Quantitative Investigations of Leaf Pigments From Their Inception in Buds Through Autumn
Coloration to Decomposition in Falling Leaves
Author(s): Jon E. Sanger
Reviewed work(s):
Source: Ecology, Vol. 52, No. 6 (Nov., 1971), pp. 1075-1089
Published by: Ecological Society of America
Stable URL: http://www.jstor.org/stable/1933816 .
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QUANTITATIVE INVESTIGATIONS OF LEAF PIGMENTS FROM THEIR INCEPTION
IN BUDS THROUGH AUTUMN COLORATION TO DECOMPOSITION
IN FALLING LEAVES'
JON E. SANGER2
Department of Botany, University of Minnesota, Minneapolis, Minnesota
Abstract. Pigments in hazel (Corylus americana), aspen (Populus tremuloides), and pin
oak (Quercus ellipsoidalis) leaves were measured from their inception in buds to development
of a summer maximum, and through the autumn coloration period to decomposition in dry
falling leaves. Leaves contained generally high but varying concentrations of chlorophyll and
carotenoid pigments throughout the summer months. The summer pigment variations among
the three species are discussed in the light of the usefulness of chlorophyll content as an index
of net primary productivity. During the autumn coloration period, preceding leaf desiccation
and fall, chlorophyll decays rapidly, producing low levels of pheophytin with only occasional
faint traces of pheophorbide and chlorophyllide during the period of most rapid chlorophyll
breakdown. The levels of carotenoids begin declining at the same time as chlorophyll, but at a
much slower rate. Violaxanthin disappears most rapidly, followed closely by neoxanthin.
Lutein and ,B-caroteneare the most stable carotenoids. Falling oak leaves, whether dropped
to the ground in autumn or held on the tree throughout the winter, in spring still contain
measurable amounts of lutein and 5-carotene and low concentrations of pheophytin a. Concurrent with the autumn degradation of plastid pigments is an abrupt and substantial rise in
anthocyanin of oak and hazel. At leaf-fall aspen and hazel leaves are devoid of all pigments.
(1932), who claimed autumn pigments were not carotene
in nature but "esterified xanthophylls" behaving
Each year vast quantities of plastid pigments are
produced in plant bodies during spring and early like carotenes in the partition test. Karrer and Walker
summer. Throughout summer the intense green of (1934) took the view that the autumn pigments were
the chlorophyll masks the yellow and orange colors degradation products of lutein and that the carotenes
of the carotenoids. In autumn preferential destruc- and xanthophylls both disappeared in autumn, carotion of the chlorophylls allows the more slowly dis- tenes going relatively more quickly than xanthophylls.
appearing carotenoids to color the countryside with Schertz (1929) studied the seasonal changes of the
a variety of orange and yellow hues. In many plants chloroplast pigments of several plants on the Mall
the coloration period is made even more brilliant by in Washington, D.C., but made no attempt to septhe appearance of the bronze and red colors of newly arate the individual chlorophylls and carotenoids,
formed anthocyanins. The fate of pigments follow- and his techniques in general were quite crude. Wolf
ing autumn coloration and subsequent leaf fall re- (1956) traced the changes in chlorophylls a and b
in autumn leaves, but he made no attempt to identify
mains essentially uninvestigated.
took a more careful
The subject of autumn pigment degradation has carotenoids. Goodwin (1958)
the
in
pigments, using
changes
plant
look
at
seasonal
been of particular scientific interest since the work
carotenoids, chlorodeal
with
techniques
to
modern
of Willstitter and Stoll (1918), who reported lower
phylls, and anthocyanins. His studies, however, did
chlorophyll values in yellow leaves than in green
not include measurement of pigments from their inleaves. They did not, however, follow closely the
in leaf buds, and they were not carried far
loss of either chlorophyll or carotenoids. It quickly ception
late autumn and winter to ascertain the
into
enough
became evident to early investigators that carotof the pigments that are destined to
fate
eventual
enoids as well as chlorophylls underwent changes
become part of the organic fractions of forest soils,
during the autumn period. Details of many of the
early investigations, with special emphasis on their peats, or lake sediments.
The present study was undertaken for several
errors and inconsistencies, are discussed by GoodFirst, the data have an intrinsic interest bereasons.
win (1958). He notes particularly the work of
studies are available tracing pigments from
no
cause
believed
to
Tswett (1911), who
carotenoids
be alin leaf buds to their decomposition
their
initiation
most entirely epiphasic in autumn leaves, but with
of leaf coloration and desiccation
the
period
beyond
the major fraction being separable from ,3-carotene.
several ecological studies
in
late
autumn.
Second,
Tswett termed these pigments "autumn xanthophylls."
Also discussed is the work of Kuhn and Brockmann have supported the use of chlorophyll content per
plant and per unit landscape area as a productivity
1 Received September 30, 1969; accepted July 2, 1970.
index (Odum 1959, Brougham 1960, Bray 1960,
2 Present address: Dept. of Botany-Bacteriology, Ohio
1962, Whittaker and Garfine 1962, Ovington and
Wesleyan University, Delaware, Ohio 43015.
INTRODUCTION
JON E. SANGER
1076
Lawrence 1967, Sanger, unpublished data). Data
concerning changes in pigment concentrations of
selected components of a plant community throughout the growing season are necessary to establish the reliability and comparability of this index. Third, Goodwin's (1958) observations show
that no simple generalizations can be made for
the fate of carotenoids in autumn leaves because
of the great differences among the three trees
he examined. Fourth, before any accurate assessment of the significance of pigment levels in soils,
peats, and pond muds can be made, it is necessary
to know the types and amounts of pigments which
are likely to reach the soils from various terrestrial
sources. Last, nothing is known of the effect of varying autumn climates on pigment decomposition. The
relatively short, dry autumns of Minnesota may produce conditions far different for pigment decay than
those observed by Goodwin during the relatively
long, moist autumns of England.
The three species chosen for study were pin oak
(Quercus ellipsoidalis), American hazel (Corylus
americana), and quaking aspen (Populus tremuloides). The first two were selected because they are
the chief components of the tree and shrub layers in
the oak forest where soil humus layers were studied
for pigment composition (Sanger 1971). In addition,
both species exhibit development of anthocyanin
during autumn coloration. This was considered important because falling leaves are often reddish in
color, yet no trace of anthocyanin has been found
by the author or reported by others in soils, peats, or
pond muds. Aspen was chosen because it is found
commonly throughout the area in scattered clumps
and along the edges of the oak woodlands. In addition, anthocyanin does not develop in aspen leaves
during the autumn, enabling comparisons to be made
with pigment degradation in the species that do develop anthocyanin. It is also important that pin oak
leaves tend to overwinter on the tree while hazel and
aspen leaves do not.
