THE EFFECT OF SUNLIGHT ON THE DECOMPOSITION

OF PIN OAK LEAVES

Tiffany W., Ally R., Emma M., and Piya S.

 April 14th, 2024
BIOL 271.01

Abstract

Leaf packs are a vital part of stream ecology; they provide shelter and nutrients for

macroinvertebrates who then break the packs down into fine particulate matter that can be used

as an energy source for other consumers. Leaf decomposition is heavily influenced by UV

radiation. More radiation will cause leaves to decompose faster. We hypothesized that leaf packs

placed in sunny areas of a stream would decompose at a quicker rate than those placed in shade,

and they would attract more macroinvertebrates who aggregate in sunny areas to avoid predation.

We created artificial leaf packs and placed some in sunny areas and some in shaded areas of the

same stream. We observed the decomposition rates and macroinvertebrate abundances of these

leaf packs over time. Our results proved to be inconclusive due to the inconclusive results from

our linear regression and the ANOVA test that proved the final sunny leaf packs to have a

significantly lower ash-free dry weight than the rest of the leaf packs. This study provides new

knowledge on the effects of photodegradation on Pin Oak (Quercus palustris) leaves and may be

used to conduct further research.

Introduction

Leaves that fall into streams and are caught on structures within a stream can sometimes

form bunches of detritus, which are referred to as leaf packs (Benfield 1996). These leaf packs

temporarily become home to a variety of different macroinvertebrates that feed on and break

down the detritus (Benfield 1996). Eventually, these leaves are broken down into fine-particulate

matter, which is then used as an energy source for various consumers throughout the stream

(Benfield 1996). Sunlight is an important factor in the rate of decomposition of leaves. Visible
wavelengths of the sun’s rays convert inorganic carbon into organic forms through

photosynthesis, and ultraviolet light may disintegrate organic matter through photodegradation

(Rentala et al. 2021). Therefore, leaf packs gathered in areas of a stream that receive more

sunlight than others will tend to break down at a quicker rate than those gathered in shaded areas.

Previous research finds that ultraviolet radiation is a significant abiotic factor that plays

an important role in the turnover of carbon and decomposition rates of detritus (Austin and

Vivanco 2006). They found that by using an artificial sunlight blocker, decomposition rates of

litter decrease by up to 60% when not exposed to ultraviolet radiation (Austin and Vivanco

2006). They also found that the removal of organisms or limiting soil resources did not affect

decomposition rates, photodegradation was the sole influence over decomposition rates of leaf

litter (Austin and Vivanco 2006). Other studies on macroinvertebrates have found that most

species prefer sunny over shaded areas. Presumably, this is because of the more abundant algal

community that arises from increased photosynthetic opportunity (Wellnitz & Ward 1998).

Macroinvertebrates may also aggregate in sunny areas to avoid fish, who are more vulnerable to

predation in sunny areas. (Helfman 1981).

Our study aimed to determine what effect sunlight has on Pin Oak (Quercus palustris)

leaves’ decomposition rates and on macroinvertebrate habitation. We hypothesized that leaf

packs placed in a sunny part of the steam would exhibit quicker decomposition rates than those

placed in shade and attract more macroinvertebrates. We speculated that leaf packs placed in the

sun would have a lower percentage of organic matter at the end of the experiment than those

placed in the shade, due to the effects of photodegradation, and increased breakdown by

shredders attracted by the sunlight. Our null hypothesis was that sunlight would have no impact

on the decomposition rates or macroinvertebrate colonization rates of our leaf packs. We sought
to expand on existing knowledge of the decomposition rates of Pin Oak leaves since there is not

currently a lot of research on the topic.

Methods

Study Site

The study site we chose was the St. John Dam site (38.18552 N, 76.42679 W), located on

the campus of St. Mary’s College of Maryland (47645 College Dr, St Mary's City, MD 20686).

This site was a first-order stream located roughly 250 meters into a wooded area. The stream was

surrounded by a variety of upper-story trees (such as Pin Oaks, Sweet Gums, and maple trees),

creating shaded areas, while still having areas that receive decent sunlight throughout the day.

The depth of the stream was roughly sixty centimeters in the deepest parts and thirty centimeters

in its’ shallower areas. The width varies, but on average it was a little over a meter wide. The

salinity of this stream was usually close to zero, with higher deviations from this average

downstream. The temperature tended to be uniform throughout, with slight variations in different

parts of the stream. The dissolved oxygen in this stream was typically higher in the upstream

area due to the abundance of riffles, whereas the downstream area consists of more pools.

