BLOOD ANALYSIS 299 



i. Fat. Nephelometric Method of Bloor, 1 Principle. The protein is precipi- 

 tated with alcohol and ether and the fatty acid in the extract determined nephelo- 

 metrically after saponification. 



Procedure. Extraction. About 2 c.c. of blood are drawn from the vein with a 

 graduated syringe and run at once with stirring into a weighed graduated flask 

 containing about 40 volumes of a mixture of 3 parts alcohol and i part ether. 

 After again weighing to find the weight of blood added, the solution is raised to 

 boiling in a water-bath, cooled under the tap, made to volume with alcohol-ether 

 mixture, mixed and filtered. The filtrate is water clear and almost colorless. 



Determination. From 5-20 c.c. of the extract (containing about 2 mg. of fat) 

 are measured with a pipette into a small beaker and saponified by evaporating 

 nearly but not quite to dryness with 2 c.c. of N/i sodium ethylate. The residue is 

 heated just to boiling after the addition of 5 c.c. of alcohol-ether, and 50 c.c. of 

 distilled water are added. 



A similar solution of the standard is prepared by adding 5 c.c. of the standard 

 fatty acid solution 2 from a pipette with stirring to 50 c.c. of distilled water. To 

 the standard and to the test solutions are added simultaneously from pipettes and 

 with stirring 10 c.c. portions of dilute (1:3) hydrochloric acid and the solutions 

 allowed to stand for five minutes, after which they are transferred to the comparison 

 tubes of the nephelometer (see Fig. 94, p. 294). Several readings should be taken 

 and averaged. The standard tube should always be on the same side. See dis- 

 cussion of nephelometer (page 294) for details as to reading. The results repre- 

 sent the amount of total fat (fatty acids and cholesterol) in the blood, expressed 

 as oleic acid. The fat of the corpuscles is not completely extracted, and it should 

 be borne in mind that other lipoids as cholesterol are included in the results. 

 Cholesterol may be determined separately and subtracted from the result for total 

 fat. It may also be determined in a part of the blood extract as prepared above 

 by a modified Autenrieth-Funk procedure. 3 Methods have also been devised for 

 the determination of the phosphatides of blood. 4 



Other Methods of Blood Analysis 



Methods for determining the alkali reserve of the blood 

 will be found in the following chapter. Important methods 

 have been developed also for magnesium, 5 sodium, 6 potassium, 7 



1 Bloor: Jour. Biol. Chem., 17, 377, 1914; 23, 317, 1915. 



2 The standard solution used is an alqohol-ether solution of pure oleic acid of which 

 5 c.c. contain about 2 mg. of the acid. The alcohol and ether used for the standard are 

 freshly redistilled absolute alcohol and pure dry ether. 



3 Bloor: Jour. Biol. Chem., 23, 317, 1915. 



4 Green wald: Jour. Biol. Chem., 21, 29, 1915. 



Bloor: Jour. Biol. Chem., 22, 133, 1915, 23, 317, 1915. 

 Kober and Egerer: /. Am. Chem. Soc., 37, 2373, 1915. 

 Taylor and Miller: Jour. Biol. Chem., 18, 215, 1914. 

 For other nephelometric methods see Chapters XVIII and XXVII. 

 6 Denis, W.: Jour. Biol. Chem., 41, 363, 1920; Marriot, W. McK., and Rowland, J. 

 Jour. Biol. Chem., 32, 233, 1917. 



6 Kramer, B.: Jour. Biol. Chem., 41, < 2'63, 1920; Doisey, E. A., and Bell, R. D.: Jour. 

 Biol. Chem., 45, 313, 1921; Kramer and Tisdall: Jour. Biol. Chem., 46, 467, 1921. 



7 Kramer, B.: Jour. Biol. Chem., 41, 263, 1920; Kramer, B. and Tisdall, F.F.: Jour. 

 Biol. Chem., 46, 339, 1921; Klausen, S. W.; Jour. Biol. Chem:, 36, 479, 1918. 



