STECTROPHOTOMETRIC DATA. 235 
of light (as determined by the extinction-coefficient) in a definite spectral 
region, exerted by a mixture of two or more colouring matters, is the sum of 
the extinction-coefficients of each of its coloured constituents ; and that in 
the case of a solution containing t\oo colouring matters, if we are acquainted 
with the optical constants of each in two and the same spectral regions, we are 
ahle by the spectrophotometer to determine the relative and absolute amount 
of each constituent. In a similar manner we should, according to theory, 
be able to determine the amounts of three or of x colouring matters coexisting 
in a solution, if we were acquainted with the value of A in three and the same, 
or in x and the same spectral regions. The immense importance of a method 
which permits of the accurate determination of oxy- and reduced haemoglobin 
in blood, and which furnishes us with essential data for calculating the 
amount of oxygen present in combination with haemoglobin, makes it 
necessary that we should explain the nature of the very simple calculations 
which enable us, from the determination of the extinction-coefficients in two 
spectral regions, to effect a determination which, so far as I know, cannot 
be carried out with any pretence to scientific accuracy, or even with any claim 
to be presumably correct, by any other process whatsoever. 
We shall assume that, by following methods which we shall not attempt 
to describe, but for which the reader is referred to Hiifner's original papers, 
blood has been diluted with 0*1 per cent, of aqueous solution of NaOH, 
under condition* which preclude the possibility of contact with oxygen, and 
that in the diluted blood solution the extinction-coefficients have been deter- 
mined in the first and in the second regions selected by Hiifner. These 
extinction-coefficients of a mixture of two colouring matters, we shall represent 
by E and E.' 
Let A r be the absorption relation of (reduced) haemoglobin in the first 
region (A 554 - A 556). 
A' r that of the same body in the second spectral region (A 531 '5 - 
A 542-5). 
A the absorption relation of oxyhemoglobin in the first spectral 
region. 
A' that of the same body in the second spectral region. 
Then the percentage of (reduced) haemoglobin, which we may designate x, will 
be found by the equation — 
x _ A r A' r (E'A' -EA ) 
A' A r -A A' r 
and the percentage of oxyhemoglobin by the following equation — - 
A A' (EA r -E'A' r ) 
V- 
A A r — A A r 
Having thus determined by spectrophotometry the amount of oxyhemoglobin 
by weight existing in a known volume, say 100 c.c. of blood, we can ascertain 
the volume of the respiratory oxygen measured at 0° C. and 760 mm. pressure 
(which could, but probably with less accuracy, be likewise determined with the 
aid of the mercurial pump and subsequent analyses of the gases boiled out of the 
blood) by multiplying each gramme of oxyhemoglobin found by 1-338 (or T34). 
In this manner Hiifner, having determined the relative and absolute amounts of 
haemoglobin and oxyhaemoglobin in the blood, drawn simultaneously from the 
main artery and vein of a limb, ascertained the amount of oxygen in each. 
There is a strong presumption that determinations of oxygen made in this manner 
are nearer the truth than those which the more complex and laborious methods 
by means of the mercurial pump and gas analysis are capable of giving. In the 
process of raising the blood to a temperature of at least 40° C. in the exhausted 
