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Clathrate hydrate of oxygen: does it occur in deep-sea fish?
D~p-Se,a Research, 1975, Vol. 22, pp. 145 to 149. Pergamon Press. Printed in Great Britain.
Clathrate hydrate of oxygen: does it occur in deep-sea fish? EDVARD A. HEMMINGSEN*
(Received 20 May 1974; accepted5 October 1974) Abslraet--Oxygen exists in the swimbladder of some deep-sea fish at partial pressures that are more than twice the hydrate equilibrium pressure in water. Observations suggest that no hydrate forms to depths of 3500-4000 m. At greater depths, gas secretion may not occur because of the formation of hydrate. INTRODUCTION THE GAS-FILLEDswimbladder of fish functions as a buoyancy organ, the volume of which is regulated by secretion or reabsorption of gas. Because the gas pressure in the swimbladder essentially equals the hydrostatic pressure acting on the fish, gas pressures of several hundred atmospheres exist in many deep-sea fish. About 90%, or more, of this swimbladder gas is oxygen; the remainder is mainly nitrogen and some carbon dioxide (SCHOLANDERand VAN DAM, 1953; SCHOLANDER,1954; DOUGLAS,1967). The deepest samples have been obtained from specimens of Coryphaenoides abyssorum collected at 3512 m, in which case the oxygen content was equivalent to a partial pressure of about 320 atm (DOUGLAS, 1967). Even higher pressures may exist; two species of brotulid fish with well-developed swimbladders have been collected at depths from 4575 to 7160 m (NIELSENand MUNK, 1964). Many gases, including oxygen, form solid hydrates in aqueous solutions. These hydrates are ice-like crystalline clathrate compounds in which the gas molecules are enclosed in a hydrogen-bounded water lattice (JEFFREY and MCMULLAN, 1967; KLOTZ, 1970). For water at 2°C, a typical deep-sea temperature, the oxygen hydrate equilibrium, or dissociation pressure, is about 148 atm (VANCLEEFFand DIEPEN, 1965). Because the oxygen pressures in the swimbladder reach values of more than twice the hydrate equilibrium pressure, conditions for the formation of oxygen hydrate exist in these fish. Such a natural occurrence of gas hydrate, the possibility of which appears to have been overlooked in previous swimbladder studies, would be unique not only for biological systems, but also for the Earth as well, with the possible exception of gas hydrates deep in the polar ice caps (MILLER, 1969). The formation of oxygen hydrate in the swimbladder or its tissue could have major effects on gas secretion, on the maintenance of a gas phase, and possibly on the survival of the fish. One question is whether or not gas dissolved in water at pressures substantially above the gas hydrate equilibrium value can be maintained in a metastable state without formation of crystalline gas hydrate. An experimental system was set up to determine to what degree such metastability may prevail with respect to oxygen. It was also of interest to determine the gas tension in the solution at gas pressures higher than the hydrate equilibrium value, and this was accomplished indirectly by measuring the diffusion of dissolved gas from the solution through a Teflon barrier, using methane, which has a lower hydrate equilibrium pressure. *Physiological Research Laboratory, Scripps Institution of Oceanography, La Julia, California 92037, U.S.A. 145
146
EDVARD A . HEMMINGSEN
METHODS
The apparatus and the experimental procedures were as follows: within a pressure chamber, distilled water in a glass beaker was stirred with a magnetic bar and was equilibrated with oxygen or methane gas at various pressures. Each water sample usually was equilibrated at several pressures, with stepwise increases in the pressures from low to high values. Each equilibration lasted at least 2 h. The water in the chamber could be observed through a window for visual evidence of crystalline hydrate formation. A coiled Teflon tube (0.041 cm i.d., 0.085 cm o.d., and I 1 cm long) was submerged completely in the water. One end of the tube was sealed, and the other end was brought to the outside via a cut-off hypodermic needle (Gauge 26). The gas flux from the water through the Teflon barrier was measured by means 0fthe movement of a meniscus in a capillary tube attached to the hypodermic needle outlet. The temperature of the apparatus was controlled to ~0.02°C by submersion in a regulated water bath. The experiments with oxygen were carried out at l~C. Because of partial or complete collapse of the Teflon tube at pressures greater than about 100 atm, measurements of the oxygen flux through the tube wall were not attempted; only the absence or presence of hydrate was recorded. With methane, a series of experiments was carried out at 14°C and at pressures below the methane hydrate equilibrium pressure, which is 118 arm at this temperature; a second series was carried out at 5°C where the equilibrium pressure is 44 atm (VILLARD, 1888). RESULTS