Study Species

Quercus palustris (Pin Oak) is a member of the Black Oak group. They are distributed

throughout most of North America, but primarily they are found in moist lowlands like St.

Mary’s College of Maryland. We identified the leaves as being glossy, between three to six

inches long, having five to seven lobes, and wide U-shaped sinuses (Gilman and Watson 2015).

The major chemical components of the Pin Oak leaves are lignin (~25%), cellulose (~45%), and

hemicellulose (~20%) (Le Floch et al. 2015). We picked Pin Oak leaves that had fallen in the

North Woods of St. Mary’s College of Maryland's campus. We chose this tree species due to its
wide availability and ease of identification. We used Pin Oak leaves to create the leaf packs used

in our study.

We looked at various macroinvertebrate species that inhabited our leaf packs during the

experiment. Some species of macroinvertebrates, notably shredders, feed on leaf detritus that has

fallen into stream ecosystems. They break large, coarse particulate matter down into finer

particles while decreasing the total organic matter of the leaves. Some species we primarily

expected to find were caddisflies (Trichoptera), craneflies (Tipuloidea), and stoneflies

(Plecoptera), all of these being shredders (Anderson and Sedell, 1979).

Experimental Design

Pin Oak leaves were collected from the North Woods of the St. Mary's College campus.

We chose leaves that were full, with little to no holes in them. They were then dried at 70° C in

the drying ovens for 24 hours. With a top-loading balance, the leaves were weighed in

approximately 10-gram groups and then placed in warm water to soften them slightly to prevent

breakage, then they were placed inside the mesh bags. The bags used were made out of nylon

with 3mm mesh openings. We recorded the dry weights of each leaf pack and labeled them

numerically. Repeating these methods, 40 leaf packs were created.

We then transported the leaf packs to the study site. We placed them in two locations of

the stream, one that gets sunlight for most of the day and one that had artificially created shade

we made using a trash bag. We created the shaded area close to the sunny area to minimize

differences in water quality measurements. The packs were placed into the ground, in close

proximity to others of the same treatment group, using stakes.

To account for loss due to handling and to create a control to compare the treated leaf

packs against, at week 0 we retrieved 5 leaf packs. We placed a D-frame kick net downstream
from the leaf packs and removed the stakes. Each leaf pack was collected into the kick net and

then transferred to a plastic gallon Zip-Lock bag. Each Ziplock bag contained one leaf pack to

avoid cross-contamination with other packs. Back in the lab, the leaves were carefully removed

from their bags and placed in a pan with tap water to rinse off the silt and debris. The

invertebrates were counted and stored for later identification. We then placed the leaves into an

aluminum foil boat to be dried overnight (24 hours) and collected the new dry weight.

After the leaf packs were weighed, we placed the dried leaves into new Zip-Lock bags.

The leaves were then crushed into tiny fragments, and using a spatula, a subsample was placed

into a pre-weighed crucible until ½ to ⅔ full. We then weighed the crucible and subsample to

obtain the dry weight of the subsample (by subtracting the crucible weight from the

crucible+subsample weight). This was repeated for each leaf pack collected. The teaching

assistants burned the crucibles and the subsamples at 500° C for 12 hours in the muffle furnace

(Model 62700, Dubuque, Iowa, USA). After 12 hours, the crucibles were cooled to 105° C, and

then transferred into a desiccator. We weighed our crucibles and ash to obtain the ash weight of

each subsample. The ash was then discarded, and crucibles were again burned at 500° C for 12

hours to clean them. These leaf pack processing methods were then repeated when we removed 5

leaf packs from each treatment group at week 3 and at week 5.

Data Collection

We recorded the water quality measurements of our study site weekly using a YSI water

quality probe with sensors and a data logger (Model 85, Yellow Spring Instruments, Yellow

Spring, Ohio). We kept track of the salinity, temperature, and dissolved oxygen percentage. The

stream current velocity was measured with a velocity meter (Marsh-McBurney Flow-mate,
Model 2000, Frederick, Maryland). Any macroinvertebrates identified in the leaf packs were

counted and then stored in vials with ethanol for identification.

