Schneider
Lipid and Glycogen Content
INTRODUCTION
Work done on lipid and glycogen metabolism in poly¬
chaetes is scarce. However, Giese (1966) reports many studies
on other invertebrates. Table 1 presents the results of studies
on glycogen and lipid contents of various polychaetes. Changes
in the glycogen and lipid of several annelids and the sipunculids
Phascolosoma agassizii and Phascolosoma gouldii have been in-
vestigated.
Terebella lepidara showed a marked decrease in body
wall lipid content after ten days of starvation. Arenicola
marina showed a body wall lipid content increase after six
weeks of starvation and a four fold increase in the gut lipids
during the same period. Dales (1957a)
Von Brand (1927) studied glycogen contents in polychaetes
and concluded that the substance in the worms he studied is
used in times of oxygen stress. Scheer (1969) reports on
conversion of glycogen to lactic acid under anaerobic conditions
in oligochaetes.
Dales (1958) found that Arenicola marina uses its
glycogen stores during anaerobiosis, while Owenia fusiformis,
which has a lower glycogen content, does not show an appreciable
depletion of glycogen reserves. Instead, Dales suspects a
reduction in metabolic rate. No evidence of lactic or pyruvic
acid was found.
Towle and Giese (1966) measured the effect of starvation
on Phascolosoma agassizii. The original carbohydrate level of
4.1% and glycogen of .8% (dry weight) did not appreciably de¬
Lipid and Glycogen Content
Schneider
crease. Instead, the lipids were metabolised and fell from
4.1% to 3.3% in seventeen days and to 2.7% in seventy-one days.
Wilbur (1947) has done starvation studies on the sip¬
unculid Phascoloma gouldii. After four weeks the lipid content
decreased from 3.86% to .8% but the muscle lipid content in¬
creased from 1.32% to 2.02%, possibly indicating the utilisation
of proteins found there.
The above data posed an interesting question. Some
polychaetes such as Terebella lepidara showed decreases in
lipid contents during starvation while increases in lipid con¬
tents were noted in Arenicola marina. Thus, the metabolism of
lipids during starvation did not seem to follow any general
pattern. Neither did the usage of glycogen during starvation
seem to be a well explored topic. The study reported here
is an investigation of lipid and glycogen content of the
polychaetes, Myxicola infundibulum (Renier 1804), Cirriformia
luxuriosa (Moore 1904), Halosydna brevisetosa (Kinberg 1855),
and Phragmatopoma californica (Fewkes 1899), and the effect
of short term starvation on these reserves.
MATERIALS AND METHODS
All animals were collected from the Monterey area
in central California. M. infundibulum and H. brevisetosa
were collected subtidally at the Monterey marina. P. cali¬
fornica was taken by chiseling a portion of a colony found
past the east end of Aggassiz beach at Hopkins Marine Station.
C. luxuriosa was taken from the mud around the roots of the
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Lipid and Glycogen Content
eel grass Phyllospadix. The worms were either analysed fresh
from the field or deprived of food as follows. M. infundibulum
and C. luxuriosa were placed in a plastic pan supplied with
running, glass fiber filtered sea water at 12.0-13.5° C.
Sixteen worms were analysed immediately or after one
and two weeks of starvation. Each of sixteen worms of each
species was homogenised in 1 ml of distilled water per wet
weight gram of worm except for P. californica. Sixteen samples
of five to eight P. californica were substituted because of
their small size. Lipids were first extracted using three
ten ml portions of a 50:50, volume to volume, mixture of
95% ethyl alcohol and diethyl ether. The insoluble material
was removed by centrifuging at 2000 rpm for 10 minutes. The
ethanol-ether was poured into a beaker, evaporated to dryness,
and re-extracted with two five ml portions of chloroform.
The chloroform was then poured into a pre-weighed pan and evap¬
orated to dryness.
The material insoluble in ethanol-ether was then assayed
for glycogen by the method of Somogyi (1934). Two ml of 50%
sodium hydroxide were added for each gram of original tissue
and digested for three hours at 100° C. After digestion the
alkaline mixture was centrifuged for 10 minutes at 2000 rpm.
The clear alkaline fluid was decanted and the remaining matter
was washed with two ml of distilled water for each gram of
tissue, shaken, and centrifuged for 10 minutes at 2000 rpm.
This fluid was then united with the first and one ml of 95%
ethanol added for each two ml of fluid. The glycogen was
allowed to precipitate overnight.
Lipid and Giycogen Content