Figure 1 shows the results obtained with oxygen. At pressures whicll were up to 50% higher than the hydrate equilibrium pressure, the samples always remained in a metastable state, without any visible hydrate present. At higher pressures, from 200 to 328 atm, hydrate formed in seven of the ten samples, although most of the samples could be maintained in a metastable state in this pressure range as well. One sample at 300 atm and one at 314 atm remained metastable for 24 h, alter which time the experiments were terminated. Similar results were obtained with methane. From the hydrate equilibrium pressure of 44 (at 5°C) to 68 atm, hydrate rarely formed. However, at pressures higher than 75 arm, hydrate always formed. Figure 2 shows the methane gas flux from solutions in the presence and absence of methane hydrate. At 14°C, the gas flux increased proportionally with the pressure over the entire pressure range; that is, the flux directly reflects the gas tension in the water. This means that there was no change in the diffusion characteristics of the Teflon tube by, for instance, compression or partial collapse at the higher pressures. At 5°C, the gas flux remained proportional to the 0z-H20 , l°C(n ,I0)
I [ I00
u - no hydt01e • ; hydrote formed
,~ ( ) O O O O £ ) ( ~ ) q.)D OC)(~ O OOOOO©O L) g O O ( ~ @ Q ('Q . ) @ 0 @ ( 2 ) L ......... 150
]. . . . . . . . . . . . . . . .
200
L. . . . .
250
I ....... 300
C) J 350
Gos Pressure (olin) F i g . 1.
O c c u r r e n c e o f o x y g e n h y d r a t e ( d o s e d p o i n t s ) i n d i s t i l l e d w a t e r as a f u n c t i o n o f p r e s s u r e . The arrow indicates the hydrate equilibrium pressure. Temperature, P'C.
Clathrate hydrate of oxygen: does it occur in deep-sea fish?
5
Melhone_H2 0
4
j
147
14'
e= hydrofe present
I.== E
3
/
~t. ~_ 2
J
~..~o~t
~ ~'@°I
5*
... --'¢~Z"~
.
.~ "°"
" w~'
I
0 v 0 Fig. 2.
I
I 20
I
I I 40 Cos Pressure(elm)
I 60
I
I 80
Steady-state flux of m e t h a n e gas through Teflon barrier from water equilibrated with the
gas at various pressures. Open points: flux without hydrate visually present in the water. Closed points: with hydrate present. The hydrate at 52-58 atm was initially formed at 85 atm, after which the pressure was decreased. The arrow indicates the hydrate equilibrium pressure. pressure below the hydrate equilibrium pressure and in the metastable solutions without any hydrate present, indicating a normal activity and motility of the dissolved gas molecules. In contrast, with hydrate present, the gas flux remained constant and equal to the flux at the hydrate equilibrium pressure, irrespective of the gas pressure above the water. DISCUSSION
Solutions of oxygen in some cases may remain in a metastable state at pressures much higher than the hydrate equilibrium value, and in other cases under equivalent conditions hydrate readily forms. In addition, the results reveal that when the metastable state prevails, there are no major anomalies in the activity of the dissolved gas, such as might be the case if the gas molecules formed a liquid crystalline type structure with the water, analogous to what happens when the gas forms solid crystalline hydrate. The results have some implications for deep-sea fishes. They suggest that there is a range of ocean depths where oxygen remains in a metastable equilibrium with water present in the swimbladder as pools, droplets or a surface film, without the formation of any hydrate. If any secretion of oxygen is to take place at depths below that of the hydrate equilibrium pressure, it is necessary that the blood, and probably also the intracellular fluids of the secretory system, remain in a metastable state as well. The blood issupplied to the swimbladder gas secretion gland through a counter-current network of capillaries, the rete mirabile, in which the blood gas tensions presumably are built up from approximately atmospheric value* to values equal to, or slightly higher than, the partial pressures inside the swimbladder (ScHOLANDER, 1954; KUHN and KUHN, 1961 ; KUHN, MOSER and KUHN, 1962; STEEN, 1963; ENNS, DOUGLASand SCHOLANDER, 1967). The pressures that can be attained by the counter-current secretion mechanism, in principle, are limited only by the gas tensions possible in the blood. Because the gas tensions probably are maintained normally in the metastable blood, secretion of gas would take place under these conditions. Once the oxygen tension of *The gas tensions in the arterial blood offish may be assumed to be approximately equal to that of ambient water, which is similar to the atmospheric value at all depths except for a correction of a b o u t 1 4 ~ per 100 atm hydrostatic pressure (see ENNS, SCHOLANDER and BRADSTREET, 1965).
