Asiatic Herpetological Research

1997

Vol. 7. pp. 139-146

Egg Components and Utilization During Incubation in the Turtle,
Chinemys reevesii
Pei-chao Wang, Wei Ma, Bo Lu and Wen-hui You
Department of Biology, East China Normal University, Shanghai20()(->2,

China

Abstract. -The energy content, water content and ratios of egg components in the turtle. Chinemys reevesii, were
determined during 0-55 days of incubation. The total water content of the egg took 65% of egg mass on

preincubation and increased 5.66% during incubation time. The total water content of the egg on preincubation
was found 60.7% in the yolk, 35% in the albumen and 4.3% in the shell. The ratios of those varied with theembryonic age (days) growth, and the greater portion the water in the egg had transferred to the embryonictissues on 55 days of incubation when the water content of the embryo was 50.4% of the egg total water content.

The dried mass of shell, yolk and albumen on preincubation were 35.3%, 61.0% and 3.1% of the egg dry mass,
The component materials of those, except for a major portion of the remained shell, wererespectively.
transferred into the embryonic tissues with the incubation days as the total transferred rate was 79.3%. and the

The lipid content in dried yolk averaged 20.4% of the yolk's dried mass during incubation.
The energy content of the yolk, albumen and embryo were relatively stable through the incubation period andcost rate for 20.7%.

averaged 6462.5. 5087.7 and 6291.4 cal/g. respectively.

Key words:

Turtles. Chinemys reevesii. egg components, energy content. Incubation.

Materials and Methods

Introduction
Oviparous

reptilescharacteristicallyallocatethemajority of their total reproductive investment to theireggs (Fischer et al.. 1991). The eggs of the turtle.

Chinemys

reevesii,

investmentin thereflect

the

total

offspring by females.

reproductive
This post-

hatching yolk is the source of energy for the survivaland growth of the hatchlings in some reptiles duringdays of life (Kraemer and Bennett. 1981:
Congdon et al.. 1983a. 1983b: Troyer, 1987; Wang et
the

al.,

first

1990.

1995).

The component materials andOne hundred eggs of the common turtle. Chinemys
reevesii,

were collected in the morning after the ovi-

position of the females in June 1987 and 1988 at a turtle

farm

near

East

China

NormalUniversity

inShanghai. China. Each egg was removed from thenest on the day of laying, marked and weighed with atorsion balance (±0.01 mg).

Ten fresh eggs were used

for the standard (normal) level of the egg component

The remaining 90
were
incubatedinanenvironmental
eggs
temperaturematerials and the energy content.

energy in the turtle egg are first utilized for embryonicdevelopment and, secondarily, for hatchling mainte-

of 28±0.05° on a water-saturated substrate (sand), andnance after it leaves the egg. However, the utilability

misted with water. Those hatching eggs were used forsome analyses of the ratios of the component materi-rates of thecomponent materials, such as the shell,
albumen and yolk of the Chinemys reevesii egg forembryonic development at the different stages of
incubation are not clear.

How many component mate-rials of the egg are allocated andwhich are necessary

for the embryonic development and hatchling'.'

So far.

only limited data are available on changes of the shell,

albumen and yolk rations in the egg during the different stages of the incubation for reptiles.
In this paper,men, yolk and

we report some component (albu-

the inside of the incubation chamber was periodicallyals and the energy content during the different stagesof incubation (Tables 1-6).

When

the

eggs

for

assay

were cracked, theembryo, yolk, albumen and shell were carefully separated and each one was separately placed in a pre-weighed glass container and then weighed.

Theembryo was separated from the yolk and both were
freed from the surrounding choriallantoic and viteline
tissue,and then blotted dry and weighed.

The albu-

and energy values infreshly laid egg of the turtles. Chinemys reevesii, andmen was cleaned away from the shell membranes, andalso the changes of thedry and weighed.

shell)

ratioscomponent ratios during thedifferent stages of incubation.

the shell

was rinsed with distilled water and blotted

The samples were then dried to a

constant mass in an oven at 60°C. All the weighings

1997

Asiatic Herpetological ResearchVol. 7. p. 140

embryos was 79.31% (SD=2.4)
and the rate of cost was 20.69% (SD=2.39).

were accurate to the nearest milligram. All of thedried materials were stored in a desiccator until

to the tissues of theassayed.