Schneider 4
The next day the carbohydrates were collected by
centrifugation at 2000 rpm for fifteen minutes, the supernatant
poured off, and the pellet washed with 95% ethanol.
Total glycogen was determined colourimetrically by
the phenol-sulphuric acid method of Dubois (1956). Values
obtained were compared with a standard curve prepared with
glucose,
RESULTS
The results of the lipid and glycogen determinations
are given in Table 2 as the mean and standard deviation of the
sixteen samples of worms. In general the levels of these
materials correspond to the values reported for other polychaetes
in Table 1.
The starvation experiments carried out with C. lux¬
uriosa had a range of glycogen contents at time O of .019% to
19%. See Figure 1. The mean was .054% +.042. After one
week the range was from .021% to .11% with a mean of .064% + .026.
After two weeks of starvation the glycogen content ranged
from .020% to .065% with a mean of .042% +.013. The decrease
over the last week was significant at p less than .02 using a
"t" test. The range of values converged on a lower limit in
each case of.019%.
The lipids in C. luxuriosa showed an increase over
two weeks of starvation; see Figure 2. At O time the lipid
content ranged from 1.6% to 2.8% with a mean of 2.24% + .41.
After one week the values ranged from 1.9% to 3.6%. The mean
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Lipid and Glycogen Content
value was 2.69% + .47. This increase is significant at p less
than .007. After two weeks the lipid content ranged from 2.1%
to 3.1% with a mean of 2.56% +.36. This was also significantly
higher than at O time with p less than.013 but not significantly
different from the value at one week.
For M. infundibulum, the glycogen was found to range
from .021% to .069% of wet body weight. The mean was found to
be .043% + .015. One week of starvation produced a range of
.014% to .064% glycogen with a mean of .034% +.021; see Figure
3. Two weeks of starvation showed the worm's glycogen to
range from .014% to .079% and the mean being .037% +.020.
None of these values showed any statistically significant
change over the two week period.
The lipids of M. infundibulum initially ranged from
2.5% to 3.8% with a mean of 3.28% +.41. One week later the
lipids comprised from 2.3% to 4.4% of body weight, the mean
being 3.22% + .67; see Figure 4. After two weeks without
nourishment, the worms showed lipid contents of from 2.0% to
3.8% with a mean of 2.84% + .53. The decrease in lipid content
in M. infundibulum is significant at p less than.02.
DISCUSSION
The subtidal worms Halosydna brevisetosa and Myxicola
infundibulum both show the two highest lipid contents and the
two lowest glycogen levels. The intertidal worms, Phragmatopoma
californica and Cirriformia luxuriosa show lower lipid contents
and slightly higher glycogen contents. Whether this is of
adaptive significance or just coincidence can not be deter¬
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Lipid and Glycogen Content
mined at this point. The levels of glycogen and lipid in these
worms corresponded to levels reported in other polychaetes.
Cirriformia luxuriosa seems to be relying on glycogen
and some other reserve material, possibly protein, to survive
periods of starvation. During the two week period the glycogen
content of the worms decreased, implying the metabolism of
this energy reserve. The upper limit of the ranges also
consistently decreased over the course of starvation while the
lower limit remained constant. This indicates a basic body
level of glycogen which may not be metabolised. As the worms
starve, they seem to use up their glycogen to the point where
it comprises.019% of wet body weight.
The lipid content as a percentage of body weight appear¬
ed to increase over the period of two weeks. This can be
explained in one of two ways. Either the annelid has syn¬
thesised lipids or has used some other reserve to a greater
extent than lipids. The latter hypothesis seems the more
likely. Unless there is a decrease in metabolic rate during
starvation, something other than glycogen must be the substrate
of this metabolism because the decrease in glycogen over the
two week period is small, 120 micrograms/gram of worm.
In Myxicola infundibulum there was no significant
change in glycogen during the period of starvation. However,
there did seem to be a lower limit to glycogen content here
as in Cirriformia luxuriosa. Myxicola infundibulum did show
a significant decrease in lipid levels over the two week period.
This decrease of .44% of body weight in lipid content is
6
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Lipid and Glycogen Content