148
EDVARD A. HEMMINGSEN
the blood becomes limited by the formation of hydrate, the gas secretion will bc abolished for swimbladder oxygen pressures any higher than that of the hydrate equilibrium; that is, no secretion will occur at depths greater than about 1500-2000 m. Without secretion, the gas of the swimbladder slowly but inevitably will be lost. Similarly, once hydrate forms and remains in the swimbladder, the gas phase will be lost by its conversion to hydrate as the water required continues to enter by diffusion. The maximum depth at which gas secretion and the gas phase can be maintained is difficult to estimate, because substances that may impede the formation of hydrate and extend the range of metastability could conceivably exist in the blood or the swimbladder. The addition of solutes and other substances to water may increase the hydrate equilibrium pressure (ScAUZILLO, 1956; MILLER, 1961), but this effect would not be substantial. It is known that gases and other hydrate-forming substances may cooperate to lower the hydrate equilibrium value (PAuLI~G, 1961; JEFFREY and MCMULLAN, 1967), but not to increase it. Different types of interactions may arise from the presence of glycoproteins and other macromolecules. Evidence indicates that water around such molecules exists in a higher ordered state and that it may attain other properties than in its pure state (PAULING,1961 ; MILLER, 1961; JEFFREY,1969; KLOTZ, 1970; DROST-HANSEN, 1972). However, any such effects are likely only to increase the tendency for hydrate formation unless the pre-structured water is excluded from interacting with the gas molecules. If, as indicated by the present data for water, hydrate formation occurs consistently at pressures that are higher than about twice the equilibrium pressure, the swimbladder is unlikely to contain gas at depths below approximately 3500 m. Observations indicate that this may be so. Whereas specimens of Coryphaenoides abyssorum collected at about 3500 m or shallower depths surfaced with greatly expanded and ruptured swimbladders, 'popped' eyes, and other evidence for the presence of substantial quantities of gas (DOUGLAS, 1967; C. L. Hubbs, personal communication) and/or gas hydrate at the original depths, none of several specimens collected at about 4000m showed any such signs of containing gas (C. L. Hubbs, personal communication). It is also to be noted that the swimbladder of the one specimen of Bassogigas profundissimus collected at 7100 m was not ruptured or abnormally expanded (NIELSEN and MUNK, 1964), and probably, therefore, was gas-free. Factors other than hydrate formation also may greatly affect the gas secretion and the function of the swimbladder at great depths. At such high pressures, the buoyancy efficiency of the swimbladder gas is much decreased; for instance, at 300 atm, the density of oxygen is about 0.40 g m1-1. Therefore, the gas-secretion process in fish at great depths, as compared to that at shallower depths, must compensate not only for the volume decrease due to compression of the gas, but also for the additional volume required because of increased gas density. Because the maximum rate at which gas can be secreted by the counter-current mechanism is limited by physical factors (ENNS, DOUGLAS and SCHOLANDER, 1967), fish with gas-filled swimbladders below certain depths may be unable to secrete adequate quantities of gas for buoyancy adjustments even at slow rates of vertical migration. One solution could alleviate this problem. Fish normally have a limited vertical migration; therefore, only a small fraction of the total swimbladder gas volume is involved in the buoyancy regulation. Thus, if the 'unused' gas volume is replaced with a substance of favorable buoyancy and little compressibility, the quantities of gas secreted or reabsorbed would be greatly
Clathrate hydrate of oxygen: does it occur in deep-sea fish .9
149
reduced. Indeed, the lipid substances which are typically present in the swimbladder o f deep-sea fish (PHLEGER and BENSON, 1971) m a y serve such a function. However, other functions o f this substance m a y be imagined. Notably, it m a y provide the m i n i m u m b u o y a n c y necessary for survival during periods when the gas is lost accidentally due to hydrate formation and refilling is taking place; or, it m a y somehow act to impede or minimize hydrate formation in the gas phase.