Water Content in Eggs. The water content in eggs

Water content was calculated as the difference inthe net wet and dry mass of the egg materials. The
contents of the dry embryo, yolk and albumen

was determined by subtracting the dried mass fromthe fresh mass of the egg. and the results are shown inenergywere then measured. Triplicates of each individual
determine thesample were assayed by combustion toenergy content with the microbamb calorimeter (JR-Table 3.

2000 type made in China) and benzoic acid standards.
The results were corrected for free ash energy content.

bation days slightly increased (Table 3) as the mean
rate my reach 9.2% until 55 days of incuba-

day of incubation, the water content ofDuringeach egg averaged 65% (SD=6.3. N=10). The rates of
those after 20 days of incubation raised with the incu-

growthThe lipid content of the yolk was removed withether from an aliquot off the dry yolk masstion.

time lasted 5 hours.

The amount of lipid was determined by subtracting the mass of the sample after
extraction from the sample mass before extraction.

The water content of the shell averages 16.1%(SD=1.3. N=76) throughout the incubation period.
The water in the yolk on
day of incubation was
65.5% (SD=7.8. N=10), but the value of that declined
as the incubation time increased (Table 3). The water

The sample weighings were accurate to the nearest

in themilligram.

increased.

petroleumin a soxhlet extractor;

each length of the extractive

embryos also declined as the incubation time
But, the water in the albumen kept in arange of 95.3-97.3% on 0-55

Results(Table 3).

The changes of the egg components of Chinemys
reevesii at the different incubation times are shown in
Tables 1,2 and 3.
the fresh
Eggshell. During day zero of incubation,
fresh
the
of
mass of the eggshell was 14.6%
egg(Table1)days of incubationand the dried massof the eggshellwas

The values of the

35.3% of the dried egg (Table 2).

shell declined as the time of incubation increased, andexhibited an age phase of incubation (Tables 1 and 2).

lipid
Lipid Content in Yolk. Table 4 shows that
content (%) in the dried yolk on the first periods ofincubation reduced slightly, and then, increased with
the incubation time did as raised to 26.51% on 55

thedays on incubation.

Energy Values of Egg Contents. The energy values
and embryo throughout(cal/g) of the yolk, albumen
incubation time are shown in Table 5.
Thecaloric

values(free-ash)of thewet-yolk

Yolk Content and Incubation Time. During the firstincreased with the incubation days and those data are

the majordays of incubation, the yolk contents took
and2). But. thosethe(Tablesofcomponents
egg
ity
contents declined as the incubation time increased.

fitted into the

When the hatchlings left their eggshells, the remain-

relatively in the range of 6343.9-6609.8 cal/g.

1of the fresh yolk mass
ing yolk was only about 6-9%

following equation: for wet yolk, cal/But. theg=2356.52±31.43 days, r=0.9745, P<0.01.
caloric values (free-ash) of the dried yolk stabilized

The caloric values of the embryonic mass also

on the zero day of incubation.

increased with the incubation time (Table 5) as theAlbumen Contents. The fresh albumen contents (%)

relationof the egg were higher after than before 20 days ofBut. dried albumen (%) of the).
incubation (Table

incubation expressed a positive linear regression correlation for Y=1306.32+97.44X, r=0.9820. P<0.01.

1

The caloric values of dried embryonic mass were sta-egg mass expressed relative stability (Table 2).

Embryonic Mass. From 0-10 days of incubation, theembryos had not been determined because of a little
tissue of each embryo. We found that the embryonic
wet and dry mass of Chinemys reevesii increased withthe time (days) of incubation (Tables 1 and 2). The
recently

emergedhatchlings

averaged

between caloric values (Y) and days (X) of

4.8

g

79.31% (SD=2.4) of the total(SD=0.78. N=10)
mass
the
in
content
compared with day zero ofeggorincubation, so that, the rate of the material transferredbilized in the range of 6257.7-6337.7 cal/g (free-ash).

The albumen energy values are the lowest among
the totalcomponents of the egg (Table 5). Such asmean energy values of wet albumen is 204.8 cal/g andis 5087.7 cal/g throughout the period
of incubation, but only equal to 5.95% of wet anddried albumen

76.7% of dried yolk energy, respectively.

