QUANTITATIVE ESTIMATION OF THE BLOOD PIGMENTS. 291 



solution we now generally use a carbon-monoxide haemoglobin solution 

 as a standard liquid because it may be kept for a long time. The blood 

 solution in this case is saturated with carbon monoxide. 1 



The quantitative estimation of the blood-coloring matters by means 

 of the spectroscope may be done in different ways, but at the present time 

 the spedrophoto metric method is chiefly used, and this seems to be the 

 most reliable. This method is based on the fact that the extinction 

 coefficient of a colored liquid for a certain region of the spectrum is directly 

 proportional to the concentration, so that C:E=C\:Ei, when C and 

 Ci represent the different concentrations and E and EI the corresponding 



C C 

 coefficients of extinction. From the equation -=W-, ft follows that for 



Jcj Hi i 



one and the same pigment this relation, which is called the absorption 

 ratio, must be constant. If the absorption ratio is represented by A, the 

 determined extinction coefficient by E, and the concentration (the amount 

 of coloring-matter in grams in 1 cc.) by C, then C=AE. 



Different forms of apparatus have been constructed (VIERORDT and 

 HuFNER 2 ) for the determination of the extinction coefficient, which is 

 equal to the negative logarithm of those rays of light which remain 

 after the passage of the light through a layer 1 cm. thick of an absorbing 

 liquid. In regard to this apparatus the reader is referred to other text- 

 books. 



For purposes of control the extinction coefficients- are determined in two dif- 

 ferent regions of the spectrum. HUFNER has selected (a) the region between the 

 two absorption-bands of oxyhaemoglobin, especially between the wave-lengths 

 554 fifi and 565 //*, and (ft) the region of the second band, especially the inter- 

 val between the wave-lengths 531.5 ///* and 542.5 ,/<. The constants or the 

 absorption ratio for these two regions of the spectrum are designated by HUFNER 

 by A and A'. Before the determination the blood must be diluted with water, 

 and if the proportion of dilution of the blood be represented by V, then the con- 

 centration or the amount of coloring-matter in 100 parts of the undiluted blood is 



C = WQ.V.A.E and 

 = 100. V.A'.E'. 



The absorption ratio or the constants in the two above-mentioned regions 

 of the spectrum have been determined for oxyhaemoglobin, haemoglobin, carbon 

 monoxide haemoglobin, and methaemoglobin, as follows: 



Oxyhsemoglobin A =0.002070 and A' =0.001312 



Haemoglobin A r =0.001354 and A' r =0.001778 



Carbon-monoxide haemoglobin . .A c =0.001383 and A' c =0.001263 

 Methfemoglobin 'A m = 0.002077 and ,!', = 0.001 754 



The quantity of each coloring matter may be determined in a mixture of 

 two blood-coloring matters by this method; this is of special importance in 

 the determination of the quantity of oxy haemoglobin and haemoglobin present 

 in blood at the same time. 



In order to facilitate these determinations, HUFNER 3 has worked out tables 

 which give the relation between the two pigments existing in a solution contain- 



1 See Haldane, Journ. of Physiol., 26. 



2 See Vierordt, Die Anwendung des Spektralapparates zu Photometrie, etc. (Tubin- 

 gen, 1873), and Hiifner, Arch. f. (Anat. u.) Physiol., 1894, and Zeitschr. f. physiol. 

 Chem., 3; v. Noorden, ibid., 4; Otto, Pfliiger's Arch., 31 and 36. 



3 Arch. f. (Anat. u.) Physiol., 1900. 