Statistical Analysis

We used our samples' ash and dry weights to calculate the percentage of organic matter

and the ash-free dry weights (AFDW) of our samples. The organic matter percentage was

calculated by subtracting the subsample ash weight from the subsample dry weight and dividing

this number by the dry weight of the subsample. We did this because the ash weight is the mass

of inorganic matter, so when subtracting this from the dry weight, we may find the percent of

organic matter that was present in the sample. The AFDW was then calculated by multiplying

the percent organic matter by the dry weight of the entire leaf pack. We conducted an ANOVA

test to determine whether there was a difference in the AFDWs between each treatment

(sun/shade and time). Then, we conducted a Tukey post hoc test to determine which treatments

had significantly different AFDWs. We compared the average AFDW of each sample using a bar

graph. Then, we also calculated the mean organic matter percentage for each treatment and

conducted a linear regression to examine the loss of organic matter over time for each treatment.

We created a graph comparing the total number of macroinvertebrates counted from the sun leaf

packs and the shade leaf packs and conducted a t-test to determine whether there was a

significant difference between the numbers of macroinvertebrates found in each treatment group.

Graphs were created for each water quality measurement as well to examine whether they stayed

consistent throughout the experiment.

Results

Throughout this study, we found that the water quality measurements taken weekly were

consistent with some slight variations (Figure 1). Dissolved oxygen displayed the most variation

with the lowest being 79.2% and the highest being 259.8%. The temperature stayed consistent,

only varying by 2-3°C. Salinity began high at 5ppt, but then dropped to 0ppt and remained at that

level (Figure 1).

Figure 1. [MOU1] Water quality measurements taken from St. John’s Creek (38.18552 N,

76.42679 W) weekly throughout the experiment, testing dissolved oxygen (left), temperature

(top right), and salinity (bottom right).

A regression was created for the organic matter percentage of both the sun and shade leaf

packs (Figure 2). This regression showed that the shaded leaf packs had a significant negative
linear relationship (F₁, ₁₃= 4.654, P=0.05). As time went on, the organic matter percentage of the

shaded leaf packs decreased (y=-0.0188x+0.903, R²=0.264). However, the sun leaf packs may

appear to have a negative linear relationship, but the regression found this relationship to not be

significant (F₁, ₁₅=3.055, P=0.10). In Figure 2, it does appear that the percentage of organic

matter decreased more for the sun leaf packs than the shaded leaf packs.

Figure 2. Percent organic matter of leaf packs in sunlight (y=-0.024x+0.884, R²=0.1692) and in

shade (y=-0.0188x+0.903, R²=0.264) over time (weeks). Displaying a significant linear

relationship in shade (F₁, ₁₃= 4.654, P=0.05) but not in sunlight (F₁, ₁₅=3.055, P=0.10
Figure 3. Average (± SE) number of macroinvertebrates found in leaf packs placed in St John’s

Creek in sunlight and shaded environment over the course of five weeks.

Figure 3 displays the number of macroinvertebrates we found each time we pulled out

leaf packs. As seen in the figure, there was no significant difference in the number of

macroinvertebrates between sunny and shaded regions at week 3 or week 5 (P=0.93). At week

three, more macroinvertebrates were found in shaded packs than in sunny, and at week five,

more were found in sunny packs. We did not examine the number of different macroinvertebrate

species, but instead focused solely on abundance.

Figure 4 displays the average AFDW of the samples over time, displaying a general

decrease, but not dramatically so. It appears that the sun leaf packs that were pulled during week

five had a much lower AFDW than the shaded leaf packs pulled at the same time. The
differences between each treatment were calculated using a one-way ANOVA test that indicated

that there were differences in AFDW between sun and shaded leaf packs over the course of the

five weeks (F₄,₂₂ = 14.29, P=6.83E-06). The Tukey post-hoc test performed indicated that there

was a significant difference between the sun leaf packs pulled during week five and the rest of

the samples (all P<0.001) and the rest of the samples had no significant difference between them

(all P>0.05).

Figure 4. Average (±SE) Ash Free Dry Weight of leaf packs in sun and shade over time (weeks).

Table 3. Results of Tukey Post-Hoc test performed on AFDW data.

