FIELD MUSEUM OF NATURAL HISTORY

FOUNDED BY MARSHALL FIELD, 1893

PUBLICATION 275

BOTANICAL SERIES VOL. IK, No. 1

THE LIBRARY Or

-

UNIVERSITY OF ILLINOIS

THE DIFFERENTIAL ANALYSIS

OF STARCHES

BY

JAMES B. McNAm

ASSISTANT CURATOR OF ECONOMIC BOTANY

B. E. DAHLGREN

ACTING CURATOR, DEPARTMENT OP BOTANY

EDITOR

CHICAGO, U. S. A.

JUNE 19, 1930

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FIELD MUSEUM OF NATURAL HISTORY

FOUNDED BY MARSHALL FIELD, 1893

PUBLICATION 275

BOTANICAL SERIES VOL. IX, No. 1

THE LIBRARY OF THE

TEB141Q38

UNIVERSITY OF ILLINOIS

THE DIFFERENTIAL ANALYSIS

OF STARCHES

BY

JAMES B. Me N AIR

ASSISTANT CURATOR OF ECONOMIC BOTANY

B. E. DAHLGREN

ACTING CURATOR, DEPARTMENT OF BOTANY

EDITOR

CHICAGO, U. S. A.

JUNE 19, 1930

FEINTED IN THE UNITED STATES OF AMERICA

BT FIELD MUSEUM PRESS

5S0.5

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THE DIFFERENTIAL ANALYSIS OF STARCHES

JAMES B. McNAiR

INTRODUCTION

Reichert, in his excellent and extensive monograph on starch,

gives a historical review dealing with the starch substance, the

primary and reverted decomposition products of starch, and the

structure, form and mechanism of formation of the starch grain.

He outlines the chief forms and classifications of starch grains and

has determined in painstaking detail histological, chemical and

physical characteristics of 350 starches. For each of these he gives

two photomicrographs showing their appearance in ordinary and

polarized light.

Extensive tables of the various histological, physical and chemical

characteristics are included in Reichert's work. These tables, as

they stand, arranged in botanical order, are of limited usefulness

to the analytical chemist. For his own convenience the author has

rearranged them according to physical and chemical properties and

it is thought that thus rearranged they will be found of interest

and utility to all who have occasion to deal with starches in an

analytical way.

It is sometimes of importance to be able to determine the botani-

cal source from which a certain starch sample is derived. This

task, formerly never attempted, is now possible because of Reichert's

work. It has become possible not only to tell the difference between

starches of different plant families, as corn starch from potato

starch, but also to determine between the starches of the members

of one family of plants as in the case of wheat, rye and barley; or

even to tell whether the starch has come from different varieties

of one species — for example, the starch of ginger root from Jamaica,

and of ginger root from Cochin China, or between the starches of

pod corn, pop corn, flint corn, dent corn, soft corn, sweet corn, and

starchy-sweet corn.

Some of these differential determinations may be made by the

appearance of the starch grains under the microscope. In some

cases chemical analysis will also aid, when, for instance, the per-

centage of hemicellulose varies between different species as was

shown by Ling and Nanji1 or when the ash is not the same either

qualitatively or quantitatively as has been found to be the case in

many instances, or when the phosphoric acid present in organic

combination varies in different starches, or when the combined

fatty acid varies in quantity or quality between the starches.2 No

1 Arthur R. Ling and Dinshaw R. Nanji, Chemical Society of London, Journal,

CXXIII (1923), 2666-88.

2T. Clinton Taylor and John M. Nelson, American Chemical Society, Journal,

XLII (Easton, 1920), 1726-38.

4 FIELD MUSEUM OF NATURAL HISTORY— BOTANY, VOL. IX

doubt the physical properties of starches, such as their qualities of

viscosity and water absorption, can also be used in their differentia-

tion.

The investigations of Reichert3 have given us methods which

will differentiate between the various starches in a minimum of time

with a minimum of materials.

Reichert was able to differentiate between 350 different starches

by means of eleven processes of analysis, namely: (1) histological

microscopic examination to determine the form and size of the grains,

position and character of the hila, characteristics of the lamellae,

orientation, etc.; (2) the degree of polarization; (3) the iodine

reaction; (4) the gentian violet reaction; (5) the safranin reaction;

(6) the temperature of gelatinization; (7) the chloral hydrate-

iodine reaction; (8) the chromic acid reaction; (9) the pyrogallic

acid reaction; (10) the ferric chloride reaction; and (11) the Purdy

solution reaction.

These tests are carried put on small amounts of material by the

aid of microscope and require individually not more than one hour's

time. Reichert's processes of analysis are summarized at the end of

this paper.

PREPARATION OF THE SPECIMEN OF STARCH

Starch, as is well known, constitutes reserve food for the plant

and is stored in the leaves, stems, and roots.

