METHODS USED IN THE STUD! OF STAR( III 



tions we a edO I >, 0.25, and at Lugol'a solution. 



Pour serial reactions were studied; two with raw 

 and two with gelatinized starch, la the firsl two, the 

 slides are prepared aa in the polarization examinations, 

 substituting solutions of iodine for the balsam and 

 examining the slides in ordinar) lighl with a fully open 

 diaphragm and low power. In the firsl reaction 2 drops 

 of 0.25 per cent Lugol's solution are placed on the 

 Btarch, tlif sink' quickly adjusted on the stage of the 

 microscope, and the color reaction in quality and quan- 



ii once determined, the quantitative value recorded 

 being taken as the standard of comparison in relation to 

 other starches. Here, as in the polarization determina- 



. it was found necessary to adopt an arbitrary scale 

 and starch standards. The same scale is used as tor 

 the polarization values, but the terms light, deep, etc., 

 were substituted for low, high, etc. Moreover, it was 

 found necessarj to modifj the selection of starches to be 

 used as standards. The Btarch of Solatium tuberosum 

 was taken as having a value of CO or " moderately deep," 

 thai of Crinum moorei as having a value of 50 or " mod- 

 erate," and that of Waisonia humilis&a having a value of 

 30 .ir "light," with corresponding intermediate figures 

 ami terms as in the polariscopic di terminations. 



The second experiment is made, using 0.125 per cenl 

 solution, often bringing oul color peculiarities which may 

 be obscured or not be observed when the reagent is 

 stronger. 



The third and fourth experiments are made with 

 boiled starch with the object of eliciting peculiarities of 



tion of the grains, solution, grain-residues, and cap- 



. After heating the grains until complete gelatiniza- 

 tion occurs a variable amount of the starch passes into 

 solution, so that both -rains ami solution give starch 

 reactions. Upon boiling the preparation for '.' minutes 

 a comparatvely large amount of the starch passes into 

 solution, and the remains of the grains appear in the 

 form of grain-residues which are made up of partially 

 disintegrated grains (capsules with variable amount- of 

 contents), together with some capsules that are almost 

 or wholly free of starch contents. 



In the thinl experiment 0.05 gram of starch is plat ed 

 in 20 c.c. of water and carefully heated over a bunsen 

 burner only to the point of complete gelatinization. To 

 •3 c.c. of tl on is added 2 c.c. of a '.' per cent 



Lugol's solution, and then th.' colorations of grains ami 

 solution are determined by microscopic examination. 



In the fourth experiment the remainder of the boiled 

 preparation is boiled for 2 minutes to further break 

 down the starch grains; then I c.c. of the 2 per cent 

 Lugol's solution added; and then microscopic deter- 

 mination made of the colorations of grain re- 

 capsules, and solution. 



7. Ami. ink Ki v nous. 



A number of anilines have 5 ad by various 



tigators to I f value in the differentiation of 



Marches from diifcrent sources, of different grains of 



ind of starch, and of different part-, of indi- 

 vidual grains. Some experimenters ha 

 double or triple staii I no doubt thai 



use of double or triple Btaina would bi 

 at least, man) points of much histological im 

 hut this would have involved the carryii the 



histological examination i detail i pro- 



hibitive in a research of this character. Safranin and 

 gentian violet wi re - lected, not because they are prob- 

 ably the I" e .-tains for differential purposes, hut 

 because they have been found verj useful in starch e 



iiialion- and a- they yield single I "lor read 



Aniline colors in solution, ly when in ■■ 



solution ami exposed to light, are notably unsb 

 in order to secure strictlj comparable results a quantity 

 of a relatively strong standard solution was pre] 

 and kept in the dark, tightly corked. The stock sol,. 

 were composed of 0.25 gram of anilini n ith 150 i 

 distilled water, from day to day dilute soluti 

 prepared by adding 33 c.c. ,,\ wal 



solution: 15 c.c. of the latter solution an in a 



test-tube containing 0.0 1 ! gram of starch, the preparation 

 agitated, 1 or 2 drops withdrawn in a minute and exam- 

 ined under the micro-cope, and a final examination made 

 at the end of half an hour. In ina- 



tions the to-, ope is used, as in the iodim 



with a fully open diaphragm and low power. Ow 



the relatively slow reaction, the value- for comparative 



purposes wen taken at thi end of a half hour h 



of immediately, a- in the first iodine reaction. The 



method of valuation is the same as in the iodine 



. hut the starch standards for t ! are: 



Solanum tuberosum, value 90, "very deep": Amaryllis 

 belladonna, value 50, " moderate " ; Fn esia reft 

 value 30, " light" 



8. Tf.mi'ii:.\ii i: Gelatinizatioit. 



While the records of various investigators ind 

 that there are more or less marked dill', a the 



temperatures of gelatinization of different kite 

 starches, and even in case of different grains of the 

 same starches, the figures applying to the same kind 

 of starch are generally so at variance that not much value 

 ,in. Thi - of fallacy in 



such observations, unless the determinal made 



with the greatest precautions, are well known to • 

 biochemist. We therefore carried out this work with 

 al care. A long quadrangular water-bath was 

 used, holding about I litersol water;oneend was: 

 over the gas tlaine, and m tie- other end was inserted a 

 thermometer which was calibrated in jrade, 



hut which could readily he read in hundredths. A small 

 quantity of starch with L0 v.r. of water v. I in a 



test-tube, into which was inserted, through a pert' 

 cork, a thi milar to the one in | 



hath, and the test-tube immersi nded wire 



basket in the part of the v h farthest from the 



flame. The temperature of the wati 



