. To 
 each add a drop of dimethyl-amido-azo-benzol. 
 No. 1 becomes a brilliant red; No. 2 becomes 
 cloudy and yellow. 
 
 Lactic Acid. — The reagent used for the de- 
 tection of lactic acid is Uffelmann's, made by 
 
36 A Laboratory Manual 
 
 taking 10 c.c. of a 5 % carbolic acid solution and 
 adding a few drops ferric chloride. Amethyst- 
 blue fluid. 
 
 1. To a portion of the reagent add a few 
 drops digestive HC1. Decolorization. 
 
 2. To another portion add some lactic acid. 
 Canary-yellow color. 
 
 3. To another portion add a mixture of 
 equal parts of digestive HC1 and lactic acid 
 solution. Canary-yellow color. With Uffel- 
 mann's reagent lactic acid can be detected 
 beside HC1, but not HC1 beside lactic acid. 
 
 4. To Uffelmann's reagent add some lactic 
 acid and place in 3 test-tubes. To No. 1 add 
 an equal amount of water ; to No. 2 an equal 
 amount cone. NaCl solution ; to No. 3 an 
 equal amount 3% NaCl solution. Only No. 
 2 becomes decolorized. The presence of NaCl 
 in small quantities does not disturb the de- 
 tection of lactic acid. 
 
 Pepsin. — 1. Place in 3 test-tubes thin slices 
 of hard-boiled egg of equal size. To No. 1 
 add 10 c.c. digestive HO ; to No. 2 add 5 c.c. 
 digestive HC1 with pepsin ; to No. 3 add 10 c.c. 
 digestive HC1 with pepsin. Place all three 
 in the brood oven for an hour or more, and 
 note the difference in the thinning of the slices, 
 due to the digestive action of the three fluids. 
 
Of Physiological Chemistry 37 
 
 2. Take the whites of i hard-boiled eggs, 
 cut fine, and place in a flask, adding 500 c.c. 
 of the digestive HC1 with pepsin (1 gramme 
 pepsin, 10 e.e. HC1, and water to 500 c.c.). 
 Place in a hrood oven for 2 to 3 days, shaking 
 occasionally, filter, neutralize the filtrate, while 
 warming gently, with Xa_,('0 3 , and filter again 
 from the acid albumen thus precipitated. Then 
 evaporate to about 200 c.c, filtering again 
 before reaching this point if more albumen 
 becomes separated. The reaction meanwhile 
 must be kept neutral by adding HOI or 
 Xa 2 C0 3 , as the case should require. Now acid- 
 ify the solution with acetic acid, saturate with 
 XaCl, and filter. The filtrate contains pep- 
 tones. Test for the same. 
 
 xn. 
 
 Clinical Analysis of the 
 Stomach Contents. 
 
 For clinical examination the stomach con- 
 tents should be filtered, or if too thick and 
 diflicult of filtration, they should be diluted 
 with a definite amount of distilled water. In 
 quantitative estimations the amount should be 
 measured with a pipette and not a graduate. 
 
38 A Laboratory Manual 
 
 1. Presence of Pepsin. — Take 10 c.c. of 
 the stomach contents, place in a test-tube, and 
 add a thin slice of hard-boiled egg or fibrin. 
 Place in a brood oven, and allow to remain a 
 few hours. If after 10 hours there is no dim- 
 inution of the size of the egg, or if there is 
 an odor of decomposition, then pepsin is 
 absent. If the stomach contents are weakly 
 acid or alkaline, add an equal amount of diges- 
 tive HC1. 
 
 2. Presence of Rennet. — Take 5 c.c. neu- 
 tralized stomach contents, and add to it an 
 equal amount of cooked, neutral cow's-milk, 
 which has been fully cooled after cooking. 
 Place in a brood oven, and if rennet be 
 present, after 20 or 30 minutes the casein of 
 the milk will have separated out in flocculent 
 masses. 
 
 3. Presence of Free HC1. — Use methyl- 
 violet — tropseolin 00 — Griinzburg's reagent or 
 dimethyl-amido-azo-benzol . 
 
 4. Quantitative Estimation of Total Acid- 
 ity.— Take 10 c.c. of the filtered stomach 
 contents, add a few drops of rosolic acid until 
 yellow, and titrate with a ^ normal solution 
 NaOH until a permanent rose-red color is 
 obtained. 
 
 Calculation. — 1 c.c. NaOH = .00365 
 
Of Physiological Chemistry 39 
 
 5. Quantitative Estimation of Total HC1 
 
 (free and combined). 
 
 Take 15 c.c. of the filtered stomach contents, 
 add 1 gramme calcium carbonate, stir well, and 
 filter. Take 10 c.c. of the filtrate, place in a 
 small flask, and pass air through until the 
 C0 2 is all driven off. Then add 5 c.c. cal- 
 cium chloride, a few drops of rosolic acid, and 
 titrate with ^ normal NaOH as before. The 
 difference between the acidity found before and 
 that found now equals the total HC1 (free and 
 combined). 
 
 1 c.c. XaOH = .00365 grammes HC1. 
 
 The principle of this method is based upon 
 the fact that the calcium carbonate combines 
 with the free and combined HC1 and the 
 organic acids to form neutral calcium salts, 
 while the acid phosphates are not affected ; 
 and by determining as in (4) the total acidity, 
 and deducting from this the acidity due to the 
 acid salts, the amount of total HC1 is obtained. 
 
 6. Test for lactic acid by Uffelmann's re- 
 agent. 
 
 7. Acetic, Butyric, and Formic Acids. — Dis- 
 til a portion of the stomach contents, and, if 
 present, these acids will be found in the dis- 
 tillate. 
 
 (a.) Test for Acetic Acid. — Accurately neu- 
 
4-0 A Laboratory Manual 
 
 tralize a portion of the distillate with NaOH 
 and then add ferric chloride. Red color, if 
 present. 
 
 (5.) Tests for Butyric Acid. — 1. Add ferric 
 chloride. Red color, which disappears on heat- 
 ing. 2. Add cone. E^SC^, and warm. Charac- 
 teristic odor of rancid butter. 
 
 (c. ) Test for Formic Acid. — Add silver nitrate. 
 White pp., which turns black in the dark. 
 
 8. Altmmose Peptone. — Neutralize, while 
 warming, with Na^CC^, filter, and test filtrate 
 by biuret test. 
 
 9. Starch and. Erythrodextrine. — Test with 
 a solution of iodine in potassium iodide. They 
 should not be present an hour after food is 
 taken. 
 
 10. Kesorptive Power of the Stomach. — 
 Give a capsule containing 0.1 gramme potas- 
 sium iodide, and test the saliva every few 
 minutes with filter-paper moistened with starch 
 solution until a blue color is obtained. It 
 ought to be absorbed in from 8 to 15 minutes in 
 a normal stomach. 
 
 11. Motor Power of the Stomach. — A 
 capsule containing 1 gramme of salol is given, 
 and separate portions of the urine, passed within 
 intervals of half an hour, are tested by ferric 
 chloride. The violet color, due to salicyluric 
 
Of Physiological Chemistry 41 
 
 acid, is obtained, under normal conditions, in 
 from 45 to 75 minutes. 
 
 Microscopic Examination. 
 
 Sarcinae. — They occur in the form of pecul- 
 iar colonies of cocci, arranged in squares, and 
 strongly resembling cotton-bales. In normal 
 conditions they are found only in small num- 
 bers. 
 
 Yeast-Fungi — have a characteristic oval 
 form, arranged in buds. They are diagnostic 
 of fermentative dyspepsia. 
 
 xm. 
 
 The Bile. 
 
