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Changes in fatty acid composition of plasma of the harp seal Pagophilus groenlandicus during the post-weaning fast
Comp. Biochem. PhysioL Vol. 70B, pp. 795 to 798, 1981
0305-0491/81/120795-04502.00/0
Printed in Great Britain. All rights reserved
Copyright © 1981 Pergamon Press Ltd
CHANGES IN FATTY ACID COMPOSITION OF PLASMA OF THE HARP SEAL PAGOPHILUS GROENLANDICUS DURING THE POST-WEANING FAST BRUCE A. BAILEY, R. G. H. DOWNER, D. M. LAVIGNE,l G. DROLET and G. A. J. WORTHY 1 Department of Biology, University of Waterloo, Waterloo, Ontario, Canada N2L 3G1 and 1Department of Zoology, University of Guelph, Guelph, Ontario, Canada N1G 2W1
(Received 18 March 1981) Abstract--1. The relative percentage composition of palmitic acid, palmitoleic acid, stearic acid, oleic acid, arachidic acid and eicosaenoic acid in plasma of Pagophilus groenlandicus was studied during the post-weaning fast. 2. Plasma fatty acid levels were higher in fasted animals than in feeding animals. 3. The ratio of unsaturated fatty acids to saturated fatty acids tended to be greater in fasting animals than in feeding animals. 4. The data suggest that some of the fatty acids that appear in plasma during starvation are derived from lipid reserves in blubber and core.
INTRODUCTION
MATERIALS AND METHODS
The first month of neonatal development of the harp seal, Pagophilus groenlandicus represents a period of extreme physiological adjustment. Pups are born without an insulating layer of blubber and rely on internal and subcutaneous reserves of thermogenic fat to maintain viable body temperatures until blubber deposition is effected (Grav et al., 1974; Gray & Blix, 1976; Blix et al., 1979). A functional blubber layer is acquired within three days post partum and prodigious accumulation of reserves and growth proceeds for about 9 days (Stewart & Lavigne, 1980). At this time the pups undergo a prolonged post-weaning fast during which energy requirements must be satisfied by endogenous reserves; much of the energy used during the fast may be derived from the body core rather than the more obvious triacylglyceroi reserves present in the blubber, although the latter source cannot be discounted (Stewart & Lavigne, 1980; Bailey et al., 1980). It is evident that the post-weaning fast constitutes a critical period in the life of the harp seal and information on biochemical and physiological changes that occur during this period is of obvious importance. The present study reports relative changes in plasma fatty acid composition associated with the post-weaning fast and the succeeding recovery-period when the animals recommence feeding. Previous studies on the fatty acid composition of selected tissues of P. groenlandicus indicate that at least 60% of the total fatty acid content is represented by saturated and monounsaturated fatty acids with chain lengths of 16, 18 and 20 carbon atoms (Jangaard & Ke, 1968; Ackman et al., 1971; Engelhardt & Walker, 1974); these six fatty acids were monitored throughout the investigation to indicate any changes in the degree of saturation of circulating fatty acids during the fast and/or recovery period.
Animals Twenty recently weaned male pups, all in the early ragged-jacket age category (Stewart & Lavigne, 1980), were removed from the pack ice in the Gulf of St. Lawrence on 17 March 1980, and transported to holding facilities at University of Guelph. The animals were randomly assigned to experimental groups which varied according to the period of fasting. For purposes of the present report the groups are identified as follows:
Group I, force-fasted, animals in this group were fasted for the duration of the study; Group 2, 36-day self-fasted, animals in this group were offered food following a 29-day fast, but did not feed until 36 days; Group 3, 29-day force-fasted, animals in this group were fasted for 29 days after which they were offered and accepted food; Group 4, 8-day fasted, animals in this group were fasted for 8 days after which they were offered and accepted food. Fatty acid analyses Fatty acids were extracted from 1.0 ml aliquots of seal plasma by a slight modification of the method described by Bailey et al. (1980). Following extraction of neutral lipid from total lipid using a mixture of isopropanol:heptane: sulphuric acid (40:10:l/v:v:v) and water (Dole, 1956), a 1.0 ml aliquot of the upper heptane layer was removed for determination of free fatty acids. Fatty acids were esterified with diazomethane (Schlenk & Gellerman, 1960) and, following methylation, nitrogen was purged through the system until the yellow colour disappeared to prevent further reactions. Samples were evaporated to dryness in an atmosphere of nitrogen, and 100/~1 hexane added. Two microlitres of the hexane solution were injected into a Carlo Erba Fractovap Model GV gas chromatograph equipped with dual hydrogen flame ionization detectors and on-column injection system. The gas chromatograph was fitted with dual silanized glass columns (2 m, 3 mm i.d.) loaded with 5% DEGS-PS liquid phase on Supelcoport 100/120 mesh (Supelco Inc., U.S.A.).