METHODS
AND MATERIALS
A series of small branches totaling approximately
100 leaves (buds in spring), of which only a portion
was actually used, was picked at random throughout
the canopies of designated trees. No particular plants
were designated for hazel. Instead, branches were
picked at random from the area near the designated
oak tree. The leaves (blade + petiole) were harvested
from the branches, sealed in airtight polyethylene
bags, and taken to the laboratory to be processed
immediately or stored for less than 1 day at 2?C.
Prior to extraction 10-15 leaves were chosen at random from the initial 100 and their areas calculated.
Five to seven leaves were taken at random for drying, and a similar number for extraction of pigments.
Ecology, Vol. 52, No. 6
Dry, weighed leaves were ashed in a muffle furnace
at 550'C for 4 hr to enable calculation of pigments
per gram organic matter.
Pigments were extracted in a Waring blender with
90% acetone and 1 g MgCO3. The solution was filtered with successive portions of 90% acetone
through a Buchner funnel with a fritted disk of
coarse porosity until all pigment was removed, and
the final volume was brought up to 100 ml.
A 25-ml portion of the acetone extract was saponified by gentle shaking with an equal volume of
20% methanolic KOH for 2 hr in a flask covered
with a black polyethylene bag. Transfer of carotenoids to petroleum ether (30?-60?)
was carried
out in a separatory funnel in a partially darkened
room. The solution of petroleum ether and carotenoid was washed with distilled water, dried with
granular, anhydrous Na2SO4, and shaken with 90%
methanol in a separatory funnel. The hypophasic
component (xanthophyll) was drawn off and its volume brought up to 50 ml. The epiphasic component
(P-carotene) was washed with an additional 100 ml
of distilled water, dried with Na2SO4, and its volume
brought up to 50 ml. Measurements of absorbance
were made on a Beckman DU spectrophotometer at
445 mV. for total xanthophyll and at 451 my. for
total carotene.
An additional 25-ml portion of the original acetone extract was diluted to 50 ml in a volumetric
flask, and a carefully measured spectrum was recorded with a Beckman DU spectrophotometer from
640 mn to 700 mp to measure total chlorophyll.
The remaining 50 ml of the original acetone extract were transferred to diethyl ether. The solution
of pigment and ether was concentrated to approximately 5 ml by rapid evaporation with continued
swirling in a fume hood. Care was taken not to allow
the pigment to go to dryness. Squares of Whatman
No. 1 filter paper, 12 by 12 cm, were spotted in the
lower left-hand corner, placed in chromatography
jars with ground glass lids, covered by black, lightproof polyethylene film, and chromatographed in the
first direction with a mixture of 10 parts hexane
(practical grade from Eastman Kodak), 2.5 parts
petroleum ether (60?-70? ), and 2 parts acetone
(reagent grade). The papers were run in a second
direction in a similar manner with 10 parts hexane
(practical grade), 2.5 parts petroleum ether (60?70?), 1 part acetone, and 0.25 part methanol (technical grade). A two-dimensional chromatogram of
oak-leaf pigments is illustrated in Fig. 1. Identification of the separate spots was made from spectral
analysis. The separated pigment spots were cut out
of the paper, eluted with suitable solvents, and the
eludates made up to constant volumes. Beta-carotene
was eluted with petroleum ether; lutein, violaxanthin,
and neoxanthin with absolute ethanol; chlorophyll
INVESTIGATIONS OF LEAF PIGMENTS
Autumn 1971
1077
solutions of 80% and 90% acetone. Although 90%
acetone was used throughout, some water is always
a
- pheophytin
present in the plant tissue so that solutions are actub
l
--pheophytin
/
\Oally between 80% and 90% acetone.)
--lutein
L2--Absorption measurements were made with a Becktail
--luteir)
._____
---violaxanthin
?---___
(fman
DU spectrophotometer with 10-mm absorption
I
------____
---chlorophyllIa
cells at the blue maximum for carotenoids and the
red maximum for the chlorophylls. Pigment values
were calculated as milligrams per 100 g organic matter with the specific absorption coefficients listed in
a
---pheophorbide
--______________
?
b
Table 1. Accuracy of the calculations depends upon
(_"J--- - -pheophorbide
a
chiorophyllide
_------__
?----- ----_
the accuracy of measurement of a given volume of
(origin)
pigment extract after it is concentrated, applied, chro2nd dimension-.
matographed, and eluted from the paper chromatoFIG. 1. Oak-leaf pigments separated on a two-dimen- grams. Because volumes of pigment extract applied
sional paper chromatogram.
to chromatograms are so small, it is very difficult to
these measurements accurately enough for remake
TABLE1. Specific absorption coefficients used to calculate
liable calculation of pigment concentrations. For this
pigment concentrations
reason carotenoids per unit organic matter were first
Wavelength
ascertained by the partition technique and chloroof maximum
absorption
Absorption
by direct measurements of the original acephylls
Solvent
Author
coefficient
Pigment
(mO
tone extract. Individual pigments were then calcuChlorophyll a
lated from the chromatograms (see Sanger 1968).
Vernon
80 %a
665
90.8
and
In general there is close agreement between the pig(1960)
acetone
Chlorophyllide a
ment concentrations calculated in this study and pubVernon
64852.5
80%
Chlorophyll b
lished values. Pigments were identified by their vis(1960)
649
acetone
ible absorption spectra, and by their relative positions
Pheophytin a
on two-dimensional chromatograms employing two
Vernon
55.2
667
and
80%
or more different solvent systems (Strain 1938, Good(1960)
acetone
Pheophorbide a
win 1952, Watson 1953, Holt and Jacobs 1954,
Pheophytin b
Smith and Benitez 1955, Holden 1962, Colman and
655
Vernon
34.8
80%
and
(1960)
acetone
Pheophorbide b
Vishniac 1964, Michel-Wolwertz and Sironval 1965,
Strain et al. 1965, Strain and Svec 1966).
451
Petroleum Zscheile
258.0
/3-carotene
Anthocyanin was extracted and chromatographed
(1934)
ether
according to the technique of Harborne (1965). The
445
90%
Total xanthophyll
255.0
anthocyanin was concentrated by evaporation in a
methanol Sanger
rotary vacuum flash-evaporator, and the final volume
Strain
445
Abs.
255.0
Lutein
was carefully measured. To remove the last traces of
ethanol
(1938)
plastid pigments, circular chromatograms were run
445
Abs.
(assumed same as
Lutein tail
on 27-cm Whatman No. 1 filter paper. The chromaethassol
lutein)
togram was spotted with 0.1 ml of anthocyanin soluKarrer and
440
Abs.
tion and developed with 1-butanol-acetic acid-water
255.0
Violaxanthin
Jucker
ethanol
(4:1:5); the top layer was used as irrigation fluid.
(1950)
The anthocyanin was eluted from the paper with 1 %
Strain
438
Abs.