 Biliary Acids (Glj'cocholic and Taurocholic) , 
 Pettenkofer's Test. — Take a few c.c. of bile, 
 add an equal amount of water and a few drops 
 of cane-sugar solution. Then pour one-half 
 the amount of cone. H 2 S0 4 down the side 
 of the test-tube, so that the latter shall under- 
 lie the bile. At the juncture of the bile and 
 the acid there is found a purple-red zone. 
 Then dip the test-tube into a glassful of water, 
 and mix the two fluids carefully by shaking or 
 swaying the tube to and fro (if the resulting 
 
42 A Laboratory Manual 
 
 heat is developed too suddenly the test is 
 spoiled), and we obtain a magnificent purple- 
 red fluid. 
 
 After the tube is fully cooled, pour one-half 
 into another tube containing alcohol, and the 
 second half into a third tube containing glacial 
 acetic acid. 
 
 («.) The acetic acid solution shows in the 
 spectroscope a line in green. 
 
 (b.) The alcoholic solution shows at first 
 only the line in green, but the solution soon 
 becomes brownish, and we find a line in blue. 
 
 The results of these tests show that we have 
 present both glycocholic and taurocholic acids, 
 as well as cholic acid, which forms the basis of 
 both. 
 
 This test can be modified by diluting the 
 bile tenfold, and taking a few drops of the 
 dilute solution, and adding a trace of sugar and 
 a few drops cone. H 2 S0 4 . Evaporate in an 
 evaporating dish upon the water-bath, and a 
 violet-red color appears upon the dish at the 
 edge of the fluid. As soon as this appears 
 cease evaporating. 
 
 Biliary Pigments (Bilirubin and Biliver- 
 din) , Gmelin's Test. — Dilute some of the bile 
 ten times, take a portion in a test-tube, and 
 pour down the side of the tube some impure 
 
Of Physiological Chemistry 43 
 
 HXO3 (nitric acid containing HN0 2 ), so 
 that the acid shall underlie the bile. There is 
 formed at the junction of the two fluids a play 
 of colors, consisting of green, blue, violet, red, 
 and reddish-yellow. The green must always 
 be present. The play of colors in this test is 
 the result of oxidation, the part nearest the 
 acid being most intensely oxidized. 
 
 Huppert's Test. — Take some dilute bile 
 solution, and add lime-water as long as a pp. 
 is formed. Filter the pp. off (which consists 
 of bilirubin lime) , wash once with water, and 
 place while still moist in a test-tube filled half 
 full with alcohol which has been acidified with 
 sulphuric acid. Heat to boiling in the water- 
 bath. The fluid takes on a brilliant green or 
 bluish-green color. 
 
 XIV. 
 
 Pancreatic Juice. 
 
 Trypsin. — Dissolve 1 gramme powdered 
 extract of pancreas in 500 c.c. water, and add 
 5 c.c. of a saturated sodium carbonate solution. 
 Place in a flask, and add the whites of two hard- 
 boiled eggs finely chopped. Add a few drops 
 
44 A Laboratory Manual 
 
 of chloroform, cork the flask tightly, and place 
 in a brood oven for a few days, shaking occa- 
 sionally. Now acidify carefully with acetic 
 acid while warming, and filter. Shake a portion 
 of the nitrate in a test-tube with bromine 
 water. It should give a violet-red color. Now 
 evaporate the filtrate to about 100 c.c. and allow 
 to stand in a cool place until the next day. 
 Separation of white crystals of tyrosin. Filter 
 these off (preserving the filtrate) and examine 
 under the microscope. Colorless, needle-shaped 
 crystals of tyrosin. Dissolve some in water in 
 test-tube, add a few drops of Millon's reagent, 
 and heat to boiling. Rose-red to deep-red color. 
 
 Scherer's Test. —Evaporate a few of the 
 tyrosin crystals to dryness on platinum foil 
 with nitric acid. A yellow residue is obtained, 
 which on adding NaOH develops a deep red- 
 dish-yellow color. 
 
 The filtrate from the above is to be evapo- 
 rated still further upon the water-bath, until 
 there is formed a scum upon the surface of the 
 fluid. This scum is leucin. Examine under 
 the microscope. Crystals of leucin, which 
 consist of globules having a pearly lustre and 
 radial lines. Easily soluble in HC1 and 
 alkalies. 
 
 Diastase. — Add 1 gramme pancreas powder 
 
Of Physiological Chemistry 45 
 
 to 50 c.o. water, digest for two hours at 40° C, 
 and filter. Take equal parts of the filtrate and 
 starch clyster (made as for saliva) and digest 
 for a short time in the brood oven. The starch 
 solution becomes transparent and shows the 
 presence of sugar by Trommer's test. 
 
PART II. 
 THE URINE. 
 
THE URINE. 
 
 I. 
 
 General Characteristics. 
 
 /- Pale. 
 Color. — Normal 1 Normal. 
 (High. 
 
 r Blood. 
 Pathological < Bile. 
 
 I Chyle. 
 Accidental — due to drugs. 
 Odor. — Accidental (due to drugs) . Uri- 
 nous. Ammoniacal. Due to acetone. 
 
 Reaction. — Acid or alkaline. The alkali 
 may be fixed or volatile. If fixed, it does not 
 disappear on heating ; if volatile, it disappears 
 on heating. 
 
 Specific Gravity. — Normal, 1015-1025. 
 Amount. — Subject to great variation. Nor- 
 mal, 1,500 c.c. 
 
 Solids by Specific Gravity. — Multiply last 
 two figures of specific gravity by 2.33, which 
 gives the number of grammes of solids in 1,000 
 c.c. 
 
50 A Laboratory Manual 
 
 n. 
 
 General Chemistry and 
 Clinical Analysis. 
 
 Urobilin. — Take a little H 2 S0 4 in a test- 
 tube and pour on it, from a height, some urine. 
 Purplish-brown color, if present. 
 
 If decreased — light color. 
 
 If normal — dark and transparent. 
 
 If increased — dark and opaque. 
 
 Indican (Stokvis' Test) . — To J- test-tube of 
 urine add an equal amount HC1 and one or 
 two drops calcium hypochlorite, and' shake. 
 Then add a few drops of chloroform and shake 
 thoroughly. If indicaniPiesent^he chloro- 
 form will be colored blue, -flf fuormal, it will 
 be about the color of litmKfcst-'^ 
 
 Albumen (Qualitative) . 
 Serum Albumen. Nitric Acid Test. — To 
 
 about £ wine-glassful of urine add with a 
 pipette about \ the amount of cone. HNO 
 so that the acid shall underlie the urine. If 
 present, there will be a distinct white zone at 
 the point of contact, which is sharply defined 
 and extends either way. If bile be present, 
 
Of Physiological Chemistry 51 
 
 the albumen zone will be tinged green. In 
 the nitric-acid test several other zones will be 
 present, viz. : 
 
 1. A nitrate of urea zone, which always 
 occurs in the nitric acid and extends down- 
 wards. 
 
 2. A zone of indican and urobilin, which is 
 exactly below the albumen zone. 
 
 3. A zone due to urates, which appears above 
 the nitric acid in the urine and extends up- 
 wards. 
 
 4. A mucin zone in the upper part of the 
 urine. 
 
 Heat Test. — To some filtered urine in a test- 
 tube add a drop of dilute acetic acid, and heat 
 the upper part of the fluid. If present, there 
 will be a cloudiness to a coagulation, which can 
 be best seen by comparing it with the clear 
 fluid below. This test, if properly performed, 
 is a very delicate one. 
 
 Mucleo- Albumen,— Add a few drops of 10% 
 acetic acid. If present, there will be a pp. 
 soluble in excess of the acid. 
 
 Mucin.— Same as for nucleo-albumen, but not 
 soluble in excess of acetic acid. 
 
 Globulin. — Dilute a small amount of urine 
 with several times its bulk of water. If pres- 
 ent, a pp. will be formed. 
 
