FISHERIES OCEANOGRAPHY
7:3/4, 265±271, 1998
Feeding activities of zooplankton in the Bohai Sea
RONG WANG,* CHAOLUN LI, KE WANG
AND WUCHANG ZHANG
Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai
Road, Qingdao 266071, China
ABSTRACT
The Bohai Sea was the site of the Chinese national
GLOBEC programme. During the June 1997 cruises of
R/V Science No.1, observations and experiments on
zooplankton feeding were conducted. At ®ve 48 h
time-series stations the following observations and
measurements on zooplankton were carried out: (1)
diurnal vertical migration, by collecting samples at
different layers every 3 h with a closing net; (2)
diurnal feeding rhythms, by gut pigment analysis; and
(3) ingestion rate, by both gut pigment analysis and the
dilution method. A classi®cation by body size was used
to deal with the diversity of species and developmental
stages of zooplankton assemblages. Samples were sep-
arated into three size groups: small (200±500 lm),
medium (500±1000 lm) and large (> 1000 lm). The
results showed that the copepods (Calanus sinicus,
Paracalanus parvus, Acartia bi®losa and Centropages
mcmurrichi) performed clear diurnal vertical migra-
tions. However, their behaviour was different at dif-
ferent stations. The variation in gut pigment content
over the 24 h cycle showed strong diurnal feeding
rhythms, particularly for the large size group. Gut
pigment contents reached their daily maximum during
the time from dusk to midnight (18:00±24:00). The
peak value was about 10 times the minimum observed
in the daytime. The in situ daily grazing rate, based on
gut pigment contents and evacuation experiments, was
4.00±12.65 ng chla ind)1 day)1 for the small size
group, 5.99±66.58 ng chla ind)1 day)1 for the medium
size group and 31.31±237.13 ng chla ind)1 day)1 for
the large size group. The copepods consumed only a
small part (2.90±13.52%) of the phytoplankton bio-
mass but about 77% of the daily production. The
*Correspondence. Fax: + 86 532 2870882;
e-mail: wangrong@ms.qdio.ac.cn
Received for publication 20 March 1998
Accepted for publication 30 June 1998
Ó 1998 Blackwell Science Ltd.
Fish. Oceanogr.
grazing mortality rate of phytoplankton by microzoo-
plankton (< 200 lm) measured by the dilution
method ranged from 0.43 to 0.69 day)1 The calculated
daily consumption of phytoplankton biomass was 35±
50%, and 85±319% of the potential production.
Key words: Bohai Sea, feeding, zooplankton
INTRODUCTION
The Bohai Sea is a shallow, semi-enclosed sea charac-
terized by strong seasonal ¯uctuation in hydrographic
condition. It is surrounded by heavily populated areas
in the northern part of China. During the past decades,
over®shing, sea farming in the nearshore water and
changes in land-derived freshwater, nutrients and sed-
iments have reached levels that put the environment
under stress and resulted in changes in structure and
function of the marine ecosystem. For these reasons,
the Bohai Sea was chosen as the site of the ®rst phase of
the China±GLOBEC (Global Ocean Ecosystem Dy-
namics) programme from 1997 to 2000, of which the
dynamics of zooplankton populations was one of the
four principal foci. The work presented here gives
preliminary results based on the data collected during
the R/V Science No.1 cruises in June 1997, focused on
zooplankton feeding. The major objective of this study
was to evaluate the importance of different size groups
of zooplankton in their grazing impact on phytoplank-
ton, so as to obtain parameters on the utilization of
primary production by herbivorous copepods.
MATERIALS AND METHODS
Observations and experiments were designed to
measure the grazing pressure of copepods on phyto-
plankton in situ. At ®ve 48 h time-series stations
(Fig. 1), diurnal vertical migration, diurnal feeding
rhythms and ingestion rate of planktonic copepods
were observed and measured in the period 3±12 June
1997. A classi®cation based on body size was used to
separate the diversity of species and developmental
stages of copepods. All observations and experiments
were conducted separately by size groups: small
(200±500 lm), medium (500±1000 lm) and large
(>1000 lm). CTD, nutrients (N, P, Si), chlorophyll
265
266 R. Wang et al.
Figure 1. Time-series (48 h) observation stations in the
Bohai Sea.
a (¯uorescence) and primary production (14C) were
measured simultaneously. Phytoplankton and zoo-
plankton samples were collected for subsequent ex-
amination of their taxonomic composition and
abundance.
Sampling
To observe diurnal vertical migration of copepods,
zooplankton samples were collected in different depth
layers (0±5 m, 5±10 m, 10±20 m, 20 m±bottom) at
different times of the day (00:00, 03:00, 06:00, ¼,
24:00) for a complete daily cycle. A single closing net
with mouth area of 0.5 m2 and mesh size of 333 lm
was used. A ¯owmeter was mounted in the centre of
the mouth to measure the volume of water ®ltered.
Numbers of different species in the samples were
counted under a dissecting microscope. Samples for
gut pigment analysis were collected by vertical tows
from the bottom to the surface every 3 h at the same
time as the sampling for diurnal vertical migration,
using two nets of different mesh sizes. One, with
500 lm mesh, was used for sampling the large and
medium size groups; the other, with 200 lm mesh, was
used for sampling the small size group. Live animals for
the gut pigment evacuation experiments were col-
lected by using the same nets as for the gut pigment
analysis, and those for the dilution experiments were
collected using a 50 L water sampler. Water samples
from different depths were collected and preserved
with Lugol's solution for studies of taxonomic com-
position and abundance of microzooplankton.
Ó 1998 Blackwell Science Ltd., Fish. Oceanogr., 7:3/4, 265±271.
Gut pigment analysis
Gut pigment analysis was used to study the diurnal
feeding rhythms (Head et al., 1984). The cod-end
content was immediately poured into soda/seawater
solution (1:5, v/v) to anaesthetize the animals (Kleppel
et al., 1988), then sieved through 1000, 500 and
200 lm mesh, and the resultant fractions ®ltered onto
GF/C ®lters. The ®lters were frozen and kept at ± 20°C
until subsequent analysis. In the laboratory, copepods
on the ®lter, 30 individuals for the large size group, 40
for the medium size group, and 50 for the small size
group, were transferred to another GF/C ®lter under a
low-light dissecting microscope without regard to spe-
cies or stage. Subsequent sample treatments and ¯uo-
rescence measurements were as described by Mackas
and Bohrer (1976). Absolute values for the chlorophyll
level and the chlorophyll equivalent weight of phaeo-
pigment were calculated according to Wang (1986)
and Wang and Conover (1986). The sum was used as
the index of gut fullness. A conversion ratio of 43 was
used for carbon:chlorophyll (Wang and Fan, 1995).
Gut pigment evacuation experiments
The gut pigment evacuation method was used to es-
timate the in situ grazing rate for the copepod assem-
blage larger than 200 lm. Evacuation experiments
were conducted on board ship in a constant-temper-
ature water bath set to the surface seawater tempera-
ture. Animals were ®rst transferred gently to a
container and then separated equally into six sub-
samples, which were kept in 2 L beakers, and then
immersed in the water bath. Initially, and after 10, 20,
30, 60 and 120 min, animals of each subsample were
anaesthetized and then sieved through 1000, 500 and
200 lm mesh gauze. Different fractions were ®ltered
onto GF/C ®lters. Subsequent treatments and mea-
surements were the same as for gut pigment analysis.
The gut pigment content at time t (Gt) was related to
time as: Gt ˆ Goe)rt, where Go is the gut pigment
content at the beginning of the experiment. Evacua-
tion rate (r) can be calculated from the plot of gut
pigment against time of incubation. Ingestion rates (I)
were calculated from the expression: I ˆ r ´ G.
Dilution technique
The dilution technique developed by Landry and
Hassett (1982) was used to estimate the grazing impact
of zooplankton of less than 200 lm in size. Natural
seawater was collected from 2 m depth. Half of the
water was ®ltered twice by GF/F ®lter. The remaining
seawater was ®ltered through a sieve with a mesh size of
200 lm. The 200 lm ®ltered water was then combined
with the GF/F ®ltered water in ratios of 1:0, 3:1, 1:1