795
796
BRUCE A. BAILEY et al. 6
_o
~i
+1+1+1+1+1+1
//
/
~
i
\
\ .
\
// ~
,"A\
//
x
\
I
,x.
, +1+1+1+1+1+1
r ~ t'Nt~
I--
+1+1+1+1+1+1 I
I I0
I
APRIL
I 20
I 30
II
I I0
I 20
MAY
I 30
I
B e~
Fig. 1. Relative fatty acid content of plasma of Pagophilus groenlandicus during and following the post-weaning fast
+1+1+1+1+1+1
(relative fatty acid content expressed as integration units obtained from Spectra Physics System I electronic integrator following separation of derivatised fatty acids by gas liquid chromatography). O O animals commenced feeding on 10.04 after 8-day fast; 0 - . . . . • animals commenced feeding after prolonged fast; x . . . . . . . x animals fasted throughout period of experiment.
+1+1+1+1+1+1
Separation was achieved by temperature programming at 3.0°C/rain for 10 min. The carrier gas, helium, was adjusted to a flow rate of 47 ml/min. The injector temperature was set at 180°C and the manifold and flame ionization detectors at 300°C. Retention times and area percentages were calculated by electronic integration (Spectra Physics System I). The relative molar response coefficients for each fatty acid methyl ester were determined using standards purchased from Sigma Chemical Co., U.S.A. Tridecanoic acid was used as an internal standard, with a known volume being added to each sample prior to derivatisation.
+l+l+l+l+l+l
+1 +1 +1 +l +1 +1 0
8 RESULTS AND DISCUSSION
The relative total fatty acid content of plasma from seals in each of the experimental groups is indicated in Fig. 1. No attempt was made to calculate absolute fatty acid concentrations for each sample; however, in a few samples for which the calculation was performed, the values are compatible with those reported by John et al. (1980). The results demonstrate that fasted animals have higher plasma fatty acid contents than fed animals, a finding that is consistent with reports on the effect of starvation on other vertebrate species (Newsholme & Start, 1973). An important consequence of elevated fatty acid levels in other vertebrate species is that the rise is accompanied by increased oxidation of fatty acids in the liver and the associated production of ketone bodies (Newsholme & Start, 1973). Ketone bodies serve as an important supplement to glucose in providing metabolic energy to the nervous system during periods of glucose deprivation such as occur during starvation. It is probable
+1 +1 +1 +1 +1 +1 a~
g
g +1
O
"Od~
.o ~=
_
~
797
Plasma FA of post-weaning seal Table 2. Fatty acid composition of plasma of Paoophilus groenlandicus during a 29 to 36-day post-weaning fast Date of sample collection Age (days) Number of determinations (n) Palm±tic acid Palmitoleic acid Stearic acid Oleic acid Arachidic acid Eicosaendic acid
27.03 23
04.04 30
5
5
16.6 _ 1.7 14.7 ± 1.9 6.8 + 0.6 33.1 ± 3.4 6.2 ± 0.8 15.5 _+ 3.8
15.0 ± 14.5 ± 7.1 ± 36.2 ± 5.4 ± 14.1 ±
10.04 36
17.04 43
5 1.4 4.8 2.4 1.9 2.5 0.6
16.1 ± 1.6 14.8 ± 2.8 11.9 ± 3.0 35.2 ± 3.9 4.3 __+1.9 11.0 ± 4.8
5
24.04 50 4
16.4 ± 17.4 ± 6.0 ± 37.0 + 5.2 ± 12.9 ±
2.2 3.5 1.7 2.5 3.5 5.8
14.5 ± 15.0 ± 13.7 ± 27.2 ± 10.9 ± 8.5 ±
02.05 58 2
1.1 5.4 7.6 11.0 5.4 5.4
15.5 ___2.5 23.2 ± 6.0 7.0 ± 2.2 41.1 ± 1.1 2.3 ± 1.3 7.3 ± 0.3
Values indicate mean (± SD) percentage of total fatty acid content for n determinations.
