METHODS USED IN THE STUDY OF STARCHES. 299 



experimenters in studies of the structural peculiarities of starch-grains or in the differentia- 

 tion of different kinds of starch or for other incidental purposes. This method of differ- 

 entiating starches seemed so promising that five such reagents were selected. For obvious 

 reasons choice was made of those which differ widely in chemical composition and which 

 yield sufficiently prompt and characteristic results. Those selected include chloral hydrate- 

 iodine, pyrogallic acid, chi-omic acid, ferric chloride, and Purdy's solution. 



The chloral hydrate-iodine solution was prepared by saturating a saturated solution 

 of chloral hydi'ate with iodine. This solution, sooner or later, not only causes swelling and 

 ultimate partial dissolution of the grains, but also, owing to the presence of iodine, yields 

 important accompanying color reactions; and it is on the whole to be regarded as a very 

 ^•aluable reagent. 



Chromic acid was used in the form of a 25 per cent solution, and it is the only one of 

 the five reagents that causes within the periods of observation a complete disintegration 

 of the grains. It gives rise to gas bubbles during the decomposition processes. 



The pyi-ogallic acid solution was prepared by making a saturated solution and diluting 

 this with 3 parts of water, adding oxalic acid in the proportion of 4 per cent to prevent 

 oxidation. 



The ferric chloride solution consisted of equal parts of a saturated solution and water. 



Purdy's solution was made of equal volumes of the standard solution and water. 



The last reagent was usually found to be the least active of the five, and it is, so far 

 as the effects on the grains are concerned, probably essentially an aqueous solution of i)o- 

 tassium hydroxide, and therefore hkely possesses no advantages, except perhaps in keeping 

 qualities, over the simple aqueous solution. Oxygen or exposure to the air favors the ac- 

 tions of pyrogallic acid, but hinders those of clJoral hydrate and ferric chloride. In the 

 former case, the grains near the edge, or on the outside, of the cover-slip are decidedly 

 more affected than those within, while with the latter the opposite is true. 



There are some forms of commercial chloral hydrate that have very little action, 

 which may be due to uuder-hydration or over-hydration. The crystals put up by Schering 

 were used throughout this investigation. It is important that fresh solutions of the 

 reagents be ])repared at short intervals, as all tend to deteriorate, and it is well to let them 

 stand over night before using (see page 311). 



In using these reagents a small amount of starch is placed on a slide, several drops 

 of the reagent added, a cover glass put on, and the progress of events examined under the 

 microscope. In using a given reagent with a given kind of starch, it was found that there 

 occurred a certain amount of variation in the effects from time to time, which are prob- 

 ably to be attributed chiefly to variations in temperature, so that these studies were made 

 as far as possible under constant temperature conditions. The variations were unim- 

 portant. These agents give rise to gelatinization and swelling of the gi-ain, and cause the 

 existence of the outer and inner parts of the grains to become very conspicuous — the outer 

 part becoming sac-like and inclosing a less dense or semifluid substance. 



PREPARATION OF THE STARCHES. 



The starches used in this research were for the most part prepared from the under- 

 ground portions of the plants, that is, from tubers, rhizomes, bulbs, corms, etc. In a few 

 instances the specimens were obtained from the stalk, as from Diejfenbachia; occasionally 

 from both the stalk and fruit, as from Musa; occasionally from pseudo-tubers, as from 

 Orchis; or from the fruit, as from Castanea and Quercus; or from tiie side-shoots, as from 

 Cycas; or from the seeds, as in Graminacew. Unless otherwise stated it will be understood 

 that the starch is from underground i)arts. 



The starches were prepared by comminuting the specimen by the aid of an ordinary 

 kitchen grater or nutmeg grater, or of very coarse sand-paper, or of a small drug-mill. 