attributed to lipid utilisation by the worm. Thus, it appears
that this worm does not appreciably utilise glycogen at the
onset of starvation, but instead metabolises its lipids for
energy.
It seems that these polychaetes do not rely solely
on glycogen as an energy reserve. One species, Cirriformia
luxuriosa, seems to call upon proteins or some other reserve
to make up for an energy deficit during early starvation.
Another polychaete, Myxicola infundibulum, seems to rely upon
lipid stores at the onset of starvation. This is entirely
consistent with the previous observations of Dales (1957a),
Towle and Giese (1966), and Wilbur (1947).
Lipid and Glycogen Conter
Schneider
SUMMARY
1.
Lipid and glycogen content determinations were done
on Myxicola infundibulum, Cirriformia luxuriosa,
Halosydna brevisetosa, and Phragmatopoma californica
collected from the Monterey Bay area of central California.
Myxicola infundibulum and Cirriformia luxuriosa
were starved for two weeks and their lipid and
glycogen contents assayed.
Myxicola infundibulum appears to rely on stored lipids
at the onset of starvation.
Cirriformia luxuriosa appears to use some other
reserve at the onset of starvation.
ACKNOWLEDGEMENTS
I would like to thank Dr. John Phillips for his
invaluable help and encouragement during the course of this
project.
Lipid and Glycogen Contents
Schneider 9
REFERENCES
Brand, T. von 1927
2. Vergleich Physiol. 5: 643-698
Preliminary Observations on the Role
Dales, R. P. 1957a
of the Coelomic Cells in Food Storage
and Transport in Certain Polychaetes.
J. Mar. Biol. U. K. 36: 91-110
Feeding in Sabellids and Serpulids.
Dales, R.P. 1957b
J. Mar. Biol. U.K. 36: 309-316
Survival of Anaerobic Periods by two
Dales, R.P. 1958
Intertidal Polychaetes: Arenicola marina
and Owenia fusiformis.
J. Mar. Biol. U.K. 37: 521-529
J.K. Hamilton, P.A. Rebers, & F. Smith 1956
Dubois, M., K.A. Giles,
Colorimetric Method for Determination
of Sugars and Related Substances.
Analyt. Chem. 28: 350-356.
Constitution et Proprietes Physico
Fremiet, E.F. 1929
Chemiques des Eleocytes D'Amphritrite
johnstoni (Malmgren).
Protoplasma 5: 321-337
Lipids in Marine Invertebrates.
Giese, A.C. 1966
Physiological Review 46: 244-298
Chemical Zoology Academic Press,
Scheer, B. & M. Florkin
New York, 1969
The Solubility and Preparation of
Somogyi, M. 1934
Phosphorous and Nitrogen Free Glycogen.
J. Biol. Chem. 104: 245-253
Biochemical Changes During
Towle, A. & A.C. Giese
1966
Reproduction and Starvation in the
Sipunculid Worm Phascolosoma aggassizi.
Comp. Biochem. Physiol. 19: 667-680
Effect of Prolonged Starvation on the
Wilbur, C.G. 1947
Lipids in Phascolosoma gouldii.
J. Cell. Comp. Physiol. 29: 179-183
Wilbur, C.G. & W.M. Bayors 1947 A Comparative Study of the
Lipids in Some Marine Annelids.
Biol. Bull. 93: 99-101
Table 1
Species
% Lipid % Glycogen
Amphritrite
johnstoni
wet wt.
.8%- 4% 1.2%
Nereis
diversicolor
wet wt.
1.43% -1.73%
Nereis
pelagica
wet wt.
2.17%
Glycera
americana
wet wt.
2.75%
Amphritrite
ornata
wet wt.
3.15%
Arenicola
marina
wet wt.
1.22%
rite
Amphri
johnstoni
dry wt.
27.6%
Nere
pelagica
3.81%
wet wt.
Arenicola
marina
wet wt.
.54%
Phoronopsis
viridis
4.9%
dry wt.
Sabella
starki magnificans
1.5%
dry wt.
9.2%
Glycera rugosa
dry wt.
8.3%
1.4%
Reference
Dales (1957a)
Dales (1957a)
Wilbur & Bayors (1947)
Wilbur & Bayors (1947)
Wilbur & Bayors (1947)
Wilbur & Bayors (1947)
Fremiet (1929)
von Brand (1927)
von Brand (1927)
Giese (1966)
Giese (1966)
Giese (1966)
Table 2
Lipid and Glycogen Content
species
% ipid
%o glycogen
Cirriformia
054% -042
2.24%-41
luxuriosa
Myxicola
Q43%- 015
3.28%—41
infundibuum
Halosydna
3.66% 51
018% - .008
previsetosa
Phragma¬
137%- 073
2.95%.60
topoma
californica
VALUES ARE MEAN PERCENT OF WET WEIGHT
FOLLOWED BY STANDARD DEVIATIONS
Figure 1.
Change in the glycogen content of Cirriformia
luxuriosa with starvation. Each point
represents the total glycogen content of
one worm expressed as a percent of wet
weight.
20
18
16
14
12
00
08
glyc.
06
04
02
O0
e
Figure 2.
Change in lipid content of Cirriformia
luxuriosa with starvation. Each point
represents the total lipid content of
one worm expressed as a percent of wet
weight.
40
G.O
lipid
2.0
1.0
o
o
o
O
o8

Weeks

2
0
Figure 3.
Change in the glycogen content of Myxicola
infundibulum with starvation. Each point
represents the total glycogen content of
one worm expressed as a percent of wet
weight.
087
O6
glyc.
04
02
OO
o

2
Weeks
c
Figure 4. Change in the lipid content of Myxicola
infundibulum with starvation. Each point
presents the total lipid content of one
worm expressed as a percent of wet weight.
5.0
40
lipid
O3.C
2.0
1.0
088
0

2

Weeks