5 2 A Laboratory Manual 
 
 Alburaose-peptone. — Remove the other albu- 
 mens by 10 % acetic acid and heating ; filter, 
 and test filtrate by : (a) Biuret reaction, (b) 
 Take 10 c.c. of above urine, add 1 c.c. HC1, and 
 then precipitate with phospho-tungstic acid. 
 Warm slightly, let pp. fall to the bottom of the 
 tube, pour off the fluid above, and wash pp. 
 with water. Dissolve pp. in NaOH and add 
 a weak solution CuS0 4 . Biuret reaction. 
 
 Albumen (Quantitative) . 
 Esbach's Test. — Fill to the mark U with 
 clear, filtered urine, and then add Esbach's re- 
 agent to the mark R. Let stand for 24 hours 
 or shake down in a centrifuge. Every mark 
 occupied by the pp. represents Jj- of lf of 
 albumen, or 1 gramme to the liter. 
 
 Sugar (Qualitative). 
 
 Tronimer's Test. — Take | test-tube of urine, 
 add an equal amount of NaOH, and then add 
 CuS0 4 until it no longer dissolves. Heat the 
 upper part, and if sugar be present there will 
 be a reduction of the copper to the red 
 suboxide. 
 
 Fehling-'s Test. — Take a few c.c. of Fehling's 
 solution, dilute with about 4 times the amount 
 of water, and heat to boiling. Then add a 
 
Of Physiological Chemistry $3 
 
 little urine and heat once more near the upper 
 part of the fluid. Reduction of copper, if sugar 
 is present, to red suboxide. 
 
 Nylander's Test. — Add to the urine about \ 
 the amount of Nylander's solution, and boil. 
 Reduction of metallic bismuth if sugar is pres- 
 ent. 
 
 Sugar (Quantitative). 
 
 Fehling's Method. — First ascertain the spe- 
 cific gravity. 
 
 If 1.030 or below, dilute 5 times, by taking 
 10 c.c. of urine and adding 40 c.c. water. 
 
 If above 1.030, dilute 10 times, by taking 5 
 c.c. urine and adding 45 c.c. water. 
 
 Place the urine in a burette. Take 10 c.c. 
 Fehling's solution, place in a small flask, and 
 add 40 c.c. water. Bring to a boiling point, 
 and add the urine a little at a time, boiling 
 between each addition. Continue this until 
 the blue color has entirely disappeared. 
 
 Calculation. — 10 c.c. Fehling's solution = 
 .050 grammes sugar ; therefore the amount of 
 mixed urine used in the titration, divided by 
 its degree of dilution, contains exactly .050 
 grammes sugar. Calculate entire amount for 
 24 hours. 
 
 Fermentation Test. — Ascertain accurately 
 the specific gravity of the urine. Then take 50 
 
54 A Laboratory Manual 
 
 c.c. of the urine and add 2 grammes yeast. 
 Place in a bottle which is closed tightly and 
 through the cork of which is passed a reversed 
 pipette. Place in a warm place and allow to 
 stand for 24 hours. Again take the specific 
 gravity after fully cooled, and the difference 
 multiplied by 230 gives the percentage of 
 sugar. 
 
 Note. — In all quantitative estimations albu- 
 men must be removed, and the urine measured 
 with a pipette and not a graduate. 
 
 Urea (Qualitative). 
 
 To a few drops of urine on a microscopic 
 slide add a drop of cone. HN0 3 , and in a 
 few minutes crystals of urea nitrate will have 
 separated out. Examine with a microscope. 
 
 Quantitative (Liebig's Method). 
 
 Take 20 c.c. urine and add to it 10 c.c. 
 baryta mixture. Then filter, take 15 c.c. of 
 the filtrate, which should be clear, place in a 
 beaker, and titrate with a standard solution of 
 mercuric nitrate, until a drop, added to a 
 saturated solution sodium carbonate, gives a 
 yellow pp. of oxide of mercury. 
 
 Calculation. — 1 c.c. mercuric nitrate = .010 
 Gives amount of urea in 10 
 
Of Physiological Chemistry 55 
 
 c.c. urine. Calculate entire amount for 24 
 hours. 
 
 Normal amount, 25 to 40 grammes in 24 
 hours. 
 
 Uric Acid (Qualitative). 
 
 Take 20 c.c. urine and add to it 2 c.c. cone. 
 HC1. Set aside for 24 hours in a cool place. 
 The uric acid which has separated out is coU 
 lected on a filter and subjected to the murexid 
 test. 
 
 Murexid Test. — Place the crystals in an 
 evaporating dish, and dissolve them in a few 
 drops of HNO3 with the application of heat. 
 Evaporate carefully to dryness, when a yellow- 
 ish-red spot will be found to remain. Cool, and 
 add a drop of ammonia, when a beautiful pur- 
 plish-red color will develop, owing to the 
 formation of ammonium purpurate (murexid) . 
 
 Quantitative (Hopkins' Method) . 
 
 Take 100 c.c. urine and add 30 grammes of 
 finely powdered ammonium chloride. The 
 mixture is then allowed to stand for two hours, 
 and is stirred from time -to time. Then filter 
 and wash the pp. two or three times with a satu- 
 rated solution (NH 4 )C1. "Wash the pp. from 
 the filter, with a little boiling water, into an 
 
56 A Laboratory Manual 
 
 evaporating dish, and add 5 c.c. HC1. Evapo- 
 rate the solution until crystals of uric acid 
 begin to separate out. These are collected on 
 a filter, the filter-paper with the adherent pp. 
 is placed in a beaker, and the crystals are 
 dissolved in as small an amount of a 30 % solu- 
 tion of sodium carbonate as possible, with the 
 aid of heat. Cool, dilute with water to 100 
 c.c, add 20 c.c. cone. H 2 S0 4 , and warm. 
 Titrate, while warm, with a ^ normal solution 
 potassium permanganate, until the rose-red 
 color no longer disappears on stirring. 
 
 Calculation. — 1 c.c. potassium permanga- 
 nate =3.61 milligrammes uric acid. Gives 
 amount of uric acid in 100 c.c. urine. Calcu- 
 late entire amount for 2 4 hours . Normal amount 
 eliminated in 24 hours, about 1 gramme. 
 
 Chlorides (Qualitative) . 
 
 To a few c.c. urine which has been acidified 
 with a few drops HN0 8 add a few drops AgN0 3 . 
 White pp. of silver chloride. 
 
 Quantitative (Mohr's Method). 
 
 Take 10 c.c. urine, acidify with a few drops 
 HN0 3 , and dilute with distilled water to 50 
 c.c. Then add very carefully sodium carbo- 
 
Of Physiological Chemistry 57 
 
 nate in substance until just neutral. Then 
 add a few drops yellow K 2 Cr0 4 , which gives 
 the solution a yellow color, and titrate with a 
 standard solution of silver nitrate until a distinct 
 orange tinge is obtained after brisk stirring. 
 
 Calculation. — 1 c.c. AgN0 3 = .010 gramme 
 chlorides. Gives amount of chlorides in 10 
 c.c. urine. Calculate entire amount for 24 
 hours. 
 
 Normal amount eliminated in 24 hours, 
 11-15 grammes. 
 
 Sulphates. 
 
 To a few c.c. of urine add a few drops of 
 solution of barium chloride and HC1. A white 
 pp. of barium sulphate. 
 
 Phosphates (Qualitative) . 
 
 Total Phosphates. — A few c.c. of urine are 
 acidified with a few drops of acetic acid and 
 then treated with a solution of ferric cbloride. 
 A yellowish- white pp. of ferric phosphate. 
 
 Earthy Phosphates. — To a few c.c. of 
 urine add ammonia in excess. A flocculent 
 pp. indicates their presence. 
 
 Alkaline Phosphates.— Precipitate the earthy 
 phosphates with NH 4 OH, filter, and to the clear 
 filtrate add magnesium sulphate solution. A 
 heavy white pp. 
 
58 A Laboratory Manual 
 
 Quantitative Estimation of Phos- 
 phoric Acid. 
 