Acknowledgements--This investigation was supported by grant No. 1 K04 HL-50263-02 from the U.S. Public Health Service. REFERENCES DOUGLASE. L. (1967) Studies of gas secretion in the swimhladder of fishes. Ph.D. Dissertation, University of California, San Diego, pp. 1-134. DROSToHANSENW. (1972) Effects of pressure on the structure of water in various aqueous systems. In: The effects of pressure on organisms, M. A. SLEIGHand A. G. MACDONALD, editors, Academic Press, pp. 61-101. ENNS T., E. DOUGLASand P. F. SCHOLANDER(1967) Role of the swimbladder fete of fish in secretion of inert gas and oxygen. Advances in Biological and MedicalPhysics, 11, 231-244. ENNS T., P. F. SCHOLANDERand E. D. BRADSTREET(1965) Effect of hydrostatic pressure on gases dissolved in water. Journal of Physical Chemistry, 69, 389-391. JEFFREY G. A. (1969) Water structure in organic hydrates. Accounts of Chemical Research, 2, 344-352. JEFFREY G. A. and R. K. McMULLAN (1967) The clathrate hydrates. In: Progress in inorganic chemistry, F. A. COTTON, editor, Interscience, Voi. 8, pp. 43-108. KLOTZ I. M. (1970) Polyhedral clathrate hydrates. In; The frozen cell, G. E. W. WOLSTENHOLME and M. O'CONNOR, editors, Churchill, pp. 5-26. KUHN H. J., P. MOSER and W. KUHN (1962) Haarnadelgegenstrom als Grundlage zur Erzeugung hoher Gasdriicke in der Schwimmblase yon Tiefseefischen. Pfliigers Archly fiir die gesamte Physiologie, 275, 231-237. KUHN W. and H. J. KUHN (1961) Multiplikation yon Aussaltz- und anderen Einzeleffekten fiJr die Bereitung hoher Gasdriike in der Schwimmblase. Zeitschrift fiir Elektrochemie, 65, 426-439. MILLER S. L. (1961) A theory of gaseous anesthesia. Proceedings of the National Academy of Sciences of the United States of America, 47, 1515-1524. MILLER S. L. (1969) Clathrate hydrates of air in Antarctic ice. Science, 165, 489-490. NIELSENJ. G. and O. MUNK (1964) A haddal fish (Bassogigas profundissimus) with a functional swimbladder. Nature, 204, 594-595. PAULINGL. (1961) A molecular theory of general anesthesia. Science, 134, 15-21. PHLEGER C. F. and A. A. BENSON(1971) Cholesterol and hyperbaric oxygen in swimbladders of deep sea fishes. Nature, 230, 122. SCAUZILLO F. R. (1956) Inhibiting hydrate formations in hydrocarbon gases. Chemical Engineering Progress, 52, 324-328. SCHOLANDERP. F. (1954) Secretion of gases against high pressures in the swimbladder of deep sea fishes. II. The fete mirabile. Biological Bulletin, Woods Hole, 107, 260-277. SCHOLANDER P. F. and L. VAN DAM (1953) Composition of the swimbladder gas in deep sea fishes. Biological Bulletin, Woods Hole, 104, 75-86. STEEN J. B. (1963) The physiology of the swimbladder in the eel Anguilla vulgar&. III. The mechanism of gas secretion. Acta physiologica scandinavica, 59, 221-241. VAN CLEEEA. and G. A. M. DIEPEN (1965) Gas hydrates of nitrogen and oxygen. II. Recueil des travaux chimiques des Pays-Bas et de la Belgique, 84, 1085-1093. VIt LARD M. (1888) Sur les hydrates de mrthane et d'rthyl~ne. Compte rendu hebdomadaire des s~ances de l'Acad~mie des sciences, 107, 395-397.