and 1:3. Three 2 L glass reagent bottles were ®lled
with each dilution mixture and then incubated in situ
at the same depth for 24 h. The apparent growth rates
of phytoplankton in individual bottles were calculated
based on the changes in chlorophyll concentration.
RESULTS
Diurnal vertical migration
The relative abundance of copepods in different layers
at different times of the day clearly showed diurnal
vertical migration for most species (Fig. 2), moving
upwards after dusk and migrating downwards before
dawn. For the larger species, Calanus sinicus and Cen-
tropages mcmurrichi, the amplitude of displacement was
about 5±10 m. The behaviour of the same species
showed some differences at different stations. For in-
stance, at Station 4, where a thermocline was well
developed, Calanus sinicus performed more distinct
diurnal vertical migrations (Fig. 2c). However, at
Station 5 (Fig. 2d), where no clear thermocline was
found, the amplitude of their migration was much less
than those at Station 4. Acartia bi®losa, a dominant
species in the small group, tended to live in the upper
layer, moving down to the middle layer during the
Figure 2. Diurnal vertical migration of copepods and chaetognath species in the Bohai Sea (solid lines show the diurnal
variation of the depth of distribution centre).
Ó 1998 Blackwell Science Ltd., Fish. Oceanogr., 7:3/4, 265±271.
Zooplankton feeding in the Bohai Sea 267
daytime (Fig. 2 m). At Station 5, this species re-
mained in the upper layer (0±10 m) all day (Fig. 2n).
Paracalanus parvus, another small copepod, performed
a similar vertical migration as Calanus sinicus but the
amplitude was somewhat less for P. parvus (Fig. 2i,k).
The major carnivore, Sagitta crassa, performed a similar
diurnal vertical migration to that of its prey (Fig. 2h,j).
Diurnal feeding rhythms
The diurnal variation in gut pigment content (Fig. 3)
showed a strong diurnal feeding rhythm, particularly
for the large size group. High gut pigment content
occurred from dusk to midnight (18:00±24:00). The
peak value was about 10 times the minimum occurring
in the daytime. The daily averages for all size groups
were much lower than expected (Table 1), possibly
owing to the low chlorophyll concentration in the
water at the time of year. For the small size group, the
diurnal feeding rhythms were less apparent.
Grazing rate and grazing pressure on phytoplankton
Daily grazing rates (Table 1) were calculated from the
data on gut pigment contents and evacuation rate, and
then converted to the ration for all copepods in the
water column (Table 2) by using the population den-
268 R. Wang et al.
Figure 2h±k.
sity data. The daily consumption of phytoplankton
biomass by copepods (> 200 lm) was low because of
the low phytoplankton biomass, but it represented a
signi®cant proportion of the primary production (Ta-
ble 2). To estimate the grazing impact of microzoo-
plankton less than 200 lm in size, we used the dilution
technique (Fig.4). By this method, the daily grazing was
estimated at 35±50% of the phytoplankton biomass or
85±319% of primary production, which approached or
exceeded the potential production (Table 3).
DISCUSSION
Copepods, especially the larger species such as Calanus
sinicus and Centropages mcmurrichi, performed clear di-
urnal vertical migrations in the Bohai Sea, but their
behaviour was somewhat different at different stations.
They were typical at Station 4 (Fig. 2c), with the bulk
of the population staying near the bottom during the
day and ascending during the night. Compared with
other stations, a strong thermocline was present at
Station 4. At Stations 1, 3 and 5, where no clear the-
rmocline was found, the migrations of Calanus sinicus
and Centropages mcmurrichi were less strong and the
distribution centres were higher than those at Station 4.
Acartia bi®losa, a dominant species in the small group,
Ó 1998 Blackwell Science Ltd., Fish. Oceanogr., 7:3/4, 265±271.
Figure 2l±n.
also performed clear diurnal vertical migration at Sta-
tion 4. However, at Station 5, the bulk of the popula-
tion remained in the upper layer all day. The conclusion
is that there is no direct relation between food con-
centration and vertical migration. The chlorophyll a
concentration at the time of sampling was low, with an
average for the entire water column being around
1 g L)1. It was distributed either homogeneously
throughout the water column or with the concentration
in the deeper layers being higher than that in the upper
layers; thus there was no constant relationship between
chlorophyll concentration and vertical migration.
In the Bohai Sea, zooplankton biomass reaches its
annual maximum in June (Bai and Zhuang, 1991) but
the phytoplankton biomass decreases to a low level in
this month after the spring bloom (Kang, 1991). That
may be why we observed a low level of the gut pig-
ment content in all three copepod size groups. Even
though the gut pigment level was low, the diurnal
variation in gut pigment contents revealed a clear
diurnal feeding rhythm, particularly for the large size
fractions, i.e. resting from feeding during the daytime
and feeding actively from dusk to midnight for only a
short period due to the shortage of food. It may be
that the diurnal feeding rhythm is an intrinsic evo-
lutionary attribute of the herbivorous copepods and
has no direct relationship with environmental factors
(food concentration, temperature, etc.). It emphasizes
that estimations of the daily grazing rate must be
based on a complete daily feeding cycle.