 Take 50 c.c. urine and add 5 c.c. of sodium 
 acetate solution. Heat the mixture to boiling, 
 and titrate with a standard solution of uranyl 
 nitrate, until a drop, placed in a solution 
 of potassium ferrocyanide, gives rise to a 
 mahogany-brown pp. 
 
 Calculation. — 1 c.c. uranyl nitrate = .005 
 gramme P 2 6 . Gives amount of phosphates 
 in 50 c.c. urine. Calculate entire amount for 
 24 hours. 
 
 Normal amount eliminated in 24 hours, 
 1-2 grammes. 
 
 Bile Pigments. 
 
 Gmelin's or Huppert's Test. — Green must 
 always be present. 
 
 Blood Coloring Matters. 
 
 Determine with a spectroscope which color- 
 ing matter is present. 
 
 Guaiac Test. — To a few c.c. of urine add a 
 few drops of freshly prepared tincture guaiac 
 and a little old oil of turpentine. Shake thor- 
 oughly, and if blood is present the mixture 
 becomes blue, 
 
Of Physiological Chemistry 59 
 
 Kreatinin. 
 
 A few c.c. of urine are treated drop by drop 
 with a very dilute solution of sodium nitro- 
 prusside, and then drop by drop with a solu- 
 tion NaOH. In the presence of kreatinin a 
 ruby-red color develops, which is best seen in 
 the lower portion of the tube. Then add a few 
 drops glacial acetic acid and warm, and the 
 color changes to green. 
 
 Acetone. 
 
 1. Same test as for kreatinin, except that 
 on the addition of acetic acid a purple instead 
 of a green color develops. This takes place 
 without warming. 
 
 2. A few c.c. of urine are treated with a 
 few drops of iodine in potassium iodide solution 
 and XaOH. Warm, and a pp. of iodoform 
 occurs, which may be recognized by its odor. 
 If these reactions for acetone fail, take about 
 1 liter of urine, add a few drops H 2 S0 4 and 
 distil about \, and then redistil about 10 c.c. 
 As acetone is very volatile, if any is present 
 it will be found in the 10 c.c. 
 
 Aceto-Acetic Acid. 
 
 Add ferric chloride solution. There is a pp. 
 of ferric phosphate formed, which is allowed to 
 
60 A Laboratory Manual 
 
 settle. On the further addition of ferric chlo- 
 ride, a purple color is formed if the reaction 
 is positive. 
 
 Ehrlich's Diazo-Reaction. 
 
 To a few c.c. of urine add an equal amount 
 of sulphanilic acid and a few drops of sodium 
 nitrite. Shake thoroughly, and then allow 1 
 or 2 c.c. of ammonia to run down the side of 
 the tube so as to overlie the fluid below. If 
 the reaction is positive, a carmine-red ring 
 develops at the junction of the two fluids. 
 This reaction takes place in febrile urines, and 
 is due to a peculiar chromogen. The test was 
 formerly thought to be especially diagnostic of 
 typhoid fever. 
 
 m. 
 
 Urinary Sediment. 1 
 
 Unorganized Sediments. 
 
 Uric Acid. — Whetstone shape. Almost 
 always colored yellow or brown. Forms the 
 brick-dust sediment. Soluble in NaOH. 
 
 1 To obtain a urinary sediment, allow the urine to stand in a 
 conical glass over night, or, what is better, throw it down with the 
 centrifuge, and with a pipette transfer a small portion to a slide, 
 cover with a cover-glass; and examine with a high power. Do not 
 use a condenser. 
 
Of Physiological Chemistry 61 
 
 Urates. — Sodium and potassium are amor- 
 phous. Ammonium, dumb-bell shape, or round, 
 or sheaf-shaped. 
 
 Oxalate of Calcium. — Envelope, dumb-bell 
 or oval shape. Insoluble in acetic acid. 
 
 Phosphates. — Coffin-lid shape. Soluble in 
 acetic acid. 
 
 Cystin is very rare. Small, colorless, hex- 
 agonal crystals, soluble in ammonia. 
 
 Hippuric acid (in long needles), leucin 
 (spherules with concentric striations), and ty- 
 rosin (in fine needles) are not common. 
 
 Organized Sediments. 
 
 Blood. — Xorrnal and abnormal shapes of 
 corpuscles ; that is, round or shrivelled 
 (crenated) . 
 
 Leucocytes. — Resemble distended red blood 
 corpuscles. 
 
 Pus Corpuscles. — Are round, with nuclei 
 and granules. On the addition of acetic acid 
 the granular matter is destroyed and the nuclei 
 are brought out very clearly. 
 
 Epithelial Cells. —Are either normal, gran- 
 ular, or fatty. These three conditions mean 
 simply the age of the process. 
 
 Eenal (from convoluted tubules) — are 
 slightly polygonal, with nucleus or with gran- 
 ules, nucleus persisting. 
 
A Laboratory Manual 
 
 Renal (from straight tubules) — are less 
 polygonal and more circular. 
 
 Pelvic — battledore shape, if superficial ; 
 round, if from the deeper layers. 
 
 Ureter — spindle-shaped cells. 
 
 Bladder — from fundus polygonal ; from 
 neck round. 
 
 Vaginal — large and very faint, frequently 
 overlapping. 
 
 Urethral — spindle-shaped. 
 
 Prostate — resemble pelvic cells (deep). 
 
 Fat may be found as free fat, or dotting 
 epithelial cells, or as so-called compound 
 granular cells. 
 
 Casts. 
 
 Hyaline — faint, with both ends rounded. 
 
 Fibrinous — contains a substance of greater 
 density. Varies from hyaline only in intensity. 
 
 Blood — c,ast covered with blood corpuscles 
 and leucocytes. 
 
 Epithelial — cast covered with renal epithelial 
 cells. 
 
 Granular — cast covered with granules. 
 
 Fatty — cast covered with fat globules. 
 
 Waxy — are very distinct, intense, and homo- 
 geneous. Slightly yellow. 
 
Of Physiological Chemistry 63 
 
 Fungi and Bacteria. 
 
 Micrococcus Ureae. Produces ammoniacal 
 fermentation, converting urea into ammonia. 
 
 Bacterim Termo. Rod-like shape. 
 
 Pencillium Glaucum. Grows in chains. In- 
 dividuals are oval-shaped, with a little depres- 
 sion at each end. Occurs only in acid urines. 
 
 Yeast Fuugi. Found only in diabetic urines, 
 or in urines containing sugar. Ovals with de- 
 pressed centres. 
 
 Sarcinae. Analogous to the sarcinse found 
 in the stomach. 
 
 Tubercle Bacilli. Found in tuberculosis of 
 the genito-urinarv tract. Do not confound 
 with the smegma bacillus, which morphologi- 
 cally resembles the tubercle bacillus. 
 
 Spermatozoa. 
 
PART III. 
 TOXICOLOGY. 
 
I. 
 
 Acids. 
 
 If unmixed with organic matter, concentrate 
 if too dilute. 
 
 Sulphuric Acid. 
 
 A. First determine whether a free acid 
 or a sulphate is present, by methyl-violet or 
 dimethyl-amido-azo-benzol . 
 
 1. To a small portion of the fluid add 
 acetate of lead solution. White pp. 
 
 2. To another portion add barium chloride 
 solution. Heavy white pp. 
 
 3. To another portion add calcium chloride 
 solution. Insoluble white pp. 
 
 B. If mixed with organic matter (stomach 
 contents, etc.), concentrate to a syrup, extract 
 with successive portions of alcohol, and drive 
 off the alcohol by warming on the water bath. 
 A watery solution is left, and with it perform 
 tests 1, 2, 3. 
 
 Nitric Acid. 
 