227.0
Neoxanthin
acidic methanol, and measurements of absorption
ethanol
(1938)
were made with a Beckman DU spectrophotometer.
1%
520
Geissman
The anthocyanin in the leaves studied had a spec70.0
Anthocyanin
acidic
(1955)
trum very similar to that of cyanidin, and amounts
methanol
were calculated from its specific absorption coefficient, which is listed in Table 1.
aExperimentsby Vernon (1960) and by the author reveal no significant
Q- ---B-carotene
-
0~~0
differencesin the specific absorptivity of pheophytin a and b, pheophorbide
a and b, and chlorophyllide a between 80 % and 90 % acetone.
a and b, pheophytin a and b, pheophorbide a and b,
and chlorophyllide a with 90% acetone. (Experiments by the author reveal no significant difference
in the specific absorptivity of pheophytin a and b,
pheophorbide a and b, and chlorophyllide a between
RESULTS AND DISCUSSION
Oak leaves
It is first helpful to consider changes in leaf area
and in the dry, ash-free weight per leaf and per unit
area of leaf tissue throughout the sampling period.
These data are presented for oak in Fig. 2 and 3.
Ecology, Vol. 52, No. 6
JON E. SANGER
1078
8-
I
1
C?
0 30a.
.
chloro.a
buds
unit
area
-
w
l
*break
I.
LU
40-
C4
w
4w.
40
3
,
.I
leWa aeaf
areak
30
. ...,, ..
pheophytin a
1W
M
10<
J
J
A
SON
D
J
F
M A
4. Temporal changes in chlorophyll a and b per
unit organic matter (oak).
FIG.
J
J
A
S
0 N
D
JF
MA
2. Temporal changes in organic matter per unit
da20area and average area per leaf (oak).
_
20<t^
leaf
FIG.
*
010
0IG 5. Temporal
p
Ua.
0
_1
500Ci
changes
chlorophyltot
Inin total
carotenoid
_
-l200'U
0
O 100
0
~
iorophy
~ ~
rl
~
~
~
~
~
10
4
all
f
ey.
preenphtin F
0M J J A S 0 N D J F M A
FIG. 5. Temporal changes in total chlorophyll and
carotenoid per unit organic matter of oak leaves.
M J
J A
SON
D J
F MA
FIG. 3. Temporaltchanges in organic matter per leaf
of oak.
Buds break in late May and leafing takes place
quickly. By mid-July leaves reach their average
maximum size and weight. The seasonal changes in
absolute leaf organic matter parallel the gradual
build-up of organic matter per unit leaf area as the
mesophyll tissue develops. There is a rise from 4.5
mg/cm2 in immature leaves to an average summer
value, beginning in early July, of about 7.8 mg/cm2.
At the initiation of autumn coloration, organic matter values drop somewhat, both per unit area and on
an absolute basis, and considerable variation is observed as the leaves are drying and the pigments are
decomposing. In late autumn and throughout winter,
when leaves are dry and brittle on the tree, the individual values become less erratic and do not decline appreciably by the end of the sampling period
in April from the average autumn value. In leaves
decaying on trees, pigments are broken down at a
rate far greater than that for total organic matter.
This is an important comparison to make with leaves
decaying on the forest floor (Sanger 1971). Changes
in pigment concentrations throughout the season are
expressed as milligrams per 100 g organic matter to
make the data more comparable with existing data
of pigments in soil humus layers and lake sediments.
Changes in the chlorophyll a and b content of oak
leaves from spring buds through the autumn coloration period to leaf desiccation and drop the following
spring are presented in Fig. 4. In early spring, before
the leaf buds open, chlorophylls are low. As soon as
buds open, and at the time of rapid increases in leaf
area and leaf weight per unit area, there is an abrupt
and substantial rise in both chlorophyll a and b. Although total chlorophyll concentrations (Fig. 5)
are consistently high throughout the summer period,
considerable fluctuation is apparent, as was noted
earlier by Schertz (1929) and Goodwin (1958).
For buds in early spring the ratio of chlorophyll
a to chlorophyll b (Fig. 6) is relatively low. After
leaf-blade expansion begins, the ratio increases rapidly to a high level, which persists throughout the
entire summer. The high summer levels (approximately 2.6) correspond to the average values for
green tissue of higher land plants given by Smith and
Benitez (1955).
Autumn 1971
INVESTIGATIONS OF LEAF PIGMENTS
J
MiJ
A
S O
N
S O
N
3
TTI
0
M
J
J
A
FIG. 6. Temporal changes in the ratio of chlorophyll
a to chlorophyll b in oak leaves.
At the onset of autumn coloration in October total
chlorophyll concentrations (Fig. 5) drop rapidly
while leaf weight per unit area drops only slightly
(Fig. 2). The ultimate fate of the chlorophylls destroyed in drying autumn leaves is as yet essentially
unknown. Because Seybold (1943) could not demonstrate products of high molecular weight in autumn
leaves he concluded that extensive cleavage of the
chlorophyll molecule into colorless fragments must
occur. During the period of most rapid chlorophyll
decline, traces of pheophorbide a and chlorophyllide
a can occasionally be detected, but one cannot be
certain that these are not artifacts of the extraction
or separation procedure. Pheophytin a, however, can
be detected in small but measurable quantities during rapid chlorophyll degradation, as well as throughout the entire winter in dry leaves on the tree and
in leaves overwintering on the forest floor (Fig. 4
and 5). It appears that a small fraction of chlorophyll a gets converted to pheophytin a and then is
stable enough to be present for long periods in dry
leaves and in upland soil organic matter layers (Gorham 1959, Gorham and Sanger 1964, 1967). Only
faint traces of pheophytin b are encountered during
the period of most rapid chlorophyll degradation.
Willstiitter and Stoll (1913) demonstrated that
colloidal solutions of chlorophyll in water could be
decomposed by CO2 and that chlorophyll a was more
rapidly converted to pheophytin a than chlorophyll
1079
b to pheophytin b. This apparent greater ease in
conversion of chlorophyll a to pheophytin a is suggested here in oak leaves, because only pheophytin
a is found in measurable amounts in dry leaves on
trees and in forest organic matter layers. Possibly
the slowness of reaction of chlorophyll b to pheophytin b allows all the chlorophyll b to be destroyed
before pheophytinization is able to preserve any of
it. However, because the preserved levels of pheophytin a are so very low in dried leaves compared to
the original chlorophyll concentration, it is also possible that levels of pheophytin b, even if preserved
in equal proportion to pheophytin a, would be too
low for detection of more than trace amounts by
the methods employed.