Figure 3. Diurnal variation in gut
pigment content of copepod assem-
blages in the Bohai Sea.
Grazing rates of the copepod assemblages measured
by gut pigment analysis were much lower than ex-
pected. That is explained by the high density of
copepods and low concentration of phytoplankton.
For instance, at Station 4 the average density of Cal-
anus sinicus was as high as 309 m-3, but the average
chlorophyll a concentration in the water column was
only 1.05 g L ) 1, although there might be some
sources of underestimation in the present results such
as pigment destruction by digestion in the gut (Con-
over et al., 1986; Wang and Conover, 1986). However,
it is likely to be proportional to the total pigment
quantity ingested. The copepods consumed only a
small part (2.90±13.52%) of the phytoplankton bio-
mass owing to the low feeding ef®ciency under the
unfavourable conditions of low food availability but
high competition. Nevertheless, this small part of the
phytoplankton biomass consumed is equivalent to
Ó 1998 Blackwell Science Ltd., Fish. Oceanogr., 7:3/4, 265±271.
Zooplankton feeding in the Bohai Sea 269
about 77% of daily production. The primary produc-
tion measured at the ®ve observed stations during the
feeding experiments ranged from 100 to 136 mg C
m)2 day)1, which is only about one-third of the an-
nual average (308 mg C m)2 day)1) of the entire
Bohai Sea (Fei et al., 1991). It seems that for most of
the copepods, the energy ingested cannot cover the
respiration needs estimated using Ikeda's (1985)
equation. Evidence of a population decrease caused by
food shortage in June can be found from the historical
data. The year-round observations in the Bohai Sea in
1992±93 showed that phytoplankton biomass de-
creased dramatically from May to June (Kang, 1991),
whereas zooplankton biomass reached its annual
maximum in June (255.9 mg m)3) and then decreased
sharply to a relatively low level (98.9 mg m)3) in July.
The population density of Calanus sinicus declined
from its annual maximum of 179 m)3 to 47 m)3
270 R. Wang et al.