 A. If unmixed, test for free acid, to dis- 
 tinguish from nitrates. 
 
68 A Laboratory Manual 
 
 1 . Boil a small portion with a little albumen 
 (white woolen thread or peptone) — yellow 
 color. Cool, and add NaOH — orange color. 
 (Xanthoproteic reaction.) 
 
 2. Add to a small portion in a test-tube a 
 piece of copper and some concentrated sulphuric 
 acid, and heat. Red fumes. 
 
 3. Add to a portion some sulphuric acid, and 
 pour carefully down the side of the tube some 
 ferrous sulphate solution. Brown zone at point 
 of contact. 
 
 B. If mixed with organic matter, concen- 
 trate, cool, extract rapidly with alcohol, neu- 
 tralize with potash, filter, and drive off the 
 alcohol. Use tests 2 and 3. Reacts equally 
 well with nitrates. 
 
 Hydrochloric Acid. 
 
 A. If unmixed, test for free acid. 
 
 1. Add silver nitrate. White pp. soluble 
 in ammonia, which darkens on exposure to 
 light. 
 
 2. Add lead acetate solution. Heavy, 
 white, crystalline pp. 
 
 3. Add mercurous nitrate solution. Fine 
 white pp. 
 
 B. If mixed with organic matter, distil, 
 and test distillate. 
 
Of Physiological Chemistry 69 
 
 Oxalic Acid. 
 
 Does not react to tests for free mineral acids. 
 
 A. If unmixed : 
 
 1 . Add calcium chloride or calcium sulphate 
 solution. Fine powdery pp. of calcium oxa- 
 late. Examine with the microscope. 
 
 2. Add silver nitrate solution. White pp. 
 of silver oxalate, soluble in ammonia, and with 
 some difficulty in nitric acid. Silver chloride 
 is insoluble in nitric acid. 
 
 3. Add baryta water. White pp. 
 
 B. If mixed with organic matter, acidulate 
 with HC1 ; evaporate to dryness, extract hot 
 (by boiling) with alcohol ; drive off the alcohol 
 and dissolve residue in water. Apply all the 
 tests. 
 
 Carbolic Acid. 
 
 Does not react to tests for free mineral acids. 
 A. If unmixed : 
 
 1. Add ferric chloride solution. Deep 
 amethyst-blue color. 
 
 2. Add to a small portion I its volume of 
 ammonia, a few drops calcium hypochlorite 
 solution, and warm gently, but not to boiling. 
 Blue to green color. 
 
 3. Add a few drops Millon's reagent and 
 heat to boiling. Intense dark-red color. 
 
70 A Laboratory Manual 
 
 4. Add bromine water. A gelatinous pp. of 
 tribromophenol, soluble in excess of carbolic 
 acid. 
 
 B. If mixed with organic matter, acidify 
 strongly with HC1 and distil . Apply all the tests . 
 
 Hydrocyanic Acid (Potassium Cyanide). 
 
 A. If unmixed with organic matter : 
 
 1. Add a few drops ferrous sulphate solution, 
 then a few drops of ferric chloride, and acidify 
 strongly with HC1. Blue pp. of Berlin blue. 
 
 2. Add some ammonium sulphide, evaporate 
 to dryness upon the water bath, acidify with 
 HC1, and add a couple drops of ferric chloride. 
 Blood-red color from sulphocyanide of iron. 
 
 3. Add a few drops of picric acid, and warm. 
 Blood-red color from isopurpurate of potash. 
 
 4. Add silver nitrate. White pp. of silver 
 cyanide. 
 
 B. If mixed with organic matter : 
 
 1. Place a piece of filter-paper moistened 
 with guaiac mixture in the cork of the flask, 
 and warm. Filter-paper becomes blue. 
 
 2. Acidify with tartaric acid and distil into 
 a weak potash solution. The delivery tube 
 must pass below the surface of the solution, to 
 prevent escape into the air. With the dis- 
 tillate perform the tests. 
 
Of Physiological Chemistry 71 
 
 n. 
 
 Metallic Poisons. 
 
 Arsenic (Arsenious Acid). 
 A. If unmixed with organic matter : 
 
 1. Place a small granule of white arsenic in 
 the closed end of a reduction tube, and above 
 this place a small splinter of charcoal. First 
 heat where the charcoal is, then the closed end 
 of the tube, and just above the charcoal there 
 is formed a mirror of metallic arsenic. Now 
 remove the charcoal and heat the mirror, and a 
 sublimate of white arsenic is formed above it. 
 Examine with a microscope. Octahedral crys- 
 tals. 
 
 2. Slightly acidify some of the solution with 
 HC1, and pass H 2 S through it. Yellow sul- 
 phide of arsenic is formed, which is soluble in 
 ammonia. 
 
 3. Add ammonia to some of the solution, 
 drive off the excess by warming on the water 
 bath, and divide into 2 parts. To : 
 
 (a.) Add silver nitrate, and a yellow pp. 
 of silver arsenate is formed. To : 
 
 (5.) Add copper sulphate. A pale green 
 pp. of arsenate of copper (Scheel's green). 
 
72 A Laboratory Manual 
 
 B. If mixed with organic matter : 
 Reinscli Test. — Strongly acidulate the solu- 
 tion with HC1, place in it a piece of bright 
 copper tinsel, and warm 10 to 15 minutes over 
 the water bath. The copper, which has become 
 covered with a grayish-white layer, is now to 
 be removed, washed with water, alcohol, and 
 ether, and thoroughly dried. Place in a small 
 reduction tube and heat gently. Sublimate of 
 arsenious acid formed in the cool part of the 
 tube. Examine with the microscope. Octa- 
 hedral crystals. This method, although not as 
 delicate as the Marsh test, possesses the advan- 
 tage that it can be applied in the presence of 
 organic matter. 
 
 Antimony (Tartar Emetic). 
 
 A. If unmixed: 
 
 1. Add to some of the solution ^ its vol- 
 ume of HC1 and place in the solution a piece 
 of tin. Black deposit of antimony upon the tin 
 in the cold. 
 
 2. Acidulate some of the solution with HC1 
 and pass H S through it. Orange-red pp. of 
 sulphide of antimony. 
 
 3. Add nitric acid drop by drop to some of 
 the solution. White pp. of subnitrate of 
 antimony, soluble in excess of nitric acid. 
 
Of Physiological Chemistry 73 
 
 B. If mixed with organic matter : 
 Fresenius-Babo Method. — Acidify strongly 
 with HC1 (^ its volume) , and add, while warm- 
 ing, chlorate of potash, until the solution is 
 yellow. Drive off the free chlorine, filter, 
 dilute largely with water, and pass HjS through: 
 Orange-red pp. of sulphide of antimony. As 
 arsenic may give a somewhat similar pp. and 
 organic matter still be present, filter out the 
 pp. dry, and burn with saltpeter mixture. After 
 cooling, dissolve the residue, with warming, in 
 dilute HC1 (1 : 2), and to the clear solution 
 apply the individual tests. 
 
 Mercury (Corrosive Sublimate). 
 
 A. If unmixed : 
 
 1. Pass H 2 S through a small portion, and 
 there is formed a pp. which is at first yellow 
 and finally black. 
 
 2. Add XaOH to some of the solution. 
 Yellow pp. of mercuric oxide. 
 
 3. Add (NH 4 )OH. White pp. of amido- 
 mercuric chloride, soluble in HC1. 
 
 4. Add subchloride of tin, and there is 
 formed a white pp. of subchloride of mercury 
 or a gray one of mercury. 
 
 5. Add potassium iodide. Scarlet pp. of 
 mercuric iodide, 
 
74 A Laboratory Manual 
 
 B. If mixed with organic matter : 
 Acidify with HC1, insert a piece of bright 
 copper tinsel, and warm over the water-bath. 
 Remove the copper, wash, dry, and heat in 
 small ignition tube. Sublimate of mercury 
 formed in the cool part of the tube. Place a 
 small granule of iodine above the sublimate in 
 the tube, and heat the sublimate once more. 
 Bright red zone of iodide of mercury. Ex- 
 amine with the microscope. Diamond-shaped 
 crystals. 
 
 m. 
 