that the nervous system of the seal, like that of other vertebrates, requires ketone bodies during starvation and, therefore, the observed rise in plasma fatty acid levels is likely to be accompanied by increased concentrations of ketone bodies. Studies in other vertebrates indicate that the starvation-induced elevation of plasma fatty acids results from increased lipolysis of triacylglycerol reserves (Newsholme & Start, 1973) and, although no attempt has been made to confirm this observation in the harp seal, it is reasonable to propose that a similar mechanism operates in this species. However, the source of triacylglycerol that is mobilised for this purpose remains undetermined. Morphometric (Stewart & Lavigne, 1980) and biochemical (Bailey et al., 1980) studies suggest that there is not a massive mobil±sat±on of blubber fat during the first few weeks of the post-weaning fast, but the procedures employed in these investigations are unlikely to reveal a small amount of lipolysis, thus the blubber remains as a potential source of lipid. Other possible sources of lipid are pockets of fatty material that occur in the close apposition to the intestinal tract and fatty deposits within the musculature (D. M. Lavigne, unpublished observation). Unfortunately the triacylglycerol content of these aggregations and their fate during the post-weaning fast has not been investigated. Clearly, further study of tissue lipase activity and the rate of turnover of plasma fatty acids and
ketone bodies is required in order to assess the role of stored fat in satisfying energy requirements during the first two months of neonatal development. Tables 1--4 indicate changes in the percentages of palm±tic acid, palitoleic acid, stearic acid, oleic acid, arachidic acid and eicosaenoic acid within the total plasma fatty acid content during the post-weaning fast and recovery from the fast. Summation of the six values within an experimental group for any single day of the experiment demonstrates that the six fatty acids included in the study account for a major proportion of the total fatty acid composition of plasma and, therefore, provide a valid basis for assessing changes in the nature and degree of saturation of circulating fatty acids. Several trends are immediately apparent from the results: (1) animals in Group 4 (commenced feeding after 8 days of fasting) do not show any significant differences in fatty acid composition during the month that follows their starting to feed (Table 1); (2) animals in Groups 2 and 3 (fasted for prolonged periods and then fed) tend to show an elevation of saturated fatty acids and a decrease of unsaturated fatty acids when they begin to feed (Tables 2 and 3); (3) animals in Group 1 (force-fasted for the duration of the experiment) retain relatively low percentages of saturated fatty acids and higher percentages of
Table 3. Fatty acid composition of plasma of Pagophilus groenlandicus following commencement of feeding after a 29 to 36-day fast Date of sample collection Age (days) Number of determinations (n) Palm±tic acid Palmitoleic acid Stearic acid Oleic acid Arachidic acid Eicosaendic acid
24.04 50
02.05 58
08.05 64
1
3
4
17.5 13.9 10.7 31.5 7.9 10.1
26.5 ___5.7 10.7 _ 2.3 16.2 ± 3.3 32.2 ± 4.2 2.1 ± 0.8 7.7 _ 3.7
26.1 ± 0.9 6.7 ___2.5 20.8 ± 4.7 19.3 ___4.2 8.6 _± 2.0 2.2 ± 3.0
14.05 70 4 31.7 ± 6.9 ± 21.1 ± 27.3 ± 3.0 ± 3.2 ±
23.05 79 4
3.9 2.1 3.0 3.9 3.8 1.4
22.1 + 2.7 2.8 ± 1.3 25.3 ± 2.5 13.1 ± 5.9 18.4 ± 4.7 trace
Values indicate mean (_ SD) percentage of total fatty acid content for n determinations. c.a.e. 70/4a--I
29.05 85 4 24.9 ± 7.3 + 19.4 + 22.3 ± 8.0 ± 3.8 ±
5.2 2.8 5.6 9.9 4.9 0.1
798
BRUCEA. BAILEYet al. Table 4. Fatty acid composition of plasma of Pagophilus oroenlandieus during a prolonged post-weaning fast Date of sample collection
Age (days) Number of determinations (n) Palmitic acid Palmitoleic acid Stearic acid Oleic acid Arachidic acid Eicosaendic acid
10.04 36 7 15.2 + 1.7 14.2 + 1.8 12.2 _+ 2.7 32.1 _+ 3.5 6.2 _+ 1.9 11.8 _+ 4.3
17.04 43 4 15.6 + 2.9 18.2 _+ 3.1 6.9 + 3.5 37.6 _+ 2.1 2.1 _+ 1.9 14.7 + 5.2
24.04 50 4 12.8 +_ 1.6 14.0 + 3.2 11.3 _+ 2.4 30.6 + 3.2 10.0 _+ 3.0 11.6 _+ 3.9
02.05 58
08.05 64
4
3
13.3 + 0.2 16.6 _+ 1.7 7.0 + 0.9 41.1 _+ 2.8 1.1 _+ 0.4 18.3 + 2.7
14.05 70 2
12.4 + 1.7 14.3 + 2.1 8.1 _+ 1.1 38.6 _+ 3.5 2.6 +_ 0.9 19.5 ___4.3
18.2 +_ 9.2 17.3 _+ 4.6 10.5 + 4.2 36.2 _+ 1.9 1.4 +_ 0.2 13.6 _+ 7.6
23.05 79 1 13.5 12.2 10.2 36.4 6.6 13.4
29.05 85 1 20.8 5.1 21.6 23.3 11.7 trace
Values indicate mean ( _+SD) percentage of total fatty acid content for n determinations.