1997

Asiatic Herpetological Research

Discussion

The

Vol. 7. p. 141

newly hatchlings of Chinemys reevesii
4.8
averagedg (SD=0.78. N=10) for 68.41% of the
fresh egg content (albumen
and yolk) mass at

Type and Mass of Eggshelljust

The types of eggshell in the turtles were classified to

28±0.5°C of the incubation temperature.

be both fundamental patterns of the rigid-shell (orbrittle-shelled) and the flexible-shelled; in the former,

egg contents may provide for the embryonic development, the transferred rate from the egg contents into

the shell mass take above 30% of the whole egg driedembryonic tissues was 68.41%, and the rate of costmass and below 21% of one in the latter (Congdon

The dried mass of shells averaged 35.3% of the total dried mass of eggs in Chineand Gibbons, 1985).

mys reevesii and should be classified as a type of rigid
and the mass of that is approximate to
36.44% of Gopherus polyphemus eggshell mass andis lower by 4-8% than those of Kinosternon subru-

don and Gibbons. 1985).

was 31.59% at 28±0.5°C of the incubation tempera-

Chinemys reevesii eggs. The transferred rate
Trionyx triunguis egg contents is 75.21%(Leshem et al.. 1991). But. the transferred rate isof theinfluenced by the temperature of incubation, at 30°C
for

65.54% and

at

33°C

for

62.61%

declined as the incubation days increased (Table

1

and

in

Chinemys

reevesii egg contents (Wang et al., 1990).

Movement of Water Inside Eggs. The

The shell mass in the egg in Chinemys reevesiiof theture ineggshell,hnini, K. coloratus and Clemmys marmorata (Cong-

If all

total

water

content of each Chinemys reevesii fresh eggs at prein-cubation averaged 65% that the percent water approx-

66.9-70%2),

imatescium in the eggshell that was possibly transferred into

mississippiensis egg (Fischer et al., 1991). The watercontent of the reptile eggs have been defined as thedeclining a total of 37.8% in fresh eggshell and
6.2% in the dried mass of eggshell with a part of calthe developing embryo.
tents of calcium

We have determined the con-and magnesium in whole egg (Table

6) for tested the above hypothesis.

The results (Table

to

water

of

the

Alligator

hydric condition necessary for successful completionof the incubation (Packard and Packard, 1988). We

found that the

total

lost

water about

20% laid egg6) show that ( 1 ) the calcium and magnesium contents

mass in the Grass lizard (Takydromus septentrionalis)

were less in posthatching eggshells than in preincuba-

had failed to hatch.

tion ones; (2) the calcium and magnesium contents ofmuch more water from its surrounding substrata forthe egg contents (yolk and albumen) only equaled

the embryonic development completion, and the mass

45.93% and 87.74% of those in hatchlings. respec-of eggs may be increased 342.2% of thetively.

Obviously calcium and magnesium in the egg

In addition, the egg must absorb

mass over the course of incubation
1989).

In this study, the total

initialegg(Wang et al.,water content of thecontents were not enough to provide the embryonic
The other 54.07% of calcium anddevelopment.

Chinemys

12.26% of magnesium in the newly hatchlings may(SD=2.61, N=76) of the initial egg mass during incu-

come from reserves in the eggshell.

bation. But. eggs of some species hatch successfullyeven declining appreciably in mass. The eggs of thepapers also support our results:
cium in embryos of sea turtles

Some previous

the sources of cal-came 60-80% fromreevesiiegg

increased

only

5.66%

al.,

snapping turtles (Chelydra serpentina) frequently
hatch after declining in mass by 18% over the course1969) and the 56% of calcium in embryonic snappingof incubation (Morris et al., 1983); the eggs of soft-

Chelydra serpetins, obtained also from thedevelopment required (Packard et al.. 1984b).

mately 21% water of the egg without ill effect during

stores in the eggshell (Simkiss, 1967; Bustard etturtles.

Growth of Embryos and Consumption of Yolk.

shelled turtles (Trionyx triunguis)

may lose approxi-

incubation (Leshem et al., 1991 ).

We found that the yolk content declined as the

The total water content of the Chinemys reevesii

incubation time increased, the embryo mass increased

egg on the preincubation reserved 60.7% in the yolk.
35% in the albumen and 4.3% in the shell (Table 3).

as the incubation time did.