 Metallic Poisons (Continued). 
 
 Copper (Copper Sulphate). 
 
 A. In pure solution : 
 
 1. Pass H 2 S through the solution. Black 
 sulphide of copper. 
 
 2. Add some potassium ferrocyanide. 
 Brownish-red pp. 
 
 3. Add ammonia to some of the solution. 
 Blue pp., soluble in excess of the ammonia, 
 giving a deep-blue fluid. 
 
 B. If mixed with organic matter : 
 
 1. Accidulate with HC1 and insert a piece 
 
Of Physiological Chemistry 75 
 
 of steel. Bright-red layer of copper upon the 
 steel. 
 
 2. Destroy organic matter by HC1 and 
 KCIO3 (Fresenius-Babo), filter, dilute filtrate, 
 and pass H 2 S through. Black pp., which can 
 be further verified by the above tests. 
 
 Lead (Lead Acetate) . 
 
 A. In pure solution : 
 
 1. Add sulphuric acid. White pp. of lead 
 sulphate. 
 
 2. Add HC1. White pp. of lead chloride, 
 insoluble in ammonia. (Distinction from 
 silver chloride.) 
 
 3. Add potassium ferrocyanide. White 
 pp. of ferrocyanide of lead. 
 
 4. Add potassium iodide. Yellow pp. of 
 lead iodide. 
 
 B. If mixed with organic matter : 
 Destroy organic matter with HC1 and 
 
 KC10 3 , filter while hot, and, upon cooling and 
 diluting, white crystalline lead chloride may be 
 deposited. This can be redissolved in hot 
 water and the tests applied. If the chloride 
 is not deposited, pass H 2 S through the solution, 
 filter off the black lead sulphide, and dissolve 
 in warm nitric acid. Verify by the tests. 
 
76 A Laboratory Manual 
 
 Phosphorus. 
 
 1. Fasten a piece of filter-paper, moistened 
 with silver nitrate solution, to the cork of the 
 flask containing the mixture, and warm slightly 
 (by placing the flask in warm water) . Filter- 
 paper becomes blackened, due to the formation 
 of silver phosphide. This must take place in 
 the dark, as silver nitrate is blackened by light 
 alone. It is also best to place a second piece 
 of filter-paper in the cork, moistened with lead 
 acetate, to exclude hydrogen sulphide, which 
 would also blacken silver nitrate. 
 
 2. Acidify a portion with sulphuric acid, 
 and distil into a weak solution of silver nitrate. 
 If examined in the dark, the vapor is phos- 
 phorescent in the cooler, and the silver is pre- 
 cipitated as a black phosphide of silver. 
 
 Nitro-Benzol. 
 
 A. If unmixed : 
 
 1. Mix some of the solution with alcohol, 
 add NaOH and some sulphur or the sulphide of 
 an alkali. Red color, if present. 
 
 2. Distil a portion, dissolve the oily drops 
 of the distillate in alcohol, and treat with zinc 
 and dilute H 2 S0 4 . From this aniline is formed. 
 Render alkaline with NaOH, shake with ether, 
 allow the ether to evaporate, and treat the resi- 
 
Of Physiological Chemistry 77 
 
 due with chloride of lime solution. Violet 
 color. 
 
 3. Distil a portion, add to distillate a few 
 drops KOH, boil, and then add carbolic acid. 
 Evaporate to a ring of dryness, and touch the 
 ring with calcium hypochlorite solution. Green 
 color, if present. 
 
 B. If mixed with organic matter, distil, 
 and apply all the tests. 
 
 For the detection and the isolation of the 
 metallic poisons when mixed with organic 
 matter, the ideal method is that of Fresenius- 
 Babo ; that is, the destruction of organic matter 
 with hydrochloric acid and potassium chlorate, 
 as o-iven above. In the metals under consid- 
 eration it can be applied to antimony, copper, 
 lead, arsensic, and mercury. As the two latter 
 are volatile, care must be taken against their 
 possible loss, by conducting the operation in 
 a flask containing a cork with a long tube, 
 instead of in an evaporating dish, as with anti- 
 mony, copper, and lead. In the case of 
 arsenic, the latter procedure is not absolutely 
 necessary, provided care be taken to add the 
 potassium chlorate before heating. 
 
78 A Laboratory Manual 
 
 IV. 
 Alkaloids. 
 
 If unmixed with organic matter : 
 
 Morphine. 
 
 1. Take 1 or 2 c.c. of the solution in a 
 test-tube, acidify with sulphuric acid, add a 
 couple drops of chloroform, a small granule of 
 iodic acid, and shake thoroughly. Chloroform 
 becomes colored red, showing the presence of 
 morphine. 
 
 2. Take a granule of the substance (or if 
 in pure solution, evaporate to dryness on the 
 water-bath), add dilute nitric and sulphuric 
 acids (a drop of each only), warm, and allow 
 to cool. Blood-red color. 
 
 3. Dissolve a granule in a couple drops of 
 nitric acid. Orange-red color. 
 
 4. Add to a granule a drop of Froehde's 
 reagent. Colored violet, green, blue-green, 
 and yellow. 
 
 Strychnine. 
 
 1. Dissolve a granule or two in concen- 
 trated sulphuric acid in a porcelain dish, and 
 then add a small granule of red chromate of 
 
Of Physiological Chemistry 79 
 
 potash. Colored blue, violet, and finally 
 cherry-red. 
 
 2. Add a drop of dilute nitric acid to a 
 granule, in an evaporating dish, and warm 
 carefully. Xow add to the warm solution a 
 small crystal of chlorate of potash. Intense 
 scarlet color, which changes to brown upon 
 the addition of 1 or 2 drops of ammonia, with 
 the formation of a brown pp. 
 
 Veratrine. 
 
 1. Warm with hydrochloric acid. Beauti- 
 ful red color. 
 
 2. Add to some of the dry alkaloid some 
 cane-sugar, and moisten with concentrated sul- 
 phuric acid. Yellow, blue, and finally a violet 
 color. 
 
 3. With Froehde'.s reagent. Yellow, then 
 cherry-red. 
 
 Atropine. 
 
 1. Dissolve a portion of the solid in fum- 
 ing nitric acid, evaporate to dryness upon the 
 water bath, and moisten, after cooling, with a 
 10% alcoholic KOH solution. Violet, then 
 cherry-red color. 
 
 2. Heat a portion of the dry alkaloid in a 
 small closed tube, or in an evaporating dish, 
 
80 A Laboratory Manual 
 
 over a free flame, until white fumes begin to 
 rise. Odor of violets. 
 
 Cocaine. 
 
 1. Add bf chromic acid solution to a por- 
 tion, and there is formed a pp. which at first 
 becomes redissolved, but after a time returns 
 as a crystalline orange-colored pp. of cocaine 
 chromate. This reaction takes place more 
 quickly upon the addition of a few drops of 
 cone. HC1. 
 
 2. Add 5 % chromic acid solution, and warm. 
 Green color, with fumes of benzoic acid. 
 
 3. Add drop by drop 1% potassium per- 
 manganate solution. Small violet crystals of 
 permanganate of cocaine. 
 
 4. Touch a small portion of a dilute solu- 
 tion to the tongue, and there is caused a numb- 
 ness lasting for some time. 
 
 If the alkaloid is mixed with organic matter, 
 in order to isolate it it is necessary to resort 
 to the method of Stas-Otto. 
 
 The Method of Stas-Otto. 
 
 1. Evaporate upon the water-bath to a 
 syrup. 
 
 2. Transfer to a flask containing a cork with 
 
Of Physiological Chemistry 81 
 
 a long tube, add twice as much of a 1. % alco- 
 holic tartaric acid solution, and warm upon the 
 water-bath from 30 minutes to an hour. 
 