Clathrate hydrate of oxygen: does it occur in deep-sea fish? EDVARD A. HEMMINGSEN*
(Received 20 May 1974; accepted5 October 1974) Abslraet--Oxygen exists in the swimbladder of some deep-sea fish at partial pressures that are more than twice the hydrate equilibrium pressure in water. Observations suggest that no hydrate forms to depths of 3500-4000 m. At greater depths, gas secretion may not occur because of the formation of hydrate. INTRODUCTION THE GAS-FILLEDswimbladder of fish functions as a buoyancy organ, the volume of which is regulated by secretion or reabsorption of gas. Because the gas pressure in the swimbladder essentially equals the hydrostatic pressure acting on the fish, gas pressures of several hundred atmospheres exist in many deep-sea fish. About 90%, or more, of this swimbladder gas is oxygen; the remainder is mainly nitrogen and some carbon dioxide (SCHOLANDERand VAN DAM, 1953; SCHOLANDER,1954; DOUGLAS,1967). The deepest samples have been obtained from specimens of Coryphaenoides abyssorum collected at 3512 m, in which case the oxygen content was equivalent to a partial pressure of about 320 atm (DOUGLAS, 1967). Even higher pressures may exist; two species of brotulid fish with well-developed swimbladders have been collected at depths from 4575 to 7160 m (NIELSENand MUNK, 1964). Many gases, including oxygen, form solid hydrates in aqueous solutions. These hydrates are ice-like crystalline clathrate compounds in which the gas molecules are enclosed in a hydrogen-bounded water lattice (JEFFREY and MCMULLAN, 1967; KLOTZ, 1970). For water at 2°C, a typical deep-sea temperature, the oxygen hydrate equilibrium, or dissociation pressure, is about 148 atm (VANCLEEFFand DIEPEN, 1965). Because the oxygen pressures in the swimbladder reach values of more than twice the hydrate equilibrium pressure, conditions for the formation of oxygen hydrate exist in these fish. Such a natural occurrence of gas hydrate, the possibility of which appears to have been overlooked in previous swimbladder studies, would be unique not only for biological systems, but also for the Earth as well, with the possible exception of gas hydrates deep in the polar ice caps (MILLER, 1969). The formation of oxygen hydrate in the swimbladder or its tissue could have major effects on gas secretion, on the maintenance of a gas phase, and possibly on the survival of the fish. One question is whether or not gas dissolved in water at pressures substantially above the gas hydrate equilibrium value can be maintained in a metastable state without formation of crystalline gas hydrate. An experimental system was set up to determine to what degree such metastability may prevail with respect to oxygen. It was also of interest to determine the gas tension in the solution at gas pressures higher than the hydrate equilibrium value, and this was accomplished indirectly by measuring the diffusion of dissolved gas from the solution through a Teflon barrier, using methane, which has a lower hydrate equilibrium pressure. *Physiological Research Laboratory, Scripps Institution of Oceanography, La Julia, California 92037, U.S.A. 145
146
EDVARD A . HEMMINGSEN
METHODS
The apparatus and the experimental procedures were as follows: within a pressure chamber, distilled water in a glass beaker was stirred with a magnetic bar and was equilibrated with oxygen or methane gas at various pressures. Each water sample usually was equilibrated at several pressures, with stepwise increases in the pressures from low to high values. Each equilibration lasted at least 2 h. The water in the chamber could be observed through a window for visual evidence of crystalline hydrate formation. A coiled Teflon tube (0.041 cm i.d., 0.085 cm o.d., and I 1 cm long) was submerged completely in the water. One end of the tube was sealed, and the other end was brought to the outside via a cut-off hypodermic needle (Gauge 26). The gas flux from the water through the Teflon barrier was measured by means 0fthe movement of a meniscus in a capillary tube attached to the hypodermic needle outlet. The temperature of the apparatus was controlled to ~0.02°C by submersion in a regulated water bath. The experiments with oxygen were carried out at l~C. Because of partial or complete collapse of the Teflon tube at pressures greater than about 100 atm, measurements of the oxygen flux through the tube wall were not attempted; only the absence or presence of hydrate was recorded. With methane, a series of experiments was carried out at 14°C and at pressures below the methane hydrate equilibrium pressure, which is 118 arm at this temperature; a second series was carried out at 5°C where the equilibrium pressure is 44 atm (VILLARD, 1888). RESULTS