Studies involving the extraction and measurement
of chlorophyll under a number of experimental situations have revealed the presence of isomers of
chlorophyll a and b (a' and b') (Strain and Manning
1942, Michel-Wolwertz and Sironval 1965, Bacon
and Holden 1967). It is possible that chlorophyll
isomers appear during the most rapid breakdown of
pigment. Their chromatographic and spectral properties, however, differ very slightly from those of the
parent compounds, and the techniques employed in
this study may not have been sufficiently sensitive
to detect such subtle differences. Strain and Manning
(1942) indicated that chlorophyll d is more bluegreen than the parent chlorophyll a, and b' more yellow-green than chlorophyll b. At no time was there
any indication of a dual nature of the chlorophyll
spot, nor was any color change apparent in the chlorophyll a and b spots during any of the separations.
When autumn coloration begins, the ratio of chlorophyll a to chlorophyll b again becomes low (Fig.
6). Studies of Willstitter and Stoll (1918) revealed
no change in the ratio of chlorophyll a to chlorophyll
b in autumn leaves. The more rapid breakdown of
chlorophyll a as compared to chlorophyll b under a
variety of circumstances involving the decay of green
plant tissue has since been demonstrated, however
(Rudolf 1934, Nagel 1940, Seybold 1943, Egle 1944,
Jeffrey and Griffith 1947, Strain 1949, Wolf and Wolf
1955, Wolf 1956, Goodwin 1958), so it appears that
the earlier views of Willstitter and Stoll were wrong.
Egle (1944), from his studies on silage, concluded
that acids arising during fermentation constituted an
important factor in chlorophyll destruction, with
chlorophyll a less resistant than b. Similar observations on drying plant material suggested to him that
acids normally present in leaves are very important in
the destruction of chlorophyll. Goodwin (1958) noted
in autumn leaves, and the present study of oak leaves
confirms, that only when the chlorophylls had almost
disappeared was there a tendency for chlorophyll a
to be destroyed more quickly than chlorophyll b.
The rate of destruction of chlorophyll a relative to
Ecology, Vol. 52, No. 6
JON E. SANGER
1080
4
00.5I
lutein
of 0.34
0
0
3-"
U
0 2|
-rtiolaxanthi
oFIG 7Tmolneoxanthian
M J
.
6i0
I
Dn
9utei n
i
|
~~~~~~~~~~~tail
*
M
J JA
SO
N
D
J
F
M A
7. Temporal changes in xanthophylls per unit organic matter of oak leaves.
FIG.
frn si
a
NOm
S
v
r
rw
0
0
A
SON
D J
F MA
FIG. 9. Temporal changes in the ratio of P-carotene
to xanthophyll in oak leaves.
n>
j:OJ-
J
llqoq~
Z
0
IAqdO44uox
0
8 C
FIG. 8. Temporal changes in concentrations of carotenoids per unit organic matter of oak leaves.
b may reflect the rapidity of leaf desiccation of different species in autumn. Oak leaves in this study
tended to retain moisture much longer than those of
aspen and hazel, and as a result the rapid decline of
chlorophyll a relative to chlorophyll b was more
apparent in oak.
Like the chlorophylls, the major carotenoid components are all present in spring leaf buds, but in
relatively small amounts (Fig. 5, 7, and 8). After a
sharp rise as buds open, total carotenoid concentrations (Fig. 5) increase gradually to summer maxima
at the end of July. Neoxanthin reaches maximum
concentration in early summer followed slightly later
by violaxanthin (Fig. 7). Beta-carotene and lutein,
the two components most stable in autumn, do not
reach maximal concentrations until midsummer (Fig.
7 and 8).
The ratio of P-carotene (the only carotene observed in the tree leaves studied) to total xanthophyll reveals a rather sudden increase from 0.38 in
buds to 0.49 in early July (Fig. 9). Average values
of 0.45 are recorded for summer up until autumn
coloration.
Although variations in summer pigment levels are
apparent, concentrations remain high until the initiation of autumn coloration, when the level of carotenoids begins declining at the same time as the chlorophylls. As indicated previously, the generally
accepted theory has been that the chlorophylls disappear first, unmasking the carotenoids, which disappear only gradually in late autumn. The curves for
total chlorophyll and total carotenoid (Fig. 5) show,
however, that carotenoids, although retained a few
days longer in oak leaves than are chlorophylls, decline at the same rate. The major difference is in the
ultimate amount of the decline, with very small concentrations of chlorophyll being preserved as pheophytin a throughout late autumn and winter compared to much higher concentrations of the carotenoids lutein and P-carotene during the same period.
Violaxanthin and neoxanthin are the least stable
of the carotenoids (Fig. 7), with concentrations dropping in autumn at about the same time as the chlorophylls decline (see Fig. 4). Violaxanthin is the
least stable, disappearing 1-2 weeks before neoxanthin. In Goodwin's study the level of violaxanthin
remained in oak and plum leaves with comparative
constancy until the end of his sampling period in
late November. Thus broad generalizations cannot
be drawn for the fate of xanthophylls in autumn
leaves since differences between species and between
climates may have considerable influence on pigment
decomposition.
During the period of sharp decline in total carotenoid, an additional pigment spot was noted tailing
closely behind lutein, although distinctly separate
from it (Fig. 7). The pigment could be detected in
relatively small amounts for about 2 months during
autumn, but disappeared shortly after neoxanthin.
Positive identification was not made because of its
spectral peculiarities and inconsistencies. Further
study was limited by the very low concentrations of
Autumn 1971
INVESTIGATIONS OF LEAF PIGMENTS
this pigment. The most plausible explanation is that
the lutein tail was a mixture of esters of violaxanthin
and lutein. The pigments within the spot were epiphasic initially, but on saponification became for the
most part hypophasic. These pigments are most
likely those noted by Willstitter in 1918, the autumn
carotenes of Tswett (1911), and autumn xanthophylls of Kuhn and Brockmann (1932) who, according to Goodwin (1958), correctly identified them
as xanthophyll esters. No pigments with spectra resembling lutein-5:6-epoxide, which Goodwin detected
in small quantities in autumn leaves of oak, plum,
and sycamore, could be detected in any leaves from
this study with techniques similar to those of Goodwin.
Beta-carotene and lutein are the most stable carotenoids of those examined in this study (Fig. 7 and
8). This was to be expected, because these are the
two components found commonly in the organic
layers of woodland soils (Sanger 1971). Both pigments decline rapidly during the autumn coloration
period, but once oak leaves dry on the tree the rate
of pigment loss becomes considerably less. Concentrations of 5-carotene drop to much lower levels than
lutein in autumn, but nevertheless ,3-carotene remains
preserved in measurable amounts throughout the entire winter in leaves dry on the tree, declining gradually to quite low levels by spring. Slightly higher
concentrations of 5-carotene are noted in the spring
for leaves that overwintered on the soil surface beneath the winter snows, which that year accumulated
3-4 ft. Lutein, the dominant carotenoid preserved
in dried leaves, declines slowly throughout the winter in dry leaves on the trees, but remains on the
average about 7 times as abundant as 5-carotene.