 3. Cool, filter, concentrate the filtrate until 
 the alcohol is driven off, and filter once more 
 through a filter moistened with water. 
 
 4. Evaporate the filtrate once more to a 
 syrup. 
 
 5. Add alcohol again, stirring until all is 
 separated which is not soluble. 
 
 6. Filter once more, drive off the alcohol by 
 evaporation, dissolve the residue in water, 
 render alkaline by XaOH, and shake in a sepa- 
 rating funnel with ether. Strychnine, vera- 
 trine, atropine, and cocaine are found in the 
 ether, which is allowed to evaporate spontane- 
 ously in an evaporating dish, and the residue 
 verified by the above tests. 
 
 7. Free from ether by warming upon the 
 water-bath, add ammonium chloride until am- 
 moniacal, and allow to stand over night. Then 
 warm, and shake with amyl alcohol . Morphine 
 found i n the amyl alcohol . Allow to evaporate , 
 and verify by the above tests. 
 
82 A Laboratory Manual 
 
 V.' 
 
 Scheme for the Detection of 
 an Unknown Poison. 
 
 A fresh specimen is to be taken for each test. 
 
 1. Add methyl- violet or dimethyl-amido- 
 azo-benzol. 
 
 Shows free mineral acids : 
 Sulphuric. 
 Nitric. 
 Hydrochloric (volatile) . 
 
 2. Add ferric chloride. Blue color shows 
 
 carbolic acid (volatile). 
 
 3. Take a portion in 3 test-tubes and acidify 
 withHCl. Place in: 
 
 (a.) A piece of bright copper tinsel, and 
 warm. Deposit shows arsenic, mercury, or 
 antimony. 
 
 (6.) A piece of bright tin. Black deposit 
 in the cold shows antimony. 
 
 (c.) A piece of bright steel. Red deposit 
 shows copper. 
 
 4. Add potassium iodide. Yellow pp. Or 
 moisten a piece of filter-paper with the solu- 
 
Of Physiological Chemistry 83 
 
 tion and place in a stream of H 2 S. Black 
 shining deposit shows 
 Lead. 
 
 5. Take a portion in 2 test-tubes. Place 
 in : 
 
 (a.) A strip of filter-paper moistened with 
 guaiac mixture. Warm gently. Blue color 
 shows hydrocyanic acid. 
 
 (b.) A strip of filter-paper moistened with 
 silver nitrate. Warm gently. Black deposit 
 in the dark shows phosphorus. 
 
 6. Distil into water. In distillate is found : 
 
 Hydrochloric acid. 
 Carbolic acid. 
 Hydrocyanic acid. 
 Phosphorus. 
 Nitro-benzol. 
 
 7. Extract with alcohol : 
 
 Sulphuric acid. 
 Nitric acid. 
 Oxalic acid. 
 
 8. Extract with alcohol and tartaric acid 
 (Stas-Otto) : 
 
 Morphine. 
 
 Strychnine. 
 
 Atropine. 
 
 Veratrine. 
 
 Cocaine. 
 
84 A Laboratory Manual 
 
 9. Destroy organic matter with hydrochloric 
 acid and potassium chlorate (Fresenius-Babo) : 
 Arsenic. 
 Antimony. 
 Mercury. 
 Lead. 
 Copper. 
 
APPENDIX. 
 
Reagents. 
 
 Alcohol. — The alcohol must be colorless 
 and leave no residue upon evaporation. 
 
 Sulphuric Ether. 
 
 Ammonia. — Liq. ammon. caust. 
 
 Ammonium Carbonate. 
 
 Alkaline Barium Solution (Baryta mixt- 
 ure) . — Equal parts barium water and barium 
 chloride solution. 
 
 Barium Water. — 1 part caustic barium. 
 15 parts water. 
 
 Dissolved warm, cooled, and filtered. 
 
 Barium Nitrate. — 1 : 12. 
 
 Barium Chloride. — 1 : 10. 
 
 Barium Carbonate. — Precipitated out of a 
 barium chloride solution by ammonium car- 
 bonate and well washed. 
 
 Lead Acetate. — 1 : 10. 
 
 Subacetate of Lead Solution. — (Liq. plumbi 
 subacetatis U.S. P.) 
 
 Bromine Water. — Water treated with an 
 excess of bromine and cleared by allowing to 
 stand, and decanting. 
 
 Ammonium Chloride. — 1 : 10. 
 
 Calcium Chloride. — 1 : 10. 
 (87) 
 
88 A Laboratory Manual 
 
 Sodium Chloride. — Cold saturated solution. 
 Alcoholic Zinc Chloride. —Thick syrupy 
 watery solution diluted with alcohol until the 
 spec. grav. is 1.2. 
 
 Cochineal Tincture. — 5 grammes cochineal 
 with 150 c.c. alcohol and 100 c.c. water. 
 Allow to stand some days at room temperature, 
 pour off, and filter. 
 Acetic Acid. — 30 % . 
 Ferric Chloride. — 3 : 100. 
 Potassium Ferrocyanide. — 1 : 10. 
 Yellow Chromate of Potash (KjCrO^) . — 
 1: 20. 
 
 Potassium Nitrate. — Must be free from 
 chlorides. 
 
 Potassium Hydrate. — 1 part potassium hy- 
 drate. 
 2 parts water. 
 Copper Sulphate. — 1 : 10. 
 Magnesium Sulphate. — Cold saturated solu- 
 tion. 
 
 Magnesium Mixture. — 1 part crystallized 
 
 MgS0 4 . 
 2 parts ammonium 
 
 chloride. 
 4 parts ammonia. 
 8 parts water. 
 Millon's Reagent. — 1 part mercury with 2 
 
Of Physiological Chemistry 89 
 
 parts nitric acid of spec. grav. 1.4. Gently 
 warmed until the mercury is fully dissolved. 
 One part of this solution is treated with 2 parts 
 water. 
 
 Ammonium Molybdate. — 50 grammes of 
 ammonium molybdate are dissolved in 200 
 grammes of ammonia. The solution is poured 
 into 750 c.c. nitric acid of spec. grav. 1.2; 
 allowed to stand some days in a warm place, 
 and decanted. 
 
 Sodium Hydrate. — Spec. grav. 1.17. 
 Sodium Carbonate. — Saturated solution. 
 Sodium Phosphate (NajHPC^). — 1 : 10. 
 Nessler's Reagent. 50 grammes potassium 
 iodide dissolved in an equal amount of water, 
 and as much mercuric chloride added while 
 warming until the mercury is no longer dis- 
 solved. Then add a solution of 150 grammes 
 KOH in 300 c.c. water, fill to a liter, add 5 c.c. 
 more mercuric chloride solution, and let stand. 
 Nylander's Solution. — 4 grammes tartarate 
 
 of sodium and po- 
 tassium. 
 2 grammes bismuth 
 
 subnitrate. 
 10 grammes NaOH 
 in 90 c.c. water. 
 To be kept in a dark bottle. 
 
go A Laboratory Manual 
 
 Ammonium Oxalate. — 1 : 25. 
 
 Mercuric Chloride. — 1 : 20. 
 
 Phospho-Tungrstic Acid. — 1 : 20, acidified 
 with HC1. 
 
 Potassium Sulphocyanide. — 1 : 20. 
 Rosolic Acid. — 1 : 100 alcohol. 
 Hydrochloric Acid. — 1.124 spec. grav. 
 Nitric Acid. — Must be colorless ; free from 
 nitrous acid and HC1. Spec. grav. 1.2. 
 
 Sulphuric Acid. — 200 grammes cone. H 2 S0 4 
 to 1 liter. 
 
 Silver Nitrate. — 1 : 30. 
 
 Tartaric Acid. — Crystals or powder. 
 
 Lactic Acid Solution. — 0.8 gramme : 100 
 c.c. water. 
 