Figure 1 shows the results obtained with oxygen. At pressures whicll were up to 50% higher than the hydrate equilibrium pressure, the samples always remained in a metastable state, without any visible hydrate present. At higher pressures, from 200 to 328 atm, hydrate formed in seven of the ten samples, although most of the samples could be maintained in a metastable state in this pressure range as well. One sample at 300 atm and one at 314 atm remained metastable for 24 h, alter which time the experiments were terminated. Similar results were obtained with methane. From the hydrate equilibrium pressure of 44 (at 5°C) to 68 atm, hydrate rarely formed. However, at pressures higher than 75 arm, hydrate always formed. Figure 2 shows the methane gas flux from solutions in the presence and absence of methane hydrate. At 14°C, the gas flux increased proportionally with the pressure over the entire pressure range; that is, the flux directly reflects the gas tension in the water. This means that there was no change in the diffusion characteristics of the Teflon tube by, for instance, compression or partial collapse at the higher pressures. At 5°C, the gas flux remained proportional to the 0z-H20 , l°C(n ,I0)
I [ I00
u - no hydt01e • ; hydrote formed
,~ ( ) O O O O £ ) ( ~ ) q.)D OC)(~ O OOOOO©O L) g O O ( ~ @ Q ('Q . ) @ 0 @ ( 2 ) L ......... 150
]. . . . . . . . . . . . . . . .
200
L. . . . .
250
I ....... 300
C) J 350
Gos Pressure (olin) F i g . 1.
O c c u r r e n c e o f o x y g e n h y d r a t e ( d o s e d p o i n t s ) i n d i s t i l l e d w a t e r as a f u n c t i o n o f p r e s s u r e . The arrow indicates the hydrate equilibrium pressure. Temperature, P'C.
Clathrate hydrate of oxygen: does it occur in deep-sea fish?
5
Melhone_H2 0
4
j
147
14'
e= hydrofe present
I.== E
3
/
~t. ~_ 2
J
~..~o~t
~ ~'@°I
5*
... --'¢~Z"~
.
.~ "°"
" w~'
I
0 v 0 Fig. 2.
I
I 20
I
I I 40 Cos Pressure(elm)
I 60
I
I 80
Steady-state flux of m e t h a n e gas through Teflon barrier from water equilibrated with the
gas at various pressures. Open points: flux without hydrate visually present in the water. Closed points: with hydrate present. The hydrate at 52-58 atm was initially formed at 85 atm, after which the pressure was decreased. The arrow indicates the hydrate equilibrium pressure. pressure below the hydrate equilibrium pressure and in the metastable solutions without any hydrate present, indicating a normal activity and motility of the dissolved gas molecules. In contrast, with hydrate present, the gas flux remained constant and equal to the flux at the hydrate equilibrium pressure, irrespective of the gas pressure above the water. DISCUSSION
Solutions of oxygen in some cases may remain in a metastable state at pressures much higher than the hydrate equilibrium value, and in other cases under equivalent conditions hydrate readily forms. In addition, the results reveal that when the metastable state prevails, there are no major anomalies in the activity of the dissolved gas, such as might be the case if the gas molecules formed a liquid crystalline type structure with the water, analogous to what happens when the gas forms solid crystalline hydrate. The results have some implications for deep-sea fishes. They suggest that there is a range of ocean depths where oxygen remains in a metastable equilibrium with water present in the swimbladder as pools, droplets or a surface film, without the formation of any hydrate. If any secretion of oxygen is to take place at depths below that of the hydrate equilibrium pressure, it is necessary that the blood, and probably also the intracellular fluids of the secretory system, remain in a metastable state as well. The blood issupplied to the swimbladder gas secretion gland through a counter-current network of capillaries, the rete mirabile, in which the blood gas tensions presumably are built up from approximately atmospheric value* to values equal to, or slightly higher than, the partial pressures inside the swimbladder (ScHOLANDER, 1954; KUHN and KUHN, 1961 ; KUHN, MOSER and KUHN, 1962; STEEN, 1963; ENNS, DOUGLASand SCHOLANDER, 1967). The pressures that can be attained by the counter-current secretion mechanism, in principle, are limited only by the gas tensions possible in the blood. Because the gas tensions probably are maintained normally in the metastable blood, secretion of gas would take place under these conditions. Once the oxygen tension of *The gas tensions in the arterial blood offish may be assumed to be approximately equal to that of ambient water, which is similar to the atmospheric value at all depths except for a correction of a b o u t 1 4 ~ per 100 atm hydrostatic pressure (see ENNS, SCHOLANDER and BRADSTREET, 1965).