monounsaturated fatty acids until the final stages of the fast when the situation tends to reverse (Table 4). The fatty acid composition of plasma in a fasted animal reflects that of the tissue(s) from which the fatty acids have been mobilised whereas, in a fed animal, the contribution of dietary components must be acknowledged. The results of Tables 2-4 indicate that the plasma of fasted animals shows a higher percentage of monounsaturated fatty acids than that of feeding animals, and suggests that the tissue(s) from which the fatty acids are derived during starvation may be richer in monounsaturated than saturated fatty acids. Engelhardt & Walker (1974) analysed the fatty acid composition of several tissues from harp seal pups and found that blubber showed a substantially higher percentage composition of unsaturated fatty acids than other tissues. These data, together with the present study, suggest that blubber may provide some of the fatty acid that appears in plasma during starvation although, as indicated in the previous paragraph, other possible sources cannot be eliminated. A further consideration that cannot be excluded from the present discussion is the possibility of fatty acyl specificity in the lipase responsible for hydrolysis of the triacylglycerol reserve. For example mammalian pancreatic lipase has been shown to preferentially cleave unsaturated fatty acids from the 1- and 3-positions of the triacylglycerol molecule (Morley et al., 1974). Indeed retention of unsaturated fatty acids in tissues that may be exposed to extreme environmental temperatures is likely to be advantageous. In conclusion the present study demonstrates the mobilisation of some lipid reserves during the postweaning fast of the harp seal. The site of lipid mobilisation has not been determined although comparison of fatty acid profiles of plasma and other tissues suggest that blubber may contribute to the observed increased in plasma fatty acid content. However, more detailed investigation of other possible sources and additional experimental evidence are required before a definitive statement can be made.
Acknowledgements--Funded by a contract from the Department of Supply and Services through the Research and Resource Branch, Department of Fisheries and
Oceans, Government of Canada and operating grants from the Natural Science and Engineering Research Council of Canada to R.G.H.D. and D.M.L. REFERENCES ACKMAN R. G., EPSTEIN S. & EATON C. A. (1971) Differences in the fatty acid compositions of blubber fats from Northwestern Atlantic finwhales (Balaenoptera physalus) and harp seals (Paaophilus #roenlandica). Comp. Biochem. Physiol. 40B, 683-697. BAILEYB. A., DOWNER R. G. H. & LAVIGNED. M. (1980) Neonatal changes in tissue levels of carbohydrate and lipid in the harp seal Pagophilus 9roenlandicus. Comp. Biochem. Physiol. 67B, 179-182. BLIX A. S., GRAY H. J. & RONALDK. (1979) Some aspects of temperature regulation in newborn harp seal pups. Am. J. Physiol. 236, R188-R197. DOLE V. P. (1956) A relation between non-esterified fatty acids in plasma and the metabolism of glucose. J. clin. Invest. 35, 150-154. ENGELHARDTF. R. & WALKERB. L. (1974) Fatty acid composition of the harp seal, Paoophilus 9roenlandicus (Phoca groenlandica). Comp. Biochem. Physiol. 47B, 169-179. GRAV H. J. & BLIX A. S. (1976) Brown adipose tissue--a factor in the survival of harp seal pups. Can. J. Physiol. Pharmac. 54, 409-412. GRAY H. J., BLIX A. S. & PASCHE A. (1974) How do seal pups survive birth in arctic winter? Acta physiol, scand. 