Table 1. Gut pigment content, evacuation rate and grazing rate of copepod assemblages in the Bohai Sea in June 1997.

Gut pigment
Size contents Evacuation Gut passage Grazing rate
Station Water depth group (ng chla ind.)1) rate (min)1) time(min) (ng chla ind.)1day)1)

large 1.647 0.0132 76 31.31

1 30 medium 0.341 0.0122 82 5.99
small 0.257 0.0108 93 4.00
large 3.283 0.0310 32 146.55
3 18 medium 1.578 0.0293 34 66.58
small 0.188 0.0203 49 5.50
large 1.578 0.0445 22 101.12
4 28 medium 0.348 0.0464 22 23.25
small 0.205 0.0303 33 8.94
large 8.667 0.0190 53 237.13
5 29 medium 1.299 0.0300 33 56.12
small 0.363 0.0242 41 12.65

Table 2. Phytoplankton biomass, primary production and grazing pressure by copepods in the Bohai Sea in June 1997.

Phytoplankton Primary Daily grazing rate Grazing pressure on

biomass productivity
Station mg chla m)2 (mg C m)2 day)1) mg chla m)2 mg C m)2 Biomass (%) Production (%)

1 28.97 119.03 0.84 42.00 2.90 35.29

3 17.01 136.27 2.30 115.05 13.52 84.43
4 17.68 100.34 2.38 118.75 13.46 118.35
5 31.19 93.89 1.85 92.60 5.93 70.05

Table 3. Grazing impact of microzooplankton on phytoplankton based on dilution experiments in the Bohai Sea in June 1997.

Sampling depth Grazing on Grazing on prim.

Station (m) r2 k (day)1) g (day)1) Biomass (% day)1) prod. (% day)1)

1 2 0.7786 0.6748 0.6909 49.89 101.66

2 2 0.5159 0.4307 0.4256 34.66 99.05
4 2 0.537 0.7326 0.5854 44.31 85.32
5 2 0.4356 0.2294 0.5959 44.89 319.00

k = phytoplankton growth coef®cient, g = microzooplankton grazing coef®cient.

during this same period (Bai and Zhuang, 1991).
Possibly non-living particulate organic matter from
land sources and from re-suspension of sediments may
serve as part of the food sources during this time of
year for the ®lter feeders, as we observed in the East
China Sea (Liu et al., 1997; Wang and Fan, 1997).
The grazing impact of microzooplankton on phy-
toplankton was high. It accounted for almost half of
the daily standing crop of phytoplankton and 99.05±
319.00% of the potential daily primary production.
Codonellopsis spp. (Tintinnidae), nauplii of copepods
and larvae of benthic invertebrates dominated this
Ó 1998 Blackwell Science Ltd., Fish. Oceanogr., 7:3/4, 265±271.
fraction. Size-fractionated primary production data
showed that, as the average of the ®ve observed sta-
tions, 84.33% of the measured primary production
was contributed by nano- and pico-fractions (unpub-
lished data from Dr X. Ning, Second Institute of
Oceanography, State Oceanography Administration,
Hangzhou, China). It would appear that the micro-
zooplankton may play a more important role in
transferring the primary products to higher trophic
levels at this time of year. In support of this, Burkill
et al. (1993) showed evidence that microzooplankton
and their herbivorous activity provide the major route