 Digestive HC1 Solution. — 10 c.c. 25% HC1 
 to 1 liter water. 
 
 Sol. Methyl- Violet. — 0.5 : 1,000. 
 
 Sol. Tropaeolin OO. — 0.25:1,000. 
 
 Gunzburg's Reagent. — 1 gramme vanillin. 
 
 2 grammes phloro- 
 
 glucin. 
 100 c.c. alcohol. 
 
 Artificial Gastric Juice. — 0.5 gramme pep- 
 sin dissolved in 500 c.c. digestive HC1 solu- 
 tion. 
 
Of Physiological Chemistry 91 
 
 Fronde's Reagent. — 0.1 gramme sodium 
 molybdate dissolved in 10 c.c. cone. H 2 S0 4 . 
 Pacini's Fluid. — Mercuric chloride, 2 
 grammes. 
 Sodium chloride, 4 grammes. 
 Glycerine, 26 grammes. 
 Distilled water, 226 grammes. 
 Guaiac Mixture. — 0.5 gramme gum guaiac 
 dissolved in 10 c.c. alcohol, to which are added 
 a few drops weak copper sulphate solution 
 (1 : 2,000) . The mixture must be kept tightly 
 corked. 
 
 Saltpeter Mixture. — 3 parts KN0 8 . 
 1 part NajCCV 
 
 Dimethyl-amido-azo-benzol. — 0.5 gramme 
 to 100 c.c. alcohol. 
 
 Sub-chloride of Tin. — Metallic tin dissolved 
 in excess in HC1. 
 
 Tannic Acid. — 1 : 100. 
 Indigo Carmine. — Solution. 
 Alpha-Naphthol. -15% solution in alcohol. 
 Iodine in Potassium Iodide (IKI sol.). — 
 
 Iodine, 1 gramme. 
 
 Potassium iodide, 2 grammes. 
 
 Water, 300 c.c. 
 Picric Acid. — Saturated solution. 
 
92 A Laboratory Manual 
 
 Sodium Acetate Solution. — 100 grammes 
 acetate of sodium dissolved in distilled water, 
 and 100 c.c. of a 30% solution of acetic acid, 
 the whole being diluted to a liter. 
 
 Sulphanilic Acid. — Sulphanilic acid, 2 
 grammes. 
 
 HC1, 50 c.c. 
 
 Distilled water, 1,000 c.c. 
 Diazo solution No. 1. 
 
 Sodium Hitrite. — 0.5% solution. 
 Diazo solution No. 2. 
 
 Standard Solutions. 
 
 *L Solution Tartaric Acid (for alkalinity 
 of blood) . — Contains 3 grammes tar- 
 taric acid to 1 liter distilled water. 
 1 c.c. = .0016 gramme NaOH. 
 
 N 
 Standardized with a — solution NaOH. 
 10 
 
 Tf Solution NaOH (for acidity of gastric 
 
 10 juice) . — Contains 4 grammes NaOH 
 
 to 1 liter distilled water. 
 
 1 c.c. = .00365 gramme HC1. 
 
 N 
 Standardized with — solution H 2 S0 4 . 
 
Of Physiological Chemistry 93 
 
 Esbach's Reagent (for quantitative albu- 
 men). — 10 gramme^ picric acid. 
 20 grammes citric acid. 
 Dissolved in 1,000 c.c. distilled water. 
 
 Pehling's Solution (for quantitative sugar). 
 — 34.64 grammes of pure ci'ystallized copper 
 sulphate, dissolved in about 200 c.c. distilled 
 water. 
 
 173 grammes potassium and sodium tar- 
 tarate. 
 
 125 grammes NaOH. 
 
 Both dissolved in 500 c.c. distilled water. 
 
 These two solutions are mixed together, and 
 diluted with distilled water to 1 liter. 
 
 10 c.c. = .050 gramme glucose. 
 
 Standardized with a solution of glucose, 5 
 grammes to 1 liter distilled water. 
 
 Standard Solution Mercuric Nitrate (for 
 quantitative urea). — 71.48 grammes pure 
 metallic mercury, dissolved with warming in 
 pure cone. HN0 3 , and diluted to 1 liter with 
 distilled water. 
 
 1 c.c. = .010 gramme urea. 
 
 Standardized with a solution of urea, 2 
 grammes to 100 c.c. distilled water. 
 
94 A Laboratory Manual 
 
 ^ Solution Potassium Permanganate (for 
 quantitative uric acid) . — 1.578 grammes 
 pure crystals potassium permanganate to 1 
 liter distilled water. 
 
 1 c.c. = 3.61 milligrammes uric acid. 
 
 N 
 Standardized with — solution oxalic acid. 
 20 
 
 Standard Solution Silver Nitrate (for quan- 
 titative chlorides). — 29.06 grammes pure sil- 
 ver nitrate to 1 liter distilled water. 
 
 1 c.c. = .010 gramme NaCl. 
 
 Standardized with a solution NaCl, 10 
 grammes to 1 liter distilled water. 
 
 Standard Solution Uranyl Nitrate (for quan- 
 titative phosphoric acid) . — 33 grammes ura- 
 nium oxide dissolved in pure HN0 3 and diluted 
 to 1 liter. 
 
 1 c.c. = .005 gramme P 2 6 . 
 
 Standardized with 10.085 grammes NajHPO^ 
 to 1 liter distilled water. 
 
Of Physiological Chemistry 95 
 
 Analysis of Blood. 
 
 Date. Name. 
 
 Qualitative Tests. 
 Oxy haemoglobin. Methsemoglobin. 
 
 Carbon-monoxide-hse- Hsematin. 
 
 moglobin. Hremin crystals. 
 
 Sulpho-hsemoglobin . 
 
 Quantitative Tests. 
 
 Specific gravity. Alkalinity. 
 
 Haemoglobin. (FleischFs Htemometer.) 
 Number of red corpuscles. (By Thoma-Zeiss 
 
 method and Hasmatokrit. ) 
 Number of white corpuscles. 
 Relation of white to red. 
 
 Diagnosis. 
 
 Remarks. 
 
g6 A Laboratory Manual 
 
 Analysis of Gastric Contents. 
 
 Date. Name. 
 
 Period after test meal. 
 
 Qualitative Tests. 
 Free HC1. Pepsin. Acetic acid. 
 
 Lactic acid. Amylaceous matter. Butyric acid. 
 Rennet. Albumose-peptone. Formic acid. 
 
 Quantitative Tests. 
 
 Total acidity. 
 
 Total HC1 (free and combined) . 
 
 Microscopic Analysis. 
 Sarcinse. 
 Yeast fungi, 
 i i Diagnosis. 
 Remarks. 
 
Of Physiological Chemistry 97 
 
 Analysis of Urine. 
 
 Date. Name. 
 
 Qualitative Tests. 
 Amount in 24 hours. Specific gravity. 
 Color. Solids by specific grav- 
 
 Odor. ity. 
 
 Reaction. Sediment. 
 
 Urobilin. Bile pigment. 
 
 Indican. Blood coloring matters. 
 
 Mucin. Kreatinin. 
 Nucleo-al- Acetone. 
 
 bumen. Aceto-acetic acid. 
 Serum-al- Chlorides. 
 
 bumen. Diazo-reaction. 
 Globulin. phos hates < Alkaline. 
 Albumose- C Earthy, 
 
 peptone. gulphates 
 
 Sugar. 
 
 Quantitative Tests. 
 
 Urea. Chlorides. Albumen. - 
 
 Uric acid. Phosphoric acid. Sugar. 
 
 Albu- 
 men 
 
 
 Sediment. 
 
 
 Crystals. 
 
 Blood. 
 
 Fat. 
 
 Casts. 
 
 Spermatozoa. 
 
 Fungi. 
 
 Epithelium. 
 
 Pus. 
 
 Bacteria 
 
 Remarks. 
 
 
 
 Diagnosis.