92, 427-429. JANGAARDP. M. & KE P. J. (1968) Principal fatty acids ot depot fat and milk lipids from harp seal (Pagophilus #roenlandica) and the hooded seal (Cystophora cristata). J. Fish Res. Bd Can. 25, 2419-2426. JOHN T. M., MCKEOwr~ B. A., GEORGEJ. C. & RONALDK. (1980) Plasma levels of growth hormone and free fatty acids in the harp seal. Comp. Biochem. Physiol. 66B, 159-162. MORLEYN. H., KUKSISA. & BUCHNEAD. (1974) Hydrolysis of synthetic triacylglycerols by pancreatic and lipoprotein lipase. Lipids, 9, 481-488. NEWSHOLMEE. A. & STARTC. (1973) Regulation in Metabolism. Wiley, London. SCHLENK H. & GELLERMAN J. L. (1960) Esterification ol fatty acids with diazomethane on a small scale. Analyt. Chem. 32, 1412-1414. STEWART R. E. A. 8,£ LAVIGNE D. M. (1980) Neonatal growth in Northwest Atlantic harp seals, Pagophilus #roenlandicus. J. Mammal. 60, 670-680.
0305-0491/81/120795-04502.00/0
Printed in Great Britain. All rights reserved
Copyright © 1981 Pergamon Press Ltd
CHANGES IN FATTY ACID COMPOSITION OF PLASMA OF THE HARP SEAL PAGOPHILUS GROENLANDICUS DURING THE POST-WEANING FAST BRUCE A. BAILEY, R. G. H. DOWNER, D. M. LAVIGNE,l G. DROLET and G. A. J. WORTHY 1 Department of Biology, University of Waterloo, Waterloo, Ontario, Canada N2L 3G1 and 1Department of Zoology, University of Guelph, Guelph, Ontario, Canada N1G 2W1
(Received 18 March 1981) Abstract--1. The relative percentage composition of palmitic acid, palmitoleic acid, stearic acid, oleic acid, arachidic acid and eicosaenoic acid in plasma of Pagophilus groenlandicus was studied during the post-weaning fast. 2. Plasma fatty acid levels were higher in fasted animals than in feeding animals. 3. The ratio of unsaturated fatty acids to saturated fatty acids tended to be greater in fasting animals than in feeding animals. 4. The data suggest that some of the fatty acids that appear in plasma during starvation are derived from lipid reserves in blubber and core.
INTRODUCTION
MATERIALS AND METHODS
The first month of neonatal development of the harp seal, Pagophilus groenlandicus represents a period of extreme physiological adjustment. Pups are born without an insulating layer of blubber and rely on internal and subcutaneous reserves of thermogenic fat to maintain viable body temperatures until blubber deposition is effected (Grav et al., 1974; Gray & Blix, 1976; Blix et al., 1979). A functional blubber layer is acquired within three days post partum and prodigious accumulation of reserves and growth proceeds for about 9 days (Stewart & Lavigne, 1980). At this time the pups undergo a prolonged post-weaning fast during which energy requirements must be satisfied by endogenous reserves; much of the energy used during the fast may be derived from the body core rather than the more obvious triacylglyceroi reserves present in the blubber, although the latter source cannot be discounted (Stewart & Lavigne, 1980; Bailey et al., 1980). It is evident that the post-weaning fast constitutes a critical period in the life of the harp seal and information on biochemical and physiological changes that occur during this period is of obvious importance. The present study reports relative changes in plasma fatty acid composition associated with the post-weaning fast and the succeeding recovery-period when the animals recommence feeding. Previous studies on the fatty acid composition of selected tissues of P. groenlandicus indicate that at least 60% of the total fatty acid content is represented by saturated and monounsaturated fatty acids with chain lengths of 16, 18 and 20 carbon atoms (Jangaard & Ke, 1968; Ackman et al., 1971; Engelhardt & Walker, 1974); these six fatty acids were monitored throughout the investigation to indicate any changes in the degree of saturation of circulating fatty acids during the fast and/or recovery period.