Figure 4. Micro-zooplankton (< 200 lm)
grazing by the dilution technique with
®tted regressions.
for the herbivorous fate of phytoplankton in the north-
east Atlantic Ocean in June and July.
ACKNOWLEDGEMENTS
This research was supported by National Natural
Science Foundation of China, 49790010. We thank
Captain Wu Chuanjie and the crew of the R/V Science
No.1 for invaluable assistance in collecting zoo-
plankton samples and in doing experiments on board
ship. We are also grateful to Dr Roger Harris and the
two anonymous reviewers for helpful comments on the
draft manuscript.
REFERENCES
Bai, X. and Zhuang, Z. (1991) Studies on the ¯uctuation of
zooplankton biomass and number in the Bohai Sea. Mar.
Fish. Res. 12:71±92. (In Chinese with English abstract.)
Burkill, P.H., Edwards, E.S., John, A.W.G. and Sleigh, M.A.
(1993) Microzooplankton and their herbivorous activity in
the northeast Atlantic Ocean. Deep-Sea Res. II 40:479±
493.
Chen, G., Niu, Y. and Wen, S. (1991) Marine Atlas of the Bohai
Sea, Yellow Sea and East China Sea. Beijing: China Ocean
Press, 250 pp.
Conover, R.J., Durvasula, R., Roy, S. and Wang, R. (1986)
Probable loss of chlorophyll derived pigments during passage
through the gut of zooplankton and some of the conse-
quences. Limnol. Oceanogr. 31:878±887.
Fei, Z., Mao, X. and Zhu, M. (1991) The study of primary
production in the Bohai Sea: chlorophyll a, primary
productivity and potential ®sheries resources. Mar. Fish. Res.
12:55±69. (In Chinese with English abstract.)
Ó 1998 Blackwell Science Ltd., Fish. Oceanogr., 7:3/4, 265±271.
Zooplankton feeding in the Bohai Sea 271
Head, E.J.H., Wang, R. and Conover, R.J. (1984) Comparison of
diurnal feeding rhythms in Temora longicornis and
Centropages hamatus with digestive enzyme activity. J. Plank.
Res. 6:543±551.
Ikeda, T. (1985) Metabolic rates of epipelagic marine zoo-
plankton as a function of body size and temperature. Mar.
Biol. 85:1±11.
Kang, Y. (1991) Distribution and seasonal variation of phyto-
plankton in the Bohai Sea. Mar. Fish. Res. 12:31±54. (In
Chinese with English abstract.)
Kleppel, G.S., Pieper, R.E. and Trager, G. (1988) Variability in
the gut contents of individual Acartia tonsa from water off
Southern California. Mar. Biol. 97:185±190.
Landry, M.R. and Hassett, R.P. (1982) Estimating the grazing
impact of marine micro-zooplankton. Mar. Biol. 67:283±288.
Liu, W., Wang, R. and Ji, P. (1997) Study on particulate organic
carbon in the East China Sea. Oceanologia Limnol. Sin.
28:39±43. (In Chinese with English abstract.)
Mackas, D. and Bohrer, R. (1976) Fluorescence analysis of
zooplankton gut contents and an investigation of diel feeding
patterns. J. Exp. Mar. Biol. Ecol. 25:77±85.
Wang, R. (1986) Correction of the formula to convert ¯uores-
cence measurement to phaeophorbide a concentration by
the acidi®cation method. Mar. Sci. 10:1±5. (In Chinese with
English abstract.)
Wang, R. and Conover, R.J. (1986) Dynamics of gut pigment in
the copepod Temora longicornis and the determination of in
situ grazing rates. Limnol. Oceanogr. 31:867±877.
Wang, R. and Fan, C. (1995) Gut pigments and evacuation rates
of copepod assemblages in the East China Sea. In: Global
Fluxes of Carbon and Its Related Substances in the Coastal Sea±
Ocean±Atmosphere System. Shizuo Tsunogai, Kazuo Iseki,
Isao Koike and Yadamichi Oba (eds). Yokohama: M & J
International, pp. 95±106.
Wang, R. and Fan, C. (1997) Copepod feeding activities and its
contribution to downwards vertical ¯ux of carbon in the East
China Sea. Oceanologia Limnol. Sin. 28:39±43. (In Chinese
with English abstract.)