Treatment dif f lwr upr p adj

Week Five Sun-Week Five Shade -2.546 -4.313 -0.78 0.003

Week Three Shade-Week Five Shade 0.051 -1.856 1.959 0.999

 Week Three Sun-Week Five Shade 0.991 -0.916 2.899 0.548

Week Zero Control-Week Five Shade 1.409 -0.499 3.317 0.22

Week Three Shade-Week Five Sun 2.598 0.831 4.364 0.002

Week Three Sun-Week Five Sun 3.538 1.771 5.304 0.00005

0.000010
Week Zero Control-Week Five Sun 3.955 2.189 5.722 2

Week Three Sun-Week Three Shade 0.94 -0.968 2.848 0.596

Week Zero Control-Week Three
Shade 1.358 -0.55 3.266 0.251

Week Zero Control-Week Three Sun 0.417 -1.49 2.326 0.965

Discussion

Overview

The objective of this study was to examine how the decomposition of leaf packs made

from Pin Oak leaves was affected by sunny and shaded conditions. We also examined the

abundance of invertebrates that were collected in both locations. We hypothesized that there

would be higher decomposition in the leaves in sunny areas compared to leaves in shaded areas.

Our findings deviated from this hypothesis. While our regression does make it appear that the

decomposition rate of sunny leaf packs was higher than that of the shaded leaf packs, we could

not conclude this because the regression found there to be no linear relationship between the

organic matter percentages of the sunny leaf packs. However, our analysis of the AFDWs of the

leaf packs does confirm that the sun leaf packs pulled during week five did have a significantly

lower AFDW than the rest of the leaf packs. Additionally, we hypothesized that we would

collect more macroinvertebrates from the leaf packs located in the sunny location; however, our
study found no significant differences in the number of macroinvertebrates between the two

treatments. Therefore, we could not accept either of our hypotheses.

Interpretation of Results

Our results failed to prove that the decomposition rates of sunny and shaded leaf packs

were significantly different. Since the linear regression of the percentage of organic matter

proved to be inconclusive, but the comparison of AFDWs proved that the sun leaf packs at five

weeks had significantly less AFDWs than the shaded packs, we have determined that our results

were inconclusive. We can not say for sure that the organic matter decreased more in sunny leaf

packs due to the fact that the regression showed no linear relationship between the sunny leaf

packs. This deviates from the results of a study conducted by Gallo et al. (2009) who found that

leaves in sunnier areas had significantly higher rates of decomposition than those in shaded

areas. This may be because the location of our study did not receive optimal amounts of sunlight

during the experiment. Since our study site was located in a heavily wooded area, it may have

been possible that our site did not receive enough sunlight to make the shaded area much darker

than the sunny area.

Measured invertebrate abundances showed similar mean values between the sunlit and

shaded locations (P=0.93). This could be because the sunny leaf packs and shaded leaf packs

were placed close together. Macroinvertebrates may have traveled between the sunny and shaded

leaf packs. These findings are inconsistent with the results of a study by Hawkins et al (1982)
who found that the abundance of macroinvertebrates was higher in sunny regions than in shaded

regions.

Limitations and Recommendations

In this study there were some limitations, one being we could not measure the amount of

sunlight in the sunny area and the artificial shaded area created. Some previous studies (Hawkins

et al 1982) chose areas with varying amounts of canopy cover (old -growth canopy vs clear-cut),

this may have been a method we could have utilized, and this method may be used in future

studies. Another solution to this issue would be the funding of a tool to be able to measure

sunlight for future lab experiments. In a possible follow-up study, the sunlight would be

measured and various levels of sunlight could be compared to see how much the sun impacts

decomposition rates.

Another limitation that was faced was the number of leaf packs used in the experiment.

Originally there were forty leaf packs made, but along the way, 12 leaf packs were lost, so we

were down to twenty-eight. This resulted in smaller numbers of leafpacks than anticipated to

collect at each time frame. This was an experimenter error, and in the future, more leaf packs

should be created to allow more replicates and more statistical power. If there was more time

available as well, the leaf packs could have been in the water for longer periods of time, several

months for example. A follow-up study could may be conducted over the course of a year, for

example, and would have much more data to create a more accurate analysis.

Implications

Studying the decomposition of leaf packs in both sunny and shaded conditions revealed

insight into ecosystem dynamics and functioning. It offered an understanding of abiotic factors
that influence decomposition rates, nutrient cycling, and the interactions among decomposer

organisms. This type of study can be used in ecosystem management practices such as stream

restoration and determining the effects of light availability. Further studies can involve a more

controlled or simulated environment where scientists can determine how sunlight affects leaf

packs independently.
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