Animals Twenty recently weaned male pups, all in the early ragged-jacket age category (Stewart & Lavigne, 1980), were removed from the pack ice in the Gulf of St. Lawrence on 17 March 1980, and transported to holding facilities at University of Guelph. The animals were randomly assigned to experimental groups which varied according to the period of fasting. For purposes of the present report the groups are identified as follows:
Group I, force-fasted, animals in this group were fasted for the duration of the study; Group 2, 36-day self-fasted, animals in this group were offered food following a 29-day fast, but did not feed until 36 days; Group 3, 29-day force-fasted, animals in this group were fasted for 29 days after which they were offered and accepted food; Group 4, 8-day fasted, animals in this group were fasted for 8 days after which they were offered and accepted food. Fatty acid analyses Fatty acids were extracted from 1.0 ml aliquots of seal plasma by a slight modification of the method described by Bailey et al. (1980). Following extraction of neutral lipid from total lipid using a mixture of isopropanol:heptane: sulphuric acid (40:10:l/v:v:v) and water (Dole, 1956), a 1.0 ml aliquot of the upper heptane layer was removed for determination of free fatty acids. Fatty acids were esterified with diazomethane (Schlenk & Gellerman, 1960) and, following methylation, nitrogen was purged through the system until the yellow colour disappeared to prevent further reactions. Samples were evaporated to dryness in an atmosphere of nitrogen, and 100/~1 hexane added. Two microlitres of the hexane solution were injected into a Carlo Erba Fractovap Model GV gas chromatograph equipped with dual hydrogen flame ionization detectors and on-column injection system. The gas chromatograph was fitted with dual silanized glass columns (2 m, 3 mm i.d.) loaded with 5% DEGS-PS liquid phase on Supelcoport 100/120 mesh (Supelco Inc., U.S.A.).
795
796
BRUCE A. BAILEY et al. 6
_o
~i
+1+1+1+1+1+1
//
/
~
i
\
\ .
\
// ~
,"A\
//
x
\
I
,x.
, +1+1+1+1+1+1
r ~ t'Nt~
I--
+1+1+1+1+1+1 I
I I0
I
APRIL
I 20
I 30
II
I I0
I 20
MAY
I 30
I
B e~
Fig. 1. Relative fatty acid content of plasma of Pagophilus groenlandicus during and following the post-weaning fast
+1+1+1+1+1+1
(relative fatty acid content expressed as integration units obtained from Spectra Physics System I electronic integrator following separation of derivatised fatty acids by gas liquid chromatography). O O animals commenced feeding on 10.04 after 8-day fast; 0 - . . . . • animals commenced feeding after prolonged fast; x . . . . . . . x animals fasted throughout period of experiment.
+1+1+1+1+1+1
Separation was achieved by temperature programming at 3.0°C/rain for 10 min. The carrier gas, helium, was adjusted to a flow rate of 47 ml/min. The injector temperature was set at 180°C and the manifold and flame ionization detectors at 300°C. Retention times and area percentages were calculated by electronic integration (Spectra Physics System I). The relative molar response coefficients for each fatty acid methyl ester were determined using standards purchased from Sigma Chemical Co., U.S.A. Tridecanoic acid was used as an internal standard, with a known volume being added to each sample prior to derivatisation.
