George Harlan Dungan.

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UNIVERSITY OF ILLINOIS

Agricultural Experiment Station



BULLETIN NO. 284



THE INFLUENCE OF PLANT

INJURY AND THE ROOT ROT DISEASES

UPON THE PHYSICAL AND CHEMICAL

COMPOSITION OF CORN GRAIN



BY GEORGE H. DUNCAN




URBANA, ILLINOIS, DECEMBER, 1926



SUMMARY

Constricting the ear-shanks and stalks of corn by breaking the
supporting tissue in these structures without completely severing the
vascular elements had essentially the same effect upon the yield and
composition of corn grain as premature harvesting. Constriction of the
shank was much more detrimental to ear development than the break-
ing of the stalk.

Breaking the shanks when the ears were in the soft-dough stage
caused the greatest degree of chaffiness in the grain. Chemical analy-
ses showed that there was no definite correlation between kernel
"starchiness" and quantity of starch in the grain, as has been com-
monly supposed. In fact the quantity of true starch was often less in
the "starchy" than in the horny grain. In the light of these results it
is suggested that the term "floury" be substituted for the word
"starchy" in describing the corn containing more than a normal quan-
tity of soft, dry, friable material.

The percentage of total nitrogen, hemicellulose, and non-hydro-
lyzable material was distinctly higher in grain from ears produced on
broken shanks than in grain from ears produced on sound shanks. The
mutilation of the shanks, on the other hand, resulted in a greatly re-
duced proportion of ether extract and starch in the grain.

Inoculation of the seed at planting time with corn root rot organ-
isms resulted in the production of grain having a specific gravity .027
lower than that from the adjoining uninoculated plants.

The samples of grain produced by plants that grew in the inocu-
lated hills absorbed an average of 5.78 percent more water than the
samples from plants growing in uninoculated hills, with odds .greater
than 9999 to 1 that this difference was not due to chance.

Chemical analyses did not show any significant differences be-
tween the grain produced in the inoculated and uninoculated hills.
There was slightly more nitrogen, on the average, in the corn from in-
oculated seed, and a little less ether extract and total sugar. Even
tho the average differences in total nitrogen, ether extract, and total
sugar were not large, the odds indicating the significance of these re-
sults were above 30 to 1.

Analyses of two lots of horny and two lots of floury corn of the
Learning variety showed that there was no significant distinction in
chemical composition between these two types of corn. Upon germi-
nation, however, the horny corn was found to contain a greater pro-
portion of soluble starch and dextrins than the floury corn, and starch
digestion in the horny corn was somewhat more rapid than in the
floury corn. This phenomenon offers an explanation for the superior
vigor of seedlings from horny corn, so frequently observed.



THE INFLUENCE OF PLANT

INJURY AND THE ROOT ROT DISEASES

UPON THE PHYSICAL AND CHEMICAL

COMPOSITION OF CORN GRAIN

By GEORGE H. DUNCAN, Assistant Chief in Crop Production

INTRODUCTION

For a number of years the Illinois Agricultural Experiment Sta-
tion has advised farmers to eliminate all extremely "starchy" ears from
their seed stock. This recommendation has been based upon the re-
sults of investigations into the causes of the corn rot diseases obtained
by the Station and the Office of Cereal Investigations of the U. S. De-
partment of Agriculture working cooperatively.

In these investigations by Holbert and associates 13 the fact is
established that a high degree of correlation exists between certain ear
characters in seed corn and susceptibility of seedlings developed from
such ears to infection by root rot organisms. They give the important
diagnostic features of diseased seed corn as: (a) a lack of luster
in the ear; (b) a discolored or shredded shank attachment; (c) a
bleached ear tip; (d) an extremely deep indentation at the crown
of the kernel ; (e) a relatively high proportion of soft to horny starch
in the endosperm; and (f) a dull wrinkled kernel, especially in the
region of the germ. It is evident that these characters are not en-
tirely independent of one another, for an increased percentage of soft
starch in the endosperm tends to decrease the luster of the ear, and a
high content of soft starch is often accompanied also by deep kernel
indentation. Thus it would seem that the composition of the grain,
especially the proportion of soft to horny starch in the endosperm, is
a very important factor, if not the most important single factor, in
determining susceptibility or resistance to disease, or at least in indi-
cating such susceptibility or resistance. The data from one experiment
in which horny and "starchy" kernels were selected from the same ears
and grown in comparative yield trials, showed a reduction of 12 bush-
els an acre for the "starchy" seed. The results of investigations over
a period of seven years showed a decrease of 7.9 bushels an acre for
"starchy" seed, with odds greater than one million to one that this
difference was not the result of chance.

