But while Johnson claims that neither coloring matters of normal urine nor uric acid reduce the picric acid, he admits that he has tested with picric acid and potash a large number of specimens of normal urine with the almost uniform result of a depth of color indicating the proportion of .6 of a grain of sugar to the fluidounce, the indication varying between the limits of .5 to .7 grain. The ammonio-cupric method used at the same time gave results of from .7 to .9 grain to the fluidounce, or an excess of .1 to .3 grain. Now, if my own views, the grounds for which are announced elsewhere,43 are correct, strictly normal urine contains no sugar, and any reducing action upon oxide of copper is due to uric acid, either picric acid is reduced to a degree by uric acid or by some other constituent of normal urine. This, in the light of Oliver's44 recent investigations, may be kreatinin. For he has shown that kreatinin strikes in a few seconds a red color with the cold alkaline picric solution, which is quickened by heat. From this it would seem that the exact value of the picric-acid test has as yet to be determined.
43 Tyson, Practical Examination of Urine, 4th ed., Philadelphia, 1884.
44 On Bedside Urine-Testing, including Qualitative Albumen and Sugar, by Geo. Oliver, M.D., London, Member of the Royal College of Physicians of Lond., etc., 2d ed., London, 1884.
The Indigo-Carmine Test.—The fact that indigotine, the coloring matter of commercial indigo, is converted into indigo when heated with an alkali in the presence of glucose and certain carbohydrates, has recently been applied by George Oliver of London in the construction of a test-paper. Carmine of indigo is the sulph-indigotate of sodium, an intensely blue salt, soluble in 120 parts of water. Sulph-indigotic acid is made by heating indigo with sulphuric acid, and when combined with a base, sodium, produces indigo-carmine. When sodium carbonate is mixed with a solution of indigo-carmine, the latter is precipitated in a minute state of division, but is redissolved on heating, when there results a greenish-blue solution. A freshly-made mixture of the indigo solution and sodium carbonate furnishes a fluid not unlike Fehling's solution, which gives the reaction to be described with glucose. Unfortunately, such a mixture will not keep, and the reagent would be useless but for the happy idea of Oliver of making the test-paper. In doing this bibulous paper is immersed in a solution of indigo-carmine with carbonate of sodium.45 The paper is then cut into strips an inch long and one-quarter of an inch wide.
45 No more precise directions than this are given by Oliver, either in his papers in the Lancet for 1883 or in his little book just published, On Bedside Urine-Testing. The sugar test-papers, as well as the entire series of albumen test-papers, suggested by Oliver, are now made by Parke, Davis & Co. of New York, and by Wilson & Son, Harrogate, London.
Mode of Testing.—One of the test-papers and a sodium carbonate paper46 are dropped into a half-inch test-tube, and water added until the upper end is just covered; a column of fluid one inch in height and half an inch in diameter will thus be produced, so that the solution of carmine obtained on boiling will always acquire the same concentration. Heat is now applied, the tube being gently shaken, and boiling kept up for a second or two. A beautiful blue solution will result. The test-paper may now be removed or allowed to remain.
46 Test-papers of the same size, charged with a saturated solution of sodium carbonate.
Not more than one drop of the suspected urine is let fall into the tube from a pipette held in an upright position. Drops of equal size are thus secured. The contents of the tube are again freely boiled for a few seconds, after which the tube should be raised an inch or more from the flame and held without shaking, while the solution is kept quite hot, but not boiling, for exactly one minute. If glucose be present in abnormal amount, the soft rich blue will be seen first of all to darken into violet; then, according to the quantity of sugar, there will appear in succession, purple, red, reddish-yellow, and finally straw-yellow. When the last-named color has been developed the slightest shaking of the tube will cause red streaks to fall from the surface and mingle with the pale yellowness of the solution, while further agitation will cause the return of purple and violet and the restoration of the original blue.
The time required for the commencement of the reaction after the boiling of the test liquid is in inverse proportion to the amount of glucose present. When the latter is large, over 20 grains to the ounce, it will be but a few seconds; but when small, 2 or 3 grains, from thirty to sixty seconds may elapse. If the urine do not contain more than the normal amount of sugar47—i.e. under half a grain to the ounce—the color of the solution at the end of the heating for one minute will be unchanged. The test is available by artificial light as well as by daylight.
47 It will be noted from this that Oliver accepts the view that there is a small amount of sugar in normal urine.