148
EDVARD A. HEMMINGSEN
the blood becomes limited by the formation of hydrate, the gas secretion will bc abolished for swimbladder oxygen pressures any higher than that of the hydrate equilibrium; that is, no secretion will occur at depths greater than about 1500-2000 m. Without secretion, the gas of the swimbladder slowly but inevitably will be lost. Similarly, once hydrate forms and remains in the swimbladder, the gas phase will be lost by its conversion to hydrate as the water required continues to enter by diffusion. The maximum depth at which gas secretion and the gas phase can be maintained is difficult to estimate, because substances that may impede the formation of hydrate and extend the range of metastability could conceivably exist in the blood or the swimbladder. The addition of solutes and other substances to water may increase the hydrate equilibrium pressure (ScAUZILLO, 1956; MILLER, 1961), but this effect would not be substantial. It is known that gases and other hydrate-forming substances may cooperate to lower the hydrate equilibrium value (PAuLI~G, 1961; JEFFREY and MCMULLAN, 1967), but not to increase it. Different types of interactions may arise from the presence of glycoproteins and other macromolecules. Evidence indicates that water around such molecules exists in a higher ordered state and that it may attain other properties than in its pure state (PAULING,1961 ; MILLER, 1961; JEFFREY,1969; KLOTZ, 1970; DROST-HANSEN, 1972). However, any such effects are likely only to increase the tendency for hydrate formation unless the pre-structured water is excluded from interacting with the gas molecules. If, as indicated by the present data for water, hydrate formation occurs consistently at pressures that are higher than about twice the equilibrium pressure, the swimbladder is unlikely to contain gas at depths below approximately 3500 m. Observations indicate that this may be so. Whereas specimens of Coryphaenoides abyssorum collected at about 3500 m or shallower depths surfaced with greatly expanded and ruptured swimbladders, 'popped' eyes, and other evidence for the presence of substantial quantities of gas (DOUGLAS, 1967; C. L. Hubbs, personal communication) and/or gas hydrate at the original depths, none of several specimens collected at about 4000m showed any such signs of containing gas (C. L. Hubbs, personal communication). It is also to be noted that the swimbladder of the one specimen of Bassogigas profundissimus collected at 7100 m was not ruptured or abnormally expanded (NIELSEN and MUNK, 1964), and probably, therefore, was gas-free. Factors other than hydrate formation also may greatly affect the gas secretion and the function of the swimbladder at great depths. At such high pressures, the buoyancy efficiency of the swimbladder gas is much decreased; for instance, at 300 atm, the density of oxygen is about 0.40 g m1-1. Therefore, the gas-secretion process in fish at great depths, as compared to that at shallower depths, must compensate not only for the volume decrease due to compression of the gas, but also for the additional volume required because of increased gas density. Because the maximum rate at which gas can be secreted by the counter-current mechanism is limited by physical factors (ENNS, DOUGLAS and SCHOLANDER, 1967), fish with gas-filled swimbladders below certain depths may be unable to secrete adequate quantities of gas for buoyancy adjustments even at slow rates of vertical migration. One solution could alleviate this problem. Fish normally have a limited vertical migration; therefore, only a small fraction of the total swimbladder gas volume is involved in the buoyancy regulation. Thus, if the 'unused' gas volume is replaced with a substance of favorable buoyancy and little compressibility, the quantities of gas secreted or reabsorbed would be greatly
Clathrate hydrate of oxygen: does it occur in deep-sea fish .9
149
reduced. Indeed, the lipid substances which are typically present in the swimbladder o f deep-sea fish (PHLEGER and BENSON, 1971) m a y serve such a function. However, other functions o f this substance m a y be imagined. Notably, it m a y provide the m i n i m u m b u o y a n c y necessary for survival during periods when the gas is lost accidentally due to hydrate formation and refilling is taking place; or, it m a y somehow act to impede or minimize hydrate formation in the gas phase.
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