Trost 38 found that ears of the "starchy" class were character-
ized by a larger percentage of infection on the germinator than horny

255



256 BULLETIN No. 284 [December,

ears from the same seed sample. In experiments using Reid Yellow
Dent corn as seed, the "starchy" ears produced a larger number of
weak plants and 14 percent lower yield than did ears less "starchy" in
composition.

No data, however, have as yet been obtained as to why corn con-
taining a large proportion of soft starch is more susceptible to fungus
invasion than corn containing a smaller proportion, and very little di-
rect knowledge is available as to the factors that influence the physical
composition of the endosperm in corn grain. In general it may be said
that the agencies which determine the type of endosperm in corn
divide themselves into two classes; namely, hereditary factors, and
those factors that may be included under the general term environ-
ment. The present work has been confined to a study of certain fact-
ors in the second group ; namely, the influence that stalk and shank in-
jury may have at two different stages in the development of the ear,
and the effect of the inoculation of the seed at planting time with four
organisms, viz., Diplodia zeae (Schw.) Lev., Fusarium moniliforme
Sheldon, Gibberella saubinetii (Mont.) Sacc., and Rhizopus spp., any
one of which is capable of producing a corn root rot disease. Data
are also presented on the comparative chemical composition of starchy
and horny kernels at different stages during the process of germination.

REVIEW OF LITERATURE

The literature reporting the influence of ecological factors on the
physical and chemical composition of cereal grains is voluminous, and
this summary is not presented as in any sense exhausting that general
subject. Only those papers are reviewed that show the influence of
the stage of harvesting and plant injury from mechanical or parasitic
causes on the composition of the grain.

The monumental researches of Hornberger 15 in 1882 constitute
one of the most intensive pieces of work on the corn plant up to that
time. Later Schweitzer, 28 Jones and Huston, 18 and Smith, 32 engaged in
the study of the same problem. These workers agreed with Horn-
berger, 18 who found that as the grain developed beyond the milk stage
the proportion of ether extract increased, while the percentage of
total nitrogen and ash decreased. They also noted that the relative
amount of carbohydrates increased with maturity, while that of the
crude fiber decreased. The investigations of Ince 17 also showed a larger
proportion of crude fat and nitrogen-free extract in mature grain than
in immature grain. Bushey 3 reported that corn cut short in its devel-
opment by frost contained higher percentages of ash and crube fiber
than unfrosted corn. A comparatively high proportion of the nitrogen
of the soft corn was in the amid-albumin and globulin forms. Hume,
Champlin, and Loomis 16 noted that completely mature corn contained



1926} INFLUENCE OF PLANT INJURY ON COMPOSITION OF CORN 257

the largest percentage of oil. Appleman 1 found that the grain of sweet
corn decreased markedly in content both of total sugar and of reducing
sugar as it developed from the milk to the dough stage. Kent, Patrick,
Eaton, and Heileman 20 and Curtiss and Patrick 6 found that after the
ears had reached the dough stage there was but little change in the
composition of the grain, altho there was a gain in yield per acre.

Analyses reported by Saunders 27 showed that there was a rapid
reduction in proportion of crude protein in the wheat grain during its
early development, and that afterwards there was a gradual percent-
age increase in this material. Similar results were also obtained by
McDowell 25 in both winter and spring wheat, altho the percentage in-
crease in protein following the early rapid decrease was fluctuating in
character.

LeClerc 21 working with wheat, and Failyer and Willard 10 work-
ing with both oats and wheat, found that late cutting resulted in a
slightly reduced proportion of nitrogen in the grain.

Kedzie 19 made analyses of wheat grain at a great many stages, be-
ginning when the heads were just past the "blossom" stage and con-
tinuing until the grain was dead ripe and the straw fallen. His data
showed a decrease in the percentage of ash and crude-fiber content as
the grain matured. The relative amounts of albuminoids and amid
nitrogen decreased rapidly up to the hard-dough stage, after which
they increased slightly. The proportion of ether extract decreased un-
til the grain was in the milk stage, after which it rose gradually. The
percentage of nitrogen-free extract increased up to the hard-dough
stage, then dropped off slightly.