Both reflect a potential
betweenrelationshipyolk consumption and embryonic growth.

Because the turtle egg is a semi-closed

system, all course of material transference, energy
flow and embryonic development is carried out in thesystem throughout the incubation period. The yolk of
the turtle is the main sources of energy and materials
for the embryonic development, so the yolk of Chine-

mys reevesii declined as the embryo grew during the
incubation times.

The ratios of those varied with the incubation times(Table 3). The changes of the ratios may cause theyolk consumption and the embryonic growth; forexample, on 55 days of incubation, the wet mass ofyolk was only 7.5% of egg mass and the wet embryonic mass had grown to 45.9% of the egg mass (Table
1)when the water of the yolk took only 4.9% of theegg total water content and the water of embryo took
50.4% of the egg total water content (Table 3). Sec-

Asiatic Herpetological ResearchVol. 7. p. 142

ondarily, yolk is also the approximate source of waterused by embryos (Morris et al., 1983), so a quantity ofwater was transferred to the embryo.

The albumen

and shell water content were apparently not changed(Table 3).

Literature Cited
Bustard. H. R.. K. Simkiss and N. K. Jenkibs.

Some

the

C.

reevesii

embryodeclined significantly with the embryonic age (Table3) that the changes of the relative hydration are simi-previous reports for the embryos of Greeniguana {Iguana iguana) (Ricklefs and Cullen. 1973)
toand Colubrid snake {Coluber constrictor) (Packard et1969.

incubated eggs

ofanalyses
artificially
hatchlings of green and loggerhead sea turtles.
nal of Zoology, London.

The water content of

lar

1997

and
Jour-

158:31 1-315.

Congdon, J. D. and J. W. Gibbons. 1985. Egg components and reproductive characteristics of turtles:
relationship to body size.

Herpetologica, 41(2): 194-

205.

Congdon.

J.

D.

D..

W. Tinkle andP.

C.

Rosen.

an index to the maturity of the embryo and can be1983 A. Egg components and utilization during development in aquatic turtles. Copeia 1983:265-268.

used to estimate embryonic age.

Congdon.

al.,1984).

The water content of the embryo may be

Lipid of Yolk. The rate of the lipid content in the dryyolk of Chinemys reevesii averaged 20.4% during
incubation.

The value of that is close to 21% of theegg lipid in pained turtles, Chrysemys picta (Congdon
et al., 1983).

The lipid in the yolk may play an impor-

1983B.

J.

D..

J.

W. Gibbons andinvestment

Parental

J.

Greene.

L.

the chickeninturtle(Deirocheiys reticularis). Ecology 64:419-425.
Fischer. R. U., F. J. Mazzotti. J. D. Congdon and R. E.
Jr. 1991. Post-hatching yolk reserves: pareninvestment in American Alligators from Louisiana.

Gatten,tal

providesan embryo with greater amounts of usable metabolic

Journal of Herpetology 25(2):253-256.

energy and water (Gutzke and Packard, 1987), and themost part of one is transferred to the hatchling for itof the hydric and thermal environments on eggs andcost in the first days after it leaves the egg (Fischer etPhysiological Zoology 60( 1 ):9-17.

tant role through embryonic development.

Ital„ 1991).

Gutzke. W. H. N. and G. C. Packard.

1987. Influencehatchlings of bull snakes. Pituophis melanoleucus.

Kraemer, J. E. and J. S. Bennett.

1981. Utilization ofEnergy of Egg Contents. The changes of the energy
content (cal/g) in the wet mass of albumen, yolk andposthatching yolk in loggerhead sea
caretta. Copeia 198 1 (2):406-4 I.

embryo of the Chinemys reevesii are affected by theirwater content. However, the energy content in thoseof dried mass are relatively stable through incubation(Table 5). The caloric value, 6291.4 cal/g (SD=31.8),

1991.
Leshem, A., A. Ar and R. A. Ackerman.
Growth, water and energy metabolism of the soft-of the dried yolk in Chinemys reevesii is close to thevalue of those in the snapping turtles. Chelydra ser-pentina (6.6 cal/g), and the slider turtles. Pseudemys
scripta (6.7 Kcal/g). at the time of laying (Slobodkin.
1962).

