§ 35. Spectroscopic Appearances of Blood.—If defibrinated blood[49] be diluted with water until it contains about ·01 per cent. of oxyhæmoglobin, and be examined by a spectroscope, the layer of liquid being 1 centimetre thick, a single absorption band between the wave lengths 583 and 575 is observed, and, under favourable circumstances, there is also to be seen a very weak band from 550 to 532. With solutions so dilute as this, there is no absorption at either the violet or the red end of the spectrum. A solution containing ·09 per cent. of oxyhæmoglobin shows very little absorption in the red end, but the violet end is dark up to about the wave length 428. Two absorption bands may now be distinctly seen. A solution containing ·37 per cent. of oxyhæmoglobin shows absorption of the red end to about W.L. 720; the violet is entirely, the blue partly, absorbed to about 453. The bands are considerably broader, but the centre of the bands occupies the same relative position. A solution containing as much as ·8 per cent. of oxyhæmoglobin is very dark; the two bands have amalgamated, the red end of the spectrum is absorbed nearly up to Fraunhofer’s line a; the green is just visible between W.L. 498 and 518. Venous blood, or arterial blood, which has been treated with reducing agents, such, for example, as an alkaline sulphide, gives the spectrum of reduced hæmoglobin. If the solution is equivalent to about ·2 per cent., a single broad band, with the edges very little defined, is seen to occupy the space between W.L. 595 and 538, the band being darkest about 550; both ends of the spectrum are more absorbed than by a solution of oxyhæmoglobin of the same strength. In the blood of persons or animals poisoned with hydric sulphide—to the spectrum of reduced hæmoglobin, there is added a weak absorption band in the red, with its centre nearly corresponding with the Fraunhofer line C. Blood which has been exposed to carbon oxide has a distinct spectrum, due, it would seem, to a special combination of this gas with hæmoglobin; in other words, instead of oxygen, the oxygen of oxyhæmoglobin has been displaced by carbon oxide, and crystals of carbon oxide-hæmoglobin, isomorphous with those of oxyhæmoglobin, may be obtained by suitable treatment. The spectrum of carbon oxide-hæmoglobin, however, differs so little from that of normal blood, that it is only comparison with the ordinary spectrum, or careful measurements, which will enable any person, not very familiar with the different spectra of blood, to detect it; with careful and painstaking observation the two spectra are seen to be distinct. The difference between the carbon oxide and the normal spectrum essentially consists in a slight moving of the bands nearer to E. According to the measurements of Gamgee, the band α of CO-hæmoglobin has its centre approximately at W.L. 572, and the band β has for its centre W.L. from 534 to 538, according to concentration. If a small quantity of an ammoniacal solution of ferrous tartrate or citrate be added to blood containing carbon oxide, the bands do not wholly fade, but persist more or less distinctly; whereas, if the same solution is added to bright red normal blood, the two bands vanish instantly and coalesce to form the spectrum of reduced hæmoglobin. When either a solution of hæmoglobin or blood is exposed to the air for some time, it loses its bright red colour, becomes brownish-red, and presents an acid reaction. On examining the spectrum, the two bands have become faint, or quite extinct; but there is a new band, the centre of which (according to Gamgee) occupies W.L. 632, but (according to Preyer) 634. In solutions of a certain strength, four bands may be seen, but in a strong solution only one. This change in the spectrum is due to the passing of the hæmoglobin into methæmoglobin, which may be considered as an intermediate stage of decomposition, prior to the breaking up of the hæmoglobin into hæmatin and proteids.
[49] In this brief notice of the spectroscopic appearances of the blood, the measurements in wave lengths are, for the most part, after Gamgee.—Text-Book of Physiological Chemistry, London, 1880.
A spectrum very similar to that of methæmoglobin is obtained by treating ancient blood-stains with acetic acid—viz., the spectrum of acid hæmatin, but the band is nearer to its centre, according to Gamgee, corresponding to W.L. 640 (according to Preyer, 656·6). The portion of the band is a little different in alkaline solution, the centre being about 592. Hæmatin is one of the bodies into which hæmoglobin splits up by the addition of such agents as strong acetic acid, or by the decomposing influence of exposure; the view most generally accepted being that the colouring-matter of the blood is hæmatin in combination with one or more albuminoid bodies. The hæmatin obtained by treating blood with acetic acid may be dissolved out by ether, and the ethereal solution then exhibits a remarkable distinctive spectrum. Hence, in the spectroscopic examination of blood, or solutions of blood, for medico-legal purposes, if the blood is fresh, the spectrum likely to be seen is either that of oxyhæmoglobin or hæmoglobin; but, if the blood-stain is not recent, then the spectrum of either hæmatin or methæmoglobin.
The colouring-matter of cochineal, to which alum, potassic carbonate, and tartrate have been added, gives a spectrum very similar to that of blood (see “Foods,” p. 82); but this is only the case when the solution is fresh. The colour is at once discharged by chlorine, while the colour of blood, although changed in hue, remains. The colouring-matter of certain red feathers, purpurin-sulphuric acid, and a few other reds, have some similarity to either the hæmatin or the hæmoglobin spectrum, but the bands do not strictly coincide; besides, no one would trust to a single test, and none of the colouring-matters other than blood yield hæmatin.
The blood in CO poisoning has also other characteristics. It is of a peculiar florid vermilion colour, a colour that is very persistent, lasting for days and even weeks.
Normal blood mixed with 30 per cent. potash solution forms greenish streaky clots, while blood charged with CO forms red streaky clots.
Normal blood diluted to 50 times its volume of water, and then treated successively with yellow ammonium sulphide in the proportion of 2 to 25 c.c. of blood, followed by three drops of acetic acid, gives a grey colour, while CO blood remains bright red. CO blood shaken with 4 times its volume of lead acetate remains red, but normal blood becomes brown.[50]