Data presented by Shutt 31 indicated that wheat grain injured by
frost before it was completely mature contained a lower percentage of
both total nitrogen and albuminoid nitrogen. Blish 2 reported that
"frozen wheat contains larger amounts of non-protein nitrogen, reduc-
ing sugars, and acid-reacting constituents than does sound wheat. The
non-protein nitrogen of the frozen wheat carries a considerably higher
percentage of a-amino nitrogen than that of sound wheat."

Severe rust injury hastened the ripening of wheat, and according
to Shutt 30 markedly increased the percentage of protein, crude fiber,
and ash, and lowered the proportion of fat and nitrogen-free extract.
Similar results as to the effect of rust on the ash and protein contents
of the wheat grain were obtained by Stoa. 35 Headden, 12 however,
stated that the shrunken berries resulting from rust infection were not
high in protein. He pointed out that protein and starch were laid
down simultaneously, and the effect of rust was to prevent the transfer
of the filling material to the berries.

Microchemical studies by Eckerson 9 indicated that starch was
formed in the endosperm cells of the developing wheat grain soon after
the formation of the cell walls, and that this process continued until



258 BULLETIN No. 284 [December,

desiccation began. Storage proteins were not formed in the endosperm
until the drying of the grain caused the amino acids present to con-
dense into proteins.

MATERIALS AND METHODS

In the experiment to determine the effect of breaking the stalks
and ear-shanks, yellow dent corn obtained from James R. Holbert of
the U. S. Department of Agriculture, was used as seed. It was of F^
hybrid material between pure-line strains, and consequently was much
more uniform in genetic constitution than average open-pollinated
corn. Seed of this corn was planted on the University South Farm
and received the usual cultivation under field conditions.

On August 17, when the grain was in the "milk" stage, and also on
August 31, after the grain had reached the "soft-dough" stage, a num-
ber of the ears were broken down to the extent of markedly constrict-
ing the supporting shank but not severing the ear from the stalk. Also,
on these two dates, a number of plants were broken over, so that a
definite constriction resulted between the nodes of the stalk about one
foot from the surface of the ground. The grain from plants so treated
was analyzed and the results compared with similar data from ears
produced on plants which had not been mutilated.

The seed used in the experiment to determine the effect of plant
infection with corn root, stalk, and ear rot-producing organisms on the
composition of the grain was obtained from the Plant Breeding Divi-
sion of the University of Illinois. This seed had been inbred for two
generations. Two ears possessed an extremely horny endosperm and
two an endosperm containing a high proportion of soft starch. These
were planted on May 16, in soil that had grown alfalfa for the twelve
years prior to this corn crop. At the time of planting, the alternate
hills were inoculated with a pure culture of some one of the following
organisms: Diplodia zeae (Schw.) Lev., Fusarium monitiforme Shel-
don, Gibberella saubinetii (Mont.) Sacc., and Rhizopus spp. The
grains were laid, germs up, in the hill. Two horny kernels were placed
about an inch apart on the north side of the hill and two "starchy"
kernels were similarly placed on the south side of the hill. Two drops
of a heavy spore suspension of one of the above-mentioned organisms
were placed on the germ face of each kernel in the alternate hills. In
order to make sure of getting an infection with Diplodia zeae a frag-
ment of shredded cornstalk bearing a pure culture of this organism
was placed between the kernels in the hill in addition to the spore sus-
pension placed on the kernels. Two drops of distilled water were placed
on the germ face of each kernel in the uninoculated hills. After the
moisture in the drops of spore suspension had partially dried down,
the corn was covered with moist soil which was pressed firmly down
over the seed.



1926] INFLUENCE OF PLANT INJURY ON COMPOSITION OF CORN 259

The cultures of Fusarium moniliforme and Rhizopus spp. were iso-
lated from germinating corn. The cultures of Diplodia zeae and Gib-
berella saubinetii (the latter designated as strain 259) were obtained
from Miss Helen Johann, of the U. S. Department of Agriculture.

In the study of the progressive chemical changes occurring dur-
ing germination in horny corn, as compared with those occuring in
"starchy" corn, the Learning variety was used. This lot of corn con-
sisted of twelve ears, six of which were smoothly indented and high
in percentage of horny starch and six of which were rough in indenta-
tion and relatively high in soft starch. All of these ears had been
carefully field-selected early in November and were alike in that they
had all been produced on apparently healthy plants.