The total energy content of the dry yolk in the

Chinemys reevesii increased as the lipid content of theyolk did. Both show positive correlation, r=0.6768(P<0.01 ). and the correlation is significant.

shelled turtle (Trionyx

Caretta

tri unguis)embryoeffectoftemperature. Physiological Zoology 64(2):568-594.
Morris, K. A., G. C. Packard, Boardman, G. L. Pautkstis

and M. J. Packard.

1983.

Effect of the hydricenvironment on growth of embryonic snapping turtles{Chelydra serpentina). Herpetologica 39:272-285.
1988. The physioInofreptilian eggs and embryos.
logical ecology
B
Carl
Vol.
of
the
16.
(Eds.
Ecology
Reptilia,
Biology

Packard. M. J. and M. J. Packard.

Gans and Raymond B. Huey)

Acknowledgments

turtles.

1

Alan R. Liss.

New

York. pp. 523-605.

Packard, M. J.. G. C. Packard and W. H. N. Gutzke.

This research was supported by the Natural ScientificFund of Educational Committee of China. Natural
Scientific Fund of China, and East China Normal University. We would like to thank Dr. Allan Muth, Dr.
Cary C. Packard. Dr. J. Whitfield Gibbons and Dr.
Justin D. Congdon for providing some literature, andalso special thanks to Prof. Ermi Zhao for his advice
on my manuscript.

1984.

Calcium metabolism in embryos of the ovipa-

rous snake Coluber constrictor.

Journal of Experi-mental Biology 110:99-112.
Packard, M. J., T. M. Short. G. C. Packard andGorell.

1984.

T A.

Sources of calcium for embryonicdevelopment in egg of the snapping turtles, Chelydra
serpentina. Journal of Experimental Zoology 230:8187.

Asiatic Herpetological Research

1997

Ricklefs, R.

E.

and

J.

Cullen.

1973.

Embryonic

growth of the growth of the green iguana {Iguana
iguana). Copeia 1973:269-305.
Simkiss, K.
ogy.

1967.

Appendix
Tables 1-6

Calcium in reproductive physiol-

A Comparative Study of Vertebrates.

Reinhold

Publishing Corporation. New York.
1962. Energy in animal ecology.
Advances in Ecological Research 1:69-1(11.

Slobodkin, L. B.

Troyer, K. 1987. Posthatching yolk in a lizard: internalization and contribution growth. Journal of Herpe-

tology 21(21:102-106.

Wang. P., X. Ji and W. Ma. 1989. Oxygen consumptionof
Takydromus septentrionalis eggs andhatchlings. Current Herpetologica Asia (Eds. by Matsui M., T.

Kikida and R.C. Goris).

HerpetologicalSociety of Japan, pp. 49-53.

Wang. P.. W. Ma. B. Lu and W. You. 1990. Studies on
ecology of incubation of eggs Chinemys reevesii. In
Ermi Zhao (ed.). From Water onto Land. China Forestry Publishing House. Beijing,pp.

113-119.

c(In

Chinese, with English abstract)

Wang. P., W. Ma. B. Lu and W. You.

o
.Q

1

995. Studies onOcphysiological ecology of incubation in the Chinemys
reevesii

eggs.

Asiatic

HerpetologicalResearch

6:151-156.
<D

I
C

c
ID

o

a
E
o
o

O)
Ol
iD

en

c

ro

.3!

Vol. 7. p. 143

IVol. 7, p. 144

g
'13

r:

o

c:

"a

S
a?

I
C
-£

O
c
0)

C
o
Q.

E

o
o
CD

c
CD

o

-O

Asiatic Herpetological Research

1997

1997

Asiatic Herpetological Research

.

3

E

Vol. 7. p. 145

o

*

5?

^

en

a

r-

\_i

r-

C

5

K

*t

3

"0

K

to

Si

< E

_;

<*.

K

O

—

N

^

3 C

5 E

£

Si

«J

>o

'_

fo

»

r*~.

w

r-~

—

w

p

3

in

O

-

—

—

—

c-

r-

m

Vol. 7. p. 146

Asiatic Herpetological Research

Table 4. The lipid contents in dry mass of yolk of Chinemys reevesii eggs during incubation.

Days

1997