Reichert's plant specimens were tubers, rhizomes, bulbs, corms, stalks, fruit,

pseudo-tubers, seeds, and in most cases underground parts. A specimen was

comminuted by the aid of an ordinary kitchen grater, nutmeg grater, sandpaper,

or small drug mill, in accordance with the quantity and physical characters of

the specimen. Four or five volumes of water were added to the ground specimen,

the mixture thoroughly stirred, the mass strained through four thicknesses of

cheesecloth, and the pulp then washed with sufficient water and strained as

before. The starch-water preparation is decanted in cylinders and the starch is

cleansed by repeated washing and decantation. Finally the starch is collected

in shallow dishes, the water as far as possible drained off and the preparation

dried at a temperature of 50° C. By this simple means starches can be prepared,

which are with rare exceptions practically free from gross impurities; that is,

as pure as the nature of Reichert's investigations demanded.

HISTOLOGICAL METHOD

Magnifications ranging from 85 to 400, sometimes higher, are used, according

to the size of the grains and incidental conditions. A sufficient amount of dried

starch is placed on a slide and mounted in a very dilute Lugol's solution, care

being taken not to add a larger quantity of iodine than is sufficient to accentuate

the lamellae. Since starches of different sources show wide differences in the

intensity with which they become colored with iodine it is found convenient to

3 Edward Tyson Reichert, The Differentiation and Specificity of Starch in Rela-

tion to Genera, Species, etc., Carnegie Institution of Washington, Publication No.

173, Washington, D.C., 1913; A Biochemic Basis for the Study of Problems of Tax-

onomy, Heredity, Evolution, etc., Carnegie Institution of Washington, Publication

No. 270, Washington, D.C., 1919.

STARCH ANALYSIS 5

have on hand a number of solutions ranging from 1 to 2 per cent down. By the

aid of such ordinary microscope technique there are recorded the form and size

of the grain; the position and form of the hilum, or the assumed point of origin

of growth or center of organic structure; the form, number and other characteristics

of the lamellae; the characteristics pertaining to the form of the grains, whether

singly or in doublets, triplets, or aggregates, etc. In describing the grains the

terms "proximal end" and "distal end" have been adopted, the former being

the end nearer which the hilum is located. The "longitudinal axis" corresponds

with an imaginary line, extending from the proximal end through the hilum to

the distal end. In different starches and in different grains of the same kind of

starch this may be the long or the short axis. The measurements of eccentricity

of the hilum have reference to the distance of the hilum from the proximal end

of the longitudinal axis.

REACTIONS IN POLARIZED LIGHT WITHOUT AND WITH

SELENITE

Starches have been found to exhibit not only marked differences in the degrees

with which they rotate the plane of polarized light, but also differences in the

characteristics of the "interference figure" or "cross," as it is generally termed.

The general characteristics, distinctness, shape, regularity, and position of the

interference figure, and also the approximate degree of anisotropy or intensity

of polarization were readily studied. By the aid of selenite it was determined

whether the optical properties were negative or positive, and also the size, shape

and regularity of the quadrants, as well as the intensity and pureness of the blue

and yellow colors. In spherical grains with centrally located hila, the two parts

of the "cross" intersect at the hilum, or mathematic center, of the grain, so that

the term quadrant has a proper application; but in the case of grains having

eccentric hila the position of the point of intersection of the two parts of the

cross, together with their curvatures, may destroy every semblance of quadrants

according to the conventional definition of this word. This term has therefore

been used in a very broad sense throughout Reichert's investigation to indicate

the four parts of the grain that are defined by the two parts of the cross, in pref-

erence to the great multiplicity of terms that would be required to define these

parts if great accuracy were attempted. Likewise, for convenience the term

"lines" of the interference figure may be used in preference to the "arms" of the

cross.

All starches are "optically negative," hence no special references have been

made in the text in this particular.1

The slides for polariscopic examination are prepared as follows: The end of

a small spatula is thrust into the specimen of starch and moved about, with-

drawn and sharply tapped several times in the center of the slide, and the slide

jarred in a manner to cause a practically uniform distribution of the starch grains

in a single well-disseminated layer. The margins of this layer are carefully removed

so as to leave an area 12 mm. square. An expeditious way of removing the margin

so as to insure a uniform area of starch is to use as a wiper a piece of sheet celluloid

having a 12 mm. slot, wiping transversely and then longitudinally. A few drops

of balsam are carefully added at the center of the area, a cover-slip put on, and

the slide placed on the stage of the polarizing microscope. After determining

the degree of polarization, the selenite plate is introduced and the specimen

again examined.

In order to reduce the degree of polarization into values in comparative terms

and figures it was found desirable to adopt an arbitrary scale and to select three

JThe selenite plate enables one to determine the size, shape, and regularity

of the quadrants, as well as the intensity and pureness of the blue and yellow

colors.

4 FIELD MUSEUM OF NATURAL HISTORY — BOTANY, VOL. IX