Like 3-carotene, lutein concentrations are much
greater in leaves overwintering on the soil surface.
The important point to note is that when oak leaves
drop from the trees in the desiccated condition,
whether in spring or in autumn, they contain small
but measurable amounts of lutein, ,-carotene, and
pheophytin a. Leaves dropping in autumn and becoming incorporated in the forest litter, frozen and
covered with snow, contain pigment levels in spring
only slightly lower than dry leaves on trees in late
autumn. This is probably a reflection of protection
from light and temperature extremes throughout the
winter. The higher pigment levels of oak leaves falling in autumn undoubtedly makes them greater contributors to pigments in terrestrial soils or aquatic
sediments than leaves falling in spring.
The ratio of ,8-carotene to xanthophyll (Fig. 9)
begins falling at the initiation of autumn coloration
from the average summer value of 0.45 to a lateautumn value of about 0.15. It remains about the
same throughout the winter in leaves on trees and
leaves on the forest floor. The autumn change dem-
1081
onstrates the relatively greater stability of xanthophyll (more specifically, the lutein component) during the period of pigment breakdown.
In leaves overwintering on the trees the rate of
pigment decomposition increases slightly just before
leaf drop in mid- to late April. During this time the
leaves are exposed to warmer temperatures and continued wetting and drying from spring rains and
morning dews. Also by this time the days have
lengthened considerably, and leaves are subjected to
long periods of sunlight. Probably the combined
effects of these factors cause more rapid pigment
decomposition just before leaves fall. Leaves lying
on the forest floor have by this time become fairly
tightly matted to one another and to older, more
decayed leaves of the upper soil organic horizon.
This condition is undoubtedly more conducive to
pigment preservation and is considered in greater
detail elsewhere (Sanger 1971).
During the summer months lutein represents about
35% of the total carotenoid of oak leaves, ,8-carotene about 30%, violaxanthin about 20%, and
neoxanthin about 12% (Fig. 10). These values re20M
J
J
A
S O
N
D
J
F
M A
Lutein tail
80
6040
Lutein
20-
C.
z
UJ
CX
____a_
-carotene
20C
2d
20
Violaxanthin
~~~~~~Neoxanthin]
A
J AAS S 0 NN_ D J F M A
M J
10. Temporal changes in the percentage of total
carotenoid components of oak leaves.
FIG.
Ecology, Vol. 52, No. 6
JON E. SANGER
1082
5
0 4030-
leaf area
.
-
4<
2
401
0<
(
~~~~~~~~~30
LUV-
z
20<
2-
Z 20L
0 1t.
0,
0 10-
10<&Uv0
I
<CM
i J
A
S O
N
D
J
F
M
FIG. 11. Temporal changes in concentrations of anthocyanin per unit organic matter of oak leaves.
main fairly constant from spring buds until the
initiation of the autumn coloration period, when
violaxanthin and neoxanthin disappear, leaving only
,3-carotene and lutein. The presence of the pigments
in the lutein tail can be noted during the peak of
autumn coloration, at which time they may make
up as much as 13 % of the total carotenoid. Eventually in late autumn lutein and ,3-carotene are the
only remaining carotenoids, with ,3-carotene averaging about 15% and lutein the remaining 85%
throughout much of the winter and spring.
At the initiation of autumn coloration, as soon as
chlorophyll begins to decompose, anthocyanin forms
(Fig. 11). At the time of maximum anthocyanin
concentration, levels of chlorophyll have dropped
to 20% of the average summer value. The concentration of anthocyanin rises very abruptly to a maximum and thereafter throughout the winter declines
at a slow rate. Only trace amounts remain in April
when the leaves fall from the trees. Unlike lutein
and ,3-carotene, anthocyanin is not present to any
great extent in the spring in either dry leaves on the
tree or in dry leaves overwintering on the forest floor.
Leaves on the trees remain relatively high in anthocyanin during much of the winter, probably because
they are kept continuously cold and dry. It is likely
that in spring, when temperatures increase and when
periods of wetting and drying become frequent, the
water-soluble anthocyanins are even more vulnerable
to destruction than the plastid pigments, especially
more so than lutein. The lack of anthocyanin in
dry leaves in spring is not entirely unexpected because, as previously indicated, no trace of the pigment has been found in soils, lake muds, or peats.
Hazel leaves
Changes in leaf area and in the dry, ash-free
weight per leaf and per unit area of hazel leaf tissue
throughout the sampling period are presented in Fig.
12 and 13. Leaf weight per unit area remains essentially unchanged from young leaves through leaf
desiccation and fall in autumn despite the fact that
leaf area and organic matter per leaf do not reach
J
M
A
J
A
0
S
FIG. 12. Temporal changes in dry ash-free weight per
unit area and average leaf area of hazel leaves.
UL.
100
-
_75
50
?
/
0
25-
J
J
M
0
S
A
FIG. 13. Temporal changes in organic matter per leaf
of hazel.
otal
carotenoid
18-
*900
16\
d
/
c14-
700 0
0
total
812-
\t,
chlorophyll
500:.
Vo8
08
0
0
-~~~~~~3009
Z5 6.
U4
2 i100
M
J
J
A
S
0
FIG. 14. Temporal changes in concentrations of total
chlorophyll and carotenoid per unit organic matter of
hazel leaves.
high levels until mid-July. This is contrary to the
case in oak leaves, where leaf weight per unit area
increased from low levels in buds to high summer
levels by mid-July, paralleling the seasonal changes
in absolute leaf organic matter and leaf area.
Concentrations of total chlorophylls and carotenoids increase gradually, not reaching summer maxima until the period from mid-June to mid-July (Fig.
14). As in oak leaves, all the plastid pigments inves-
Autumn 1971
INVESTIGATIONS
tigated were present in leaf buds, but in relatively
small amounts. Total pigments do not remain at consistently high concentrations throughout most of the
summer as in oak, but are high for only a short time
during midsummer. Chlorophylls a and b rise slowly,
reaching highest summer concentrations in mid-June
(Fig. 15), about 1 month before maximum leaf area
is attained (Fig. 12). Concentrations then begin to
fall at a nearly uniform rate until the total disappearance of all pigments in dry autumn leaves.
Violaxanthin, unlike the other carotenoids, rises
abruptly in late May, and for a short time exceeds
the concentration of lutein (Fig. 16). It then begins
a continual slow decline which is essentially unbroken until total decomposition in mid-October.
Beta-carotene, lutein, and neoxanthin display rates
of increase and decrease similar to the chlorophylls
and reach maximum summer concentrations in midJuly (Fig. 16 and 17).