+l+l+l+l+l+l
+1 +1 +1 +l +1 +1 0
8 RESULTS AND DISCUSSION
The relative total fatty acid content of plasma from seals in each of the experimental groups is indicated in Fig. 1. No attempt was made to calculate absolute fatty acid concentrations for each sample; however, in a few samples for which the calculation was performed, the values are compatible with those reported by John et al. (1980). The results demonstrate that fasted animals have higher plasma fatty acid contents than fed animals, a finding that is consistent with reports on the effect of starvation on other vertebrate species (Newsholme & Start, 1973). An important consequence of elevated fatty acid levels in other vertebrate species is that the rise is accompanied by increased oxidation of fatty acids in the liver and the associated production of ketone bodies (Newsholme & Start, 1973). Ketone bodies serve as an important supplement to glucose in providing metabolic energy to the nervous system during periods of glucose deprivation such as occur during starvation. It is probable
+1 +1 +1 +1 +1 +1 a~
g
g +1
O
"Od~
.o ~=
_
~
797
Plasma FA of post-weaning seal Table 2. Fatty acid composition of plasma of Paoophilus groenlandicus during a 29 to 36-day post-weaning fast Date of sample collection Age (days) Number of determinations (n) Palm±tic acid Palmitoleic acid Stearic acid Oleic acid Arachidic acid Eicosaendic acid
27.03 23
04.04 30
5
5
16.6 _ 1.7 14.7 ± 1.9 6.8 + 0.6 33.1 ± 3.4 6.2 ± 0.8 15.5 _+ 3.8
15.0 ± 14.5 ± 7.1 ± 36.2 ± 5.4 ± 14.1 ±
10.04 36
17.04 43
5 1.4 4.8 2.4 1.9 2.5 0.6
16.1 ± 1.6 14.8 ± 2.8 11.9 ± 3.0 35.2 ± 3.9 4.3 __+1.9 11.0 ± 4.8
5
24.04 50 4
16.4 ± 17.4 ± 6.0 ± 37.0 + 5.2 ± 12.9 ±
2.2 3.5 1.7 2.5 3.5 5.8
14.5 ± 15.0 ± 13.7 ± 27.2 ± 10.9 ± 8.5 ±
02.05 58 2
1.1 5.4 7.6 11.0 5.4 5.4
15.5 ___2.5 23.2 ± 6.0 7.0 ± 2.2 41.1 ± 1.1 2.3 ± 1.3 7.3 ± 0.3
Values indicate mean (± SD) percentage of total fatty acid content for n determinations.
that the nervous system of the seal, like that of other vertebrates, requires ketone bodies during starvation and, therefore, the observed rise in plasma fatty acid levels is likely to be accompanied by increased concentrations of ketone bodies. Studies in other vertebrates indicate that the starvation-induced elevation of plasma fatty acids results from increased lipolysis of triacylglycerol reserves (Newsholme & Start, 1973) and, although no attempt has been made to confirm this observation in the harp seal, it is reasonable to propose that a similar mechanism operates in this species. However, the source of triacylglycerol that is mobilised for this purpose remains undetermined. Morphometric (Stewart & Lavigne, 1980) and biochemical (Bailey et al., 1980) studies suggest that there is not a massive mobil±sat±on of blubber fat during the first few weeks of the post-weaning fast, but the procedures employed in these investigations are unlikely to reveal a small amount of lipolysis, thus the blubber remains as a potential source of lipid. Other possible sources of lipid are pockets of fatty material that occur in the close apposition to the intestinal tract and fatty deposits within the musculature (D. M. Lavigne, unpublished observation). Unfortunately the triacylglycerol content of these aggregations and their fate during the post-weaning fast has not been investigated. Clearly, further study of tissue lipase activity and the rate of turnover of plasma fatty acids and
ketone bodies is required in order to assess the role of stored fat in satisfying energy requirements during the first two months of neonatal development. Tables 1--4 indicate changes in the percentages of palm±tic acid, palitoleic acid, stearic acid, oleic acid, arachidic acid and eicosaenoic acid within the total plasma fatty acid content during the post-weaning fast and recovery from the fast. Summation of the six values within an experimental group for any single day of the experiment demonstrates that the six fatty acids included in the study account for a major proportion of the total fatty acid composition of plasma and, therefore, provide a valid basis for assessing changes in the nature and degree of saturation of circulating fatty acids. Several trends are immediately apparent from the results: (1) animals in Group 4 (commenced feeding after 8 days of fasting) do not show any significant differences in fatty acid composition during the month that follows their starting to feed (Table 1); (2) animals in Groups 2 and 3 (fasted for prolonged periods and then fed) tend to show an elevation of saturated fatty acids and a decrease of unsaturated fatty acids when they begin to feed (Tables 2 and 3); (3) animals in Group 1 (force-fasted for the duration of the experiment) retain relatively low percentages of saturated fatty acids and higher percentages of