These ears were divided into four lots of three ears each. Lots A
and C were made up of horny ears, and Lots B and D of "starchy"
ears (Figs. 1 and 2).

Five sets of germination tests were started in the temperature-
control chamber, which was maintained at 16 C. in the plant pathol-
ogy greenhouse at the University of Wisconsin a . Each test contained
20 kernels from each ear, the kernels in each case being removed from
the same rows on the ear. The corn was germinated between muslin
cloths over moist sand. At the end of the 2d, 4th, 6th, 8th, and 10th
days a set of the tests was removed and the kernels or seedlings dried
for analysis. Drying was effected by placing the samples in the oven
at 100 C. for one hour to kill the enzyms, followed by 18 hours of
drying with the oven door open (Link and Tottingham 22 ) .

CHEMICAL METHODS

All samples of corn grain analyzed were finely ground and passed
thru a 100-mesh sieve. The total nitrogen determinations were made
by the Official Gunning method modified to include the nitrogen of
nitrates. 7 The soluble nitrogen was extracted by soaking the ground
grain for four hours in 50 cc. of distilled water to each gram of sample.

The sugars were extracted from the ether-extract-free sample with
90 percent ethyl alcohol, by gently boiling the sample for one hour.
After the alcohol was evaporated, the sirupy residue was taken up with
water and the solution clarified with neutral lead acetate. The excess
lead acetate in the filtrate was then removed by adding sodium sulfate
and sodium carbonate in the proportion of nine parts of the former to
one of the latter. Phenolphthalein was used to indicate when defeca-
tion was complete. After filtering, the reducing sugar was determined,
an aliquot of the clear filtrate being used for this purpose. Total su-
gars were determined after hydrolysis in 2.5-percent hydrochloric acid,
by boiling on a sand bath for one hour.



"All the germinative tests and chemical analyses in this investigation were
made at the University of Wisconsin.



260



BULLETIN No. 284



[December,






FIG. 1. HORNY EARS OP LEAMING CORN USED IN STUDY OF CHEMICAL

CHANGES OCCURRING DURING GERMINATION

Kernels from these two lots of horny corn and from two lots of
"starchy" corn (Fig. 2) were germinated, and the chemical changes
taking place during germination were determined by analyzing samples
taken at two-day intervals. The results are described on pages 269
to 275.



Dextrins and soluble starch were extracted with cold water in
which the sugar- free sample was allowed to soak overnight. The fil-
trate was hydrolyzed by boiling in 2.5-percent hydrochloric acid for
2Y 2 hours.

The solution containing the insoluble starch was boiled for three
minutes to gelatinize the starch present. When cooled to 38 C., fresh
saliva was added to hydrolyze the starch to maltose. The complete
conversion of starch to maltose was indicated by failure of the mater-
ial to give a blue color with iodin. The maltose extract was hydro-
lyzed to glucose by adding hydrochloric acid until the concentration
of the acid in the solution was 2.5 percent and by boiling over a sand
bath for 2% hours.

The residue was washed into a flask and submitted to hydrolysis
in 2.5-percent hydrochloric acid for one hour. The filtrate was consid-
ered to be hemicellulose.



1926} INFLUENCE OF PLANT INJURY ON COMPOSITION OF CORN



261





FIG. 2. "STARCHY" EARS OF LEAMING CORN USED IN THE

GERMINATION STUDY

The relative proportion of horny and soft starch in these ears is
.indicated by the transverse and longitudinal sections of the grain. A
preliminary analysis showed practically the same percentage composi-
tion for both horny and "starchy" lots, but their specific gravity was
markedly different.



The quantity of sugars, soluble starch, and dextrins, insoluble
starch, and hemicellulose was ascertained by determining the power of
these substances to reduce the copper of Fehling's solution. The
amount of copper reduced was measured by the Shaffer-Hartman 29
iodometric titration method.

EXPERIMENTAL RESULTS
EFFECT OF BROKEN STALK AND EAR SHANKS

Weak and diseased corn plants are frequently not strong enough
to support the ears to complete maturity ; the increasing weight of the
ear may exceed the strength of the supporting tissue of the shank, or
the stalk itself may be deficient in mechanical substance and break
over under stress of wind. To simulate what so often happens in na-



262



BULLETIN No. 284



[December,




1926}



INFLUENCE OF PLANT INJURY ON COMPOSITION OF CORN



263



ture, a number of ear shanks and cornstalks were artificially broken
on August 17, when the grain was in the milk stage, and on August 31,
when the grain was in the soft-dough stage.