Instead of dropping sharply during autumn coloration, pigment concentrations begin to decline
gradually in August. The chlorophylls and carotenoids decline at essentially equal rates, reaching
~120 10
xanthophyll
08
0I-
L)
6.
<2
M
J
J
A
S
FIG. 17. Temporal changes
in cnenratio of
carot-ne
0
U
a
0.5
0.
enoid
xhperl uitognimatro hazel leaves.
3
a
hloro.
/~~~~~~<~
0 O
1083
OF LEAF PIGMENTS
0
o 5000
0.5
M
J
J
A
S
0
18. Temporal changes in the ratio ofclcarotene
antocanhrophyll
uni
n hazel leaves.
FIG.
3300 chioro.
0
..
J
A
S
0
FIG. 15. Temporal changes in chlorophylls per unit
organic matter of hazel leaves.
M
J
'4
FIG. 16.Temporalchanxanthn
FIg 1. temporhazlchavesi
organic matter of hazel leaves.
pr u
zero by the end of October (Fig. 14). In hazel leaves
lutein is preserved only slightly longer than violaxanthin and neoxanthin, which are lost about 10 days
before the last traces of lutein disappear (Fig. 16).
Only faint traces of pheophytin a are detectable
during the period of most rapid chlorophyll decline,
and no pigments are found after the leaves dry.
Hazel leaves become much more thin and brittle
when dry than do oak leaves, and all leaves fall from
the bushes in autumn free from pigment.
A comparison of Fig. 14 with Fig. 5 reveals that
hazel leaves are about twice as rich in both carotenoids and chlorophylls as oak leaves. This may be
an adaptation for photosynthesis under shade con-
Ecology, Vol. 52, No. 6
JON E. SANGER
1084
M
J
J
A
F
S
Z E^40 <0
0 ? 20j
20
Lutein
0
C
UM
j
A
0
S
FIG.21. Temporal changes in concentrations of anthocyanin per unit organic matter of hazel leaves.
20
a-carotene
Z
7
1
nit area
.
-308
U
cL<:
6 -...*
.
20Z
....
leaf area
LI
<
10<
20
Violaxanth in
M
i
A
i
0
S
FIG. 22. Temporal changes in dry ash-free weight per
unit area and average leaf area of aspen leaves.
?
M
J
J
A
S
20. Temporal changes in the percentage of total
carotenoid components of hazel leaves.
125 -
FIG.
ditions, since hazel is almost always in shady situations and locally is the dominant component of
the shrub synusia of the oak forest. For comparable
periods, however, the ratio of ,3-carotene to xanthophyll (Fig. 18) and the summer ratio of chlorophyll
a to chlorophyll b (Fig. 19) are much the same in
the two species. In autumn the ratio of chlorophyll a
to chlorophyll b does not show evidence of a decline
as in oak, so that chlorophyll b in hazel is not preserved relative to chlorophyll a. This may be a consequence of the gradual autumn decline of chlorophyll, as opposed to the very rapid decline in oak.
The carotenoid components of hazel leaves expressed as percentages of total carotenoid are shown
in Fig. 20. The summer values are almost identical
with the values for oak, with lutein representing
about 35% of the total summer carotenoid, 5-carotene from 25% to 35%, violaxanthin from about
15% to 20%, and neoxanthin from 10% to nearly
20%. Percentages of violaxanthin in hazel are somewhat higher at the beginning of the season, reflecting
the rapid development of this pigment in hazel leaves
(Fig. 16).
No traces of "esterified xanthophylls" or other
autumn pigments could be found in hazel. The "lutein tail" noted for oak could not be detected, and
no evidence was found for the presence of lutein5: 6-epoxide as noted by Goodwin (1958). Its absence may be associated with the more gradual decline of the hazel pigments in autumn, with their
UL.
75
0
all
25M
J
J
A
S
0
23. Temporal changes in organic matter per leaf
of aspen.
FIG.
subsequent complete disappearance when the leaves
dry and turn brown.
As in the case of oak, hazel leaves do develop
anthocyanin pigment in relatively large quantities
for a short period in autumn (Fig. 21). Anthocyanin
reaches maximum concentration after chlorophylls
and carotenoids decline to less than 40% of their
summer maxima. Anthocyanin disappears about the
same time as lutein, a few days later than the other
plastid pigments. Its spectrum is identical to that
recorded for the anthocyanin of oak, which in turn
is very similar to that of cyanidin.
Aspen leaves
The time required for development of maximum
aspen leaf area, and the values of organic matter per
leaf and per unit leaf area, are presented in Fig. 22
and 23. Both leaf area and leaf weight increase
rapidly after buds break in mid-May to relatively high
levels by late May. As in hazel, leaf weight per unit
INVESTIGATIONS
Autumn 1971
*
012
.
total carotenoid
OF LEAF PIGMENTS
a600:*
1085
viol!~axanthin
oi l
ll
lutein
010.0
0
0
0
AS
total chlorophyll
4000
0
zU4-
a
2000IL
0W
0
0
. .....
,;'t
neoxanthin
1
01
i
M
j
A
S
0
FIG. 24. Temporal changes in chlorophyll and carrot
enoid per unit organic matter of aspen leaves.
a
8 -:
033
0
0
S
A
J
J
M
FIG. 26. Temporal changes in xanthophylls per unit
chloro.r
organic matter of aspen leaves.
0
S200anitoro.
.otpro
b
88
~100
:Xanthophyll
;
0.
o 4
U
C
i-carotene
/
0
S
A
M
i
J
FIG. 25. Temporal changes in chlorophylls per unit
organic matter of aspen leaves.
area remains consistently high throughout the entire
growth period, and it drops only slightly during
autumn desiccation. Oak and aspen are similar in
leaf weight per unit area, with average summer
values of about 7.0 mg/cm2, whereas hazel shows
much lower values (near 3.0 mg/cm2).
Although the sampling intervals are less frequent
for aspen, the pigment concentrations and curves
show similarities to both oak and hazel. All pigments
investigated were present in low concentrations in
leaf buds (Fig. 24). Total chlorophylls and carotenoids rise very sharply after the buds open, reaching
high concentrations in late May when leaves have
nearly attained their maximum area. Chlorophyll a
and b remain consistently high throughout the summer (Fig. 25). The carotenoids reach maximal concentrations in mid- to late May, and total carotenoid
shows a slight decline thereafter through the summer. Violaxanthin, as in hazel, rises very sharply in
spring and exceeds the concentration of lutein during much of the early part of summer (Fig. 26).
Violaxanthin and neoxanthin concentrations decline
slowly from late May to early September. Beta-carotene and lutein do not show this tendency clearly
(Fig. 26 and 27).
The average summer ratio of chlorophyll a to chlo-
0
S
M
J
J
A
FIG. 27. Temporal changes in concentrations of carot-
enoids per unit organic matter of aspen leaves.