Table 3. Fatty acid composition of plasma of Pagophilus groenlandicus following commencement of feeding after a 29 to 36-day fast Date of sample collection Age (days) Number of determinations (n) Palm±tic acid Palmitoleic acid Stearic acid Oleic acid Arachidic acid Eicosaendic acid
24.04 50
02.05 58
08.05 64
1
3
4
17.5 13.9 10.7 31.5 7.9 10.1
26.5 ___5.7 10.7 _ 2.3 16.2 ± 3.3 32.2 ± 4.2 2.1 ± 0.8 7.7 _ 3.7
26.1 ± 0.9 6.7 ___2.5 20.8 ± 4.7 19.3 ___4.2 8.6 _± 2.0 2.2 ± 3.0
14.05 70 4 31.7 ± 6.9 ± 21.1 ± 27.3 ± 3.0 ± 3.2 ±
23.05 79 4
3.9 2.1 3.0 3.9 3.8 1.4
22.1 + 2.7 2.8 ± 1.3 25.3 ± 2.5 13.1 ± 5.9 18.4 ± 4.7 trace
Values indicate mean (_ SD) percentage of total fatty acid content for n determinations. c.a.e. 70/4a--I
29.05 85 4 24.9 ± 7.3 + 19.4 + 22.3 ± 8.0 ± 3.8 ±
5.2 2.8 5.6 9.9 4.9 0.1
798
BRUCEA. BAILEYet al. Table 4. Fatty acid composition of plasma of Pagophilus oroenlandieus during a prolonged post-weaning fast Date of sample collection
Age (days) Number of determinations (n) Palmitic acid Palmitoleic acid Stearic acid Oleic acid Arachidic acid Eicosaendic acid
10.04 36 7 15.2 + 1.7 14.2 + 1.8 12.2 _+ 2.7 32.1 _+ 3.5 6.2 _+ 1.9 11.8 _+ 4.3
17.04 43 4 15.6 + 2.9 18.2 _+ 3.1 6.9 + 3.5 37.6 _+ 2.1 2.1 _+ 1.9 14.7 + 5.2
24.04 50 4 12.8 +_ 1.6 14.0 + 3.2 11.3 _+ 2.4 30.6 + 3.2 10.0 _+ 3.0 11.6 _+ 3.9
02.05 58
08.05 64
4
3
13.3 + 0.2 16.6 _+ 1.7 7.0 + 0.9 41.1 _+ 2.8 1.1 _+ 0.4 18.3 + 2.7
14.05 70 2
12.4 + 1.7 14.3 + 2.1 8.1 _+ 1.1 38.6 _+ 3.5 2.6 +_ 0.9 19.5 ___4.3
18.2 +_ 9.2 17.3 _+ 4.6 10.5 + 4.2 36.2 _+ 1.9 1.4 +_ 0.2 13.6 _+ 7.6
23.05 79 1 13.5 12.2 10.2 36.4 6.6 13.4
29.05 85 1 20.8 5.1 21.6 23.3 11.7 trace
Values indicate mean ( _+SD) percentage of total fatty acid content for n determinations.
monounsaturated fatty acids until the final stages of the fast when the situation tends to reverse (Table 4). The fatty acid composition of plasma in a fasted animal reflects that of the tissue(s) from which the fatty acids have been mobilised whereas, in a fed animal, the contribution of dietary components must be acknowledged. The results of Tables 2-4 indicate that the plasma of fasted animals shows a higher percentage of monounsaturated fatty acids than that of feeding animals, and suggests that the tissue(s) from which the fatty acids are derived during starvation may be richer in monounsaturated than saturated fatty acids. Engelhardt & Walker (1974) analysed the fatty acid composition of several tissues from harp seal pups and found that blubber showed a substantially higher percentage composition of unsaturated fatty acids than other tissues. These data, together with the present study, suggest that blubber may provide some of the fatty acid that appears in plasma during starvation although, as indicated in the previous paragraph, other possible sources cannot be eliminated. A further consideration that cannot be excluded from the present discussion is the possibility of fatty acyl specificity in the lipase responsible for hydrolysis of the triacylglycerol reserve. For example mammalian pancreatic lipase has been shown to preferentially cleave unsaturated fatty acids from the 1- and 3-positions of the triacylglycerol molecule (Morley et al., 1974). Indeed retention of unsaturated fatty acids in tissues that may be exposed to extreme environmental temperatures is likely to be advantageous. In conclusion the present study demonstrates the mobilisation of some lipid reserves during the postweaning fast of the harp seal. The site of lipid mobilisation has not been determined although comparison of fatty acid profiles of plasma and other tissues suggest that blubber may contribute to the observed increased in plasma fatty acid content. However, more detailed investigation of other possible sources and additional experimental evidence are required before a definitive statement can be made.
Acknowledgements--Funded by a contract from the Department of Supply and Services through the Research and Resource Branch, Department of Fisheries and
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