The breaking-over of the stalks and ear shanks did not in any
case involve the complete severance of the ear from the shank, nor of
the upper portion of the stalk from the stub. The vascular elements,
or at least most of them, remained intact. There was, however, a dis-
tinct constriction produced, which doubtless interfered greatly with
the translocation of reserve and building materials to the ear. It would
seem that the constriction of the shank, brought about by breaking
down the ears, would be greater than that of the stalk, owing to the
fact that the break caused a departure of approximately 120 from
the normal position of the shank, whereas the break of the stalk re-
sulted in a declination of only a little over 90.

That the constriction of the shank and stalk produced a marked
effect upon the size and character of the ears and kernels may be ob-
served by reference to Fig. 3 and by a study of the data presented in
Table 1.

The breaking of the ear-shank produced a much greater hindrance
to the proper filling of the grain than the constriction of the stalk.
This is particularly pronounced in the weight of ears, the weight of



TABLE I.-W-EFFECT OF BREAKING THE SHANKS AND STALKS OF CORN ON WEIGHT OF
EARS (GRAIN AND COB), WEIGHT OF TEN KERNELS FROM EACH EAR, AND
GERMINATIVE VIGOR OF THE KERNELS















Average












Average


length of


Group


Treatment


Average
weight
of ears


Average
weight
of 10
kernels


Average
germi-
nation in
soil


number
of secon-
dary
roots
per seed-


plumule
at end of
8-day
germi-
nation












ling


test in














soil






grams


grams


perct.




mm.


A


Parent ears




2.96


98.0


2.88


120


B


Shank broken when grain














was in milk stage


78.4


.69


52.3


1.60


74


C


Shank broken when grain














was in soft-dough stage . .


191.8


1.59


97.1


1.65


75


D


Stalk broken when grain














was in milk stage


272.4


2.37


88.1


2.67


87


E


Stalk broken when grain














was in soft-dough stage . .


261.0


2.38


88.3


2.96


100


F


Ears from erect plants and














from unbroken shanks . . .


331.1


2.86


98.1


3.43


117



264



BULLETIN No. 284



[December,



ten kernels, the number of secondary roots, and the length of plumules.
Even tho the delay in breaking the shanks until the soft-dough stage
resulted in a much greater weight of ear and kernel, and also increased
the percentage of germination, the vigor of the seedlings produced by
this corn, as measured by number of secondary roots and length of
plumule, was only very slightly better than that of the corn the shanks
of which were broken when the grain was in the milk stage.

The chaffy character of the ears in Group C, Fig. 3, is striking.
All the ears in this experiment that had their shanks broken when the
grain was in the soft-dough stage contained a very high proportion of
soft starch. A comparison of the various lots of ears in respect to the
relative quantity of soft and horny starch in the endosperm of the
kernels is afforded in Table 2.

The parent ears and those that matured on erect plants and on
unbroken shanks contained the highest percentage of horny and the



TABLE 2. EFFECT OF BREAKING THE SHANKS AND STALKS OF CORN AT Two

DIFFERENT STAGES OF GROWTH, ON THE KERNEL CHARACTER

OF THE EARS PRODUCED



Group


Treatment


Number
of ears
repre-
sented


Kernel character:
percentage of ears that were


Horny


Medium


Starchy


A
B

C
D
E
F


Parent ears


5
13

7
10
12
11


80


20

8
27



30

10

18


20
70
100
70
92
55


Shank broken when grain was in
milk stage


Shank broken when grain was in
soft-dough stage ... ...


Stalk broken when grain was in
milk stage


Stalk broken when grain was in
soft- dough stage


Ears from erect plants and from
unbroken shanks



lowest percentage of "starchy" individuals. However, as may be seen
from Table 3, the chemical composition of the kernels in respect to the
quantity of true starch which they contained, did not harmonize with
their physical appearance.

The early constriction of the shanks resulted in grain high in total
nitrogen, hemicellulose, and crude fibrous material that was non-hy-


1 3

Online LibraryGeorge Harlan DunganThe influence of plant injury and the root rot diseases upon the physical and chemical composition of corn grain → online text (page 1 of 3)