15/
0.
M
J
J
A
S
0
FIG. 28. Temporal changes in the ratio of chlorophyll
a to chlorophyll b in aspen leaves.
rophyll b is about 1.9 (Fig. 28) compared to values
greater than 2.5 for oak and hazel. The general
character of the curve is similar to oak. As in oak
the ratios are relatively low in buds, rising to a summer level that is maintained more uniformly than in
oak.
The ratio of ,3-carotene to xanthophyll does not
resemble closely either that of hazel or oak (Fig.
29). Average summer values are about 0.38 in aspen,
compared to values around 0.45 for oak and hazel.
Values for aspen are at the highest recorded level in
buds and drop substantially when the young leaves
JON E. SANGER
1086
0.00.-
iU
0.
o0 0.1
M
J
J
A
S
0
FIG. 29. Temporal changes in the ratio ofa-carotene
to xanthophyll in aspen leaves.
JrJ
M
A
S
0
4
20
Lutein
O-
20
dry adflfrmtetes(i.2)Th-carotene
20Violaxant
C
in
20
M
J
A
S
0
FIG. 30. Temporal changes in the percentage of total
carotenoid components of aspen leaves.
begin to expand. This is essentially opposite to the
trend in oak and hazel.
Like hazel, and unlike oak, the level of total pigments drops to zero in late autumn when the leaves
dry and fall from the trees (Fig. 24). The level of
violaxanthin begins a rapid decline in mid-September that noticeably precedes the period of rapid decline of the other carotenoid components (Fig. 26).
Neoxanthin begins a more rapid fall at the same time
the chlorophylls begin to decline in late September,
some weeks after the initiation of a rapid fall in
violaxanthin.
As in oak, lutein and P3-caroteneappear to be the
most stable of the plastid pigments (Fig. 26, 27, and
30). Lutein in particular remains in high concentrations until the latter part of October, when it begins
to decline very rapidly as leaves dry and fall from
Ecology, Vol. 52, No. 6
the trees. The sharp drop in the level of lutein takes
place about a month later than the sharp drop in
chlorophylls a and b, violaxanthin, and neoxanthin.
Although 5-carotene begins a slight decline in late
September, it is maintained in relatively high concentrations until late October when it falls rapidly
along with lutein. Faint traces of pheophytin a are
occasionally detectable during the period of most
rapid chlorophyll decline. No traces of the lutein tail
found in oak leaves were noted, and no evidence was
found for lutein-5:6-epoxide. The absence of autumn
xanthophylls may result, as in hazel, from anatomical or chemical leaf conditions less favorable to pigment preservation than those of oak.
The ratio of chlorophyll a to chlorophyll b falls
from the summer values during late autumn when
chlorophyll concentrations drop to low levels (Fig.
28). In late autumn there is a slightly greater preservation of chlorophyll b than of chlorophyll a, a
tendency not noted at all in hazel but very evident
in oak.
The ratio of P-carotene to xanthophyll does not
decline in autumn (Fig. 29) as in hazel and oak.
This may be because of the late, sudden and extreme
drop of both ,-carotene and lutein.
The carotenoid pigment components disappear
from aspen leaves at roughly the same time in late
autumn as they do in hazel (Fig. 30). Summer values
of about 30-35% for lutein, 25% for 5-carotene, 2535% for violaxanthin, and 10-12% for neoxanthin
are similar to those for both oak and hazel.
Chlorophyll content and site productivity
In recent years several studies have suggested determination of chlorophyll load per unit of landscape area as an index to net primary productivity
of terrestrial communities (Odum 1959, Bray 1960,
1962, Brougham 1960, Whittaker and Garfine 1962,
Ovington and Lawrence 1967, Sanger, unpublished
data). The present studies of pigment levels throughout the season have demonstrated two phenomena
which may influence correlations between productivity and chlorophyll load. First, concentrations of
chlorophyll per unit leaf area exhibit distinct fluctuations about an average high summer value. This is
especially true for oak. Second, not all plants carry
maximum pigment loads throughout the entire summer per unit organic matter and per unit leaf area
(see also Ovington and Lawrence 1967). This is also
true if pigments are expressed on an absolute basis
(per leaf). Because leaf number remains essentially
unchanged throughout the canopy during summer,
any change in absolute pigment concentration involves a pigment change per unit area of landscape.
In hazel leaves, increases in absolute total chlorophyll and carotenoid (Fig. 31) parallel the relatively
slow increases in absolute values of organic matter
UI.
carotenoid
.*..*total
0.75-
1087
OF LEAF PIGMENTS
INVESTIGATIONS
Autumn 1971
o2u
t tal chlorophyll
W 0.5
..a
~~~~~~~~~~~
0
*toalchlorophyll
D 0.25 -.
M
J
J
I
.
A
0
S
FIG. 31. Temporal changes in total chlorophyll and
carotenoid per leaf of hazel.
J
-M
J
A
S
0
FIG. 3 3. Temporal changes in total chlorophyll
carotenoid
. .1.25.
total
carotenoid
00
~0.75
0
leaves from before the breaking of buds in spring
until leaf desiccation and fall. By late autumn hazel
and aspen leaves dry and drop to the ground essentially
0
u 0.2
o0i
?
U
total chlorophyll
J
J
A
S O
N
D
J
and
per leaf of aspen.
F
M
A
FIG. 32. Temporal changes in total chlorophyll and
carotenoid per leaf of oak.
(Fig. 13) throughout spring and early summer to
highest levels in mid-July. Thereafter organic matter
remains relatively constant while pigments decline
steadily to zero in late autumn. In oak total pigments
rise steadily from low levels in mid-May to high
levels in July (Fig. 32), as does organic matter (Fig.
3). Pigment concentrations then rise more slowly
until autumn, declining rapidly in late September,
when autumn coloration begins. Absolute values for
pigments and organic matter of aspen leaves show
a very rapid initial rise to high levels in late May
(Fig. 33 and 23). Thereafter aspen resembles oak
with a more gradual rise throughout the summer to
highest concentrations just before the initiation of
the autumn decline. These data suggest that chlorophyll content per unit area of landscape is likely to
vary considerably, depending upon the sampling
time. Therefore, before any truly meaningful studies
of productivity in relation to chlorophyll content are
made, the investigator should be aware of the time
of year and the duration and magnitude of fluctuation of maximum chlorophyll load for the major
components of the plant community. For some communities the period during which reliable maxima
can be estimated may be relatively short, and this
period may well differ in different communities.
CONCLUSIONS
Comparisons have been made of the changes in
pigment concentrations of oak, hazel, and aspen
devoid
of pigment.
