586 PRACTICAL ORGANIC AND BIO-CHEMISTRY 



Estimation of a Mixture of Carbohydrates. 



The analysis of a mixture of carbohydrates, mono-, di- and polysaccharides, 

 such as occur in natural materials, especially in extracts of plants, is based 

 upon the determination of the rotation and reducing value of the solutions 

 before and after hydrolysis. All di- and polysaccharides are not hydrolysed 

 equally readily, e.g. cane sugar is readily hydrolysed by organic acids, such as 

 citric acid, whereas maltose is stable. Mineral acids hydrolyse all di- and poly- 

 saccharides but not equally readily. The analysis of mixtures is based upon 

 these differences. The subject of the analysis of mixtures of carbohydrates, 

 especially those present in plant extracts, has been carefully studied by Davis 

 and Daish, 1 who emphasise the following particulars : 



(1) Glucose and fructose are destroyed by prolonged heating with 2*5- 

 5 per cent, hydrochloric acid at 70. 



(2) Maltose is not hydrolysed by boiling with 10 per cent, citric acid. 



(3) Cane sugar is completely hydrolysed by boiling with 2 per cent, citric 

 acid for 10 minutes, but in presence of acetates, etc., at least 4 per cent, citric 

 acid is required. 



They recommend the following procedure for the analysis of the carbo- 

 hydrates in a plant extract : 



(1) The material is dropped into a large volume of boiling 95 per cent, 

 alcohol to which i per cent, of its volume of ammonia of sp. gr. -880 is added, 

 so as to destroy the enzymes and prevent changes in the proportions of the 

 carbohydrates present in the material. 



(2) The liquid is separated and the residue is extracted in a large Soxhlet 

 extractor with boiling alcohol : the soluble carbohydrates are generally removed 

 as soon as the extract becomes colourless. The liquid is then pressed out 

 from the residue by a hydraulic press. The cake of solid so obtained is dried in 

 a steam oven for 18 hours, ground up and used for the analysis of starch (below). 



(3) The alcoholic liquids are concentrated in vaciio and made up to a 

 definite volume, say 500 c.c. 



(4) Total solid is determined in 2 portions of 20 c.c. each by evaporating 

 and drying in vacuo at 100 for 18 hours. 



(5) 440 c.c. are treated with basic lead acetate; the precipitate is filtered 

 off on a Buchner funnel and washed. The volume is made up to, say, 2 litres 

 = solution A. 



(6) 300 c.c. of A are treated with solid sodium carbonate to remove lead 

 and made up to 500 c.c. = solution B. 



(7) 25 c.c. of B are taken for reduction and polarisation in a 2 decimeter 

 tube. 



The result gives the amount of glucose, fructose, maltose and pentose 

 together. 



(8) 50 c.c. of solution B are hydrolysed. 



(a) By invertase. The quantity is acidified to methyl orange with a few 

 drops of concentrated sulphuric acid, 1-2 c.c. of autolysed yeast (invertase) are 

 added together with 2 or 3 drops of toluene as preservative and the mixture 

 kept at 38-40 for 24 hours. 5-10 c.c. of alumina cream are added and the 

 solution filtered. The filtrate and washings are made up to 100 c.c. The 

 reduction and rotation of 50 c.c. are taken. 



(b] By 10 per cent, citric acid. The quantity is acidified as above and 

 solid crystalline citric acid added to make 10 per cent. The solution is 

 boiled for 10 minutes, cooled, neutralised to phenolphthalein with sodium 

 hydroxide, made up to TOO c.c. and its rotation and reduction are taken. 



The results from (a) and (<) should agree closely. 



The amount of cane sugar is calculated from the increase in reduction or 

 change of rotation. 



(9) The lead is removed from 300 c.c. of solution A by hydrogen sulphide. 

 The precipitate is washed and the filtrate and washings made up to about 450 

 c.c. Excess of hydrogen sulphide is removed by drawing air through the 

 solution for about i '5 hours, a trace of ferric hydroxide being added to remove 

 the last traces, and the solution is made up to 500 c.c. 



1 J. Agric. Sci., 1913, 6. 