Pin-oak
leaves,
of which
a
large percentage remain attached to the tree throughout the entire winter, retain much of their lutein
a m-carotene
moderate percentage of their
(u25%la
aspea
small amount of chlorophyll a in the
(5a%),n
form of pheophytin a (0. 1%) in leaves remaining
on the trees well past the period of major autumn
coloration. The same three pigments are still present
in small amounts when these leaves fall in spring.
Such findings indicate that pigments in the allochthonous organic matter reaching the sediments of
lakes and ponds may be derived from a restricted
number of plants among the upland flora. This may
be an important consideration for interpreting sedimentary pigments in lakes which derive much of
their organic matter from the surrounding environment, especially from falling leaves in autumn or
from material that has weathered on the soil surface.
In most situations chlorophyll a is more rapidly
decomposed than chlorophyll b. The ratio of chlorophyll a to b is relatively low in buds and very young
leaves, remains high throughout the summer months,
and drops to low values in autumn as the chlorophylls decompose. In hazel leaves, however, where
the autumn decomposition of chlorophyll is relatively
gradual and carotenoids and chlorophylls disappear simultaneously, there is no evidence for relatively
greater autumn stability of chlorophyll b.
During the summer both carotenoids and chlorophylls differ among the three tree species in maximal concentrations as well as in the timing and
duration of maximum levels per unit organic matter
and per unit area. Such differences may need to be
considered in the use of pigment load per unit of
landscape area as a productivity index. Aspen and
oak are nearly equal in pigment concentration, but
hazel, growing entirely in a shaded environment, has
about twice the pigment concentration of oak and
1088
JON E. SANGER
Ecology, Vol. 52, No. 6
LITERATURE CITED
aspen per unit organic matter. Duration of maximum
Bacon, M. R., and M. Holden. 1967. Changes in chloropigment level is shortest for hazel.
phylls resulting from various chemical and physical
Although differences in absolute summer values
treatments of leaves and leaf extracts. Phytochemistry
are apparent, carotenoid components expressed as a
6: 193-210.
percentage of total carotenoid are remarkably con- Bray, J. R. 1960. The chlorophyll content of some native
sistent in the three species. In autumn the carotand managed plant communities in central Minnesota.
Can. J. Bot. 38: 313-333.
enoids violaxanthin and neoxanthin are more sus. 1962. The primary productivity of vegetation in
ceptible to decomposition than P-carotene and especentral Minnesota, U.S.A. and its relationship to chlocially lutein. This is most easily demonstrated in oak
rophyll content and albedo, p. 102-116. In Die Stoffleaves, where apparently anatomical or chemical conproduktion der Pflanzendecke. Gustave Fisher Verlag,
ditions in the leaves favor pigment preservation to
Stuttgart.
a greater degree than in hazel and aspen. The changes Brougham, R. W. 1960. The relationship between the
critical leaf area, total chlorophyll content, and maxin ratio of ,-carotene to xanthophyll show little simimum growth-rate of some pasture and crop plants.
ilarity among the species investigated. Curves for
Ann. Bot. 24: 463-474.
hazel and aspen leaves from spring to autumn are Colman, B., and W. Vishniac. 1964. Separation of chloroplast pigments on thin layers of sucrose. Biochim.
nearly opposite: hazel leaves show ratios high in
Biophys. Acta 82: 616-618.
summer and low in spring and autumn, whereas
K. 1944. Untersuchungen fiber die Resistenz der
Egle,
aspen leaves are low in summer and higher in spring
Plastidenfarbstoffe.Bot. Arch. 44: 93-148.
and autumn. Lutein and P-carotene remain through- Geissman, T. A. 1955. Anthocyanins, chalcones, aurones,
out the winter in dry oak leaves on the trees and in
flavones and related water-soluble plant pigments,
p. 450-498. In K. Paech and M. V. Tracey [ed.] Modoak leaves lying on the forest floor. (This indicates
ern methods of plant analysis. Vol. III. Springer-Verthe ratio n-carotene to xanthophyll in winter is in
lag,
Berlin.
fact expressed as the ratio n-carotene to lutein with
Goodwin, T. W. 1952. The comparative biochemistry of
the average value of the latter remaining essentially
the carotenoids. Chapman and Hall, London. 356 p.
unchanged throughout the winter.)
. 1958. Studies in carotenogenesis. The changes
in carotenoid and chlorophyll pigments in the leaves
An "autumn xanthophyll" (the lutein tail in Fig.
of deciduous trees during autumn necrosis. Biochem.
7 and 10) appeared temporarily in relatively small
J. 68: 503-511.
quantities in oak leaves but was not detected during
Gorham, E. 1959. Chlorophyll derivatives in woodland
the autumn coloration of hazel or aspen leaves. This
soils. Soil Sci. 87: 258-261.
was not the lutein-5:6-epoxide detected by Goodwin Gorham, E., and J. Sanger. 1964. Chlorophyll derivatives in woodland, swamp, and pond soils of Cedar
(1958), which was not found during the autumn
Creek Natural History Area, Minnesota, U.S.A., p. 1period in any of the three species examined.
12. In Y. Miyake and T. Koyama [ed.] Recent reAnthocyanin appears in both oak and hazel leaves
searches in the fields of hydrosphere, atmosphere and
during autumn. In oak, concentrations rise sharply
nuclear geochemistry. Maruzen Co., Tokyo.
. 1967. Plant pigments in woodland soils. Ecology
in early October, decline slowly throughout the win48: 306-308.
ter in leaves dry on the tree, and drop to only trace
amounts when the leaves fall from the trees in early Harborne, J. B. 1965. Flavonoids: Distribution and contribution to plant color, p. 247-278. In T. W. Goodwin
spring. No anthocyanin is present in the spring in
[ed.] Chemistry and biochemistry of plant pigments.
leaves dropping in late autumn and overwintering
Academic Press, New York.
on the forest floor. In hazel, anthocyanin begins Holden, M. 1962. Separation by paper chromatography
of chlorophylls a and b and some of their breakdown
forming in early September and reaches maximum
products.
Biochim. Biophys. Acta 56: 378-379.
concentration in early October. It declines to zero
Holt, A. S., and E. E. Jacobs. 1954. Spectroscopy of
when leaves dry and fall to the ground in late Ocplant pigments. I. Ethyl chlorophyllides A and B and
tober.
their pheophorbides. Amer. J. Bot. 41: 710-717.
Jeffrey, R. N., and R. B. Griffith. 1947. Changes in the
ACKNOWLEDGMENTS
chlorophyll and carotene contents of curing burley toI am indebted to Professor Eville Gorham for his adbacco cut at different stages of maturity. Plant Physiol.
vice and guidance throughout the research and for his
22: 34-41.
valuable suggestions on preparing the manuscript. Grate- Joslyn, M. A., and G. Mackinney. 1938. The rate of
ful acknowledgment is made to the National Science
conversion of chlorophyll to pheophytin. J. Amer.
Foundation for financial assistance (through Grants
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