Download Dissolved Organic Matter in Oceanic Waters

Journal of Oceanography, Vol. 59, pp. 129 to 147, 2003
Review
Dissolved Organic Matter in Oceanic Waters
H IROSHI OGAWA1* and EIICHIRO TANOUE2
1
2
Ocean Research Institute, University of Tokyo, Minamidai, Nakano-ku, Tokyo 164-8639, Japan
Graduate School of Environmental Studies, Nagoya University,
Furo-cho, Chigusa-ku, Nagoya 464-8601, Japan
(Received 12 February 2002; in revised form 3 July 2002; accepted 3 July 2002)
The amount of information on oceanic dissolved organic matter (DOM) has increased
dramatically in the last decade thanks to the advances in chemical characterization.
This information has supported the development of some novel and important ideas
for DOM dynamics in the ocean. Consequently, we have a better understanding of
the importance of DOM in oceanic biogeochemical cycles. Here we review studies
published mainly during 1995–2001, synthesize them and discuss unsolved problems
and future challenges. The measurement, distribution and turnover of dissolved organic carbon (DOC) are presented as the bulk dynamics of the oceanic DOM. The
size spectrum, elemental composition, and chemical compositions at molecular and
functional group levels are described. The mechanisms proposed for the survival of
biomolecules in DOM are discussed.
1. Introduction
Dissolved organic matter (DOM) in the sea is one of
the largest reservoirs of organic matter on the earth’s surface (others include soil organic matter and plant biomass
on land), holding approximately as much carbon as is
available in atmospheric carbon dioxide (Hedges, 1992).
The fact that DOM is a huge organic reservoir on the
earth’s surface has continued to impel ocean scientists to
investigate what DOM is, in terms of its source, chemical nature and function in marine environments, from
early in the 20th century.
Since homeostatic feedback among the reservoirs of
bioelements has controlled the past global environments
over the geological time scale (e.g., Berner, 1989), the
role and dynamics of dissolved organic carbon (DOC)
have become of greater interest in the present global carbon cycle (e.g., Siegenthaler and Sarmiento, 1993). Recognition of the importance of the microbial loop has also
has given us new insight into the role of DOM in marine
ecosystems (e.g., Pomeroy, 1974; Azam et al., 1983).
Primary production is the ultimate source of organic matter in the sea, but living biomass forms less than 1% of
total organic carbon in seawater, while more than 90% of
Keywords:
⋅ Dissolved organic
matter (DOM),
⋅ dissolved organic
carbon (DOC),
⋅ carbon cycle,
⋅ molecular weight
distribution,
⋅ C:N ratio,
⋅ chemical composition,
⋅ biomolecules.
organic carbon occurs as non-living DOC (e.g., Cauwet,
1979). In contrast to the organic reservoirs on land, the
processes by which DOM has been formed are unclear,
and actual sources and the chemical nature of DOM are
not well known.
DOM is still the least understood organic reservoir
on the earth, but our knowledge of DOM has been rapidly increasing. Recent advancements in DOM study have
been covered by reviews of the results of various approaches, trying to understand the dynamics and chemical nature of DOM (Trumbore and Druffel, 1995; Guo
and Santschi, 1997; Nagata, 2000; Williams, 2000;
Kepkay, 2000; Myklestad, 2000; Ogawa, 2000; Benner,
2002; Hansell and Carlson, 2002; Hedges, 2002). Some
fundamental new insights and observations have been
booked recently, which we have summarized in this review. We thus focus here on papers published since 1995,
although references to earlier fundamental work are included.
DOM has various functions and plays important roles
in chemical, biological and even physical oceanography.
For example, DOM interacts with trace metals or
radionuclides and controls their dynamics, it fuels the
microbial loop, generates gases (CO, CO2) and nutrients
with biological and photochemical reactions, absorbs and
extinguishes light, and affects satellite images, etc. The
terrestrial input of DOM is also an important topic in the
* Corresponding author. E-mail: [email protected]
Copyright © The Oceanographic Society of Japan.
129
global carbon budget as well as carbon dynamics in
coastal environments. In this review, however, we focus
on recent advances in our understanding of the dynamics
and chemical characterization of oceanic DOM. Functions
of DOM in the environment are not described.
2. What is DOM?
The first difficulty that one must confront in any
definition of DOM is to define the term “dissolved”. It
usually has an operational definition as any material that
passes through a given filter is termed “dissolved”. Glass
fiber filters (Whatman GF/F, with a nominal pore size of
0.7 µm) are widely used for the collection and analysis
of organic substances. However, vastly different filter
types have been used for study purposes. In reality, a spectrum of material sizes exists; some material, such as colloidal matter, does not fit neatly into either category.
Small-sized plankton pass through the filter; for example, 22–38% of the total bacterial biomass (Lee et al.,
1995) and 100% of viruses, if they occur alone, are not
retained on the GF/F filter. We have to be aware that “dissolved organic matter” does not exactly indicate the true
“dissolved phase” of organic matter in the sea.
3.
Distribution and Turnover Time of DOC in the
Ocean
3.1 Methodology of DOC measurement
The abundance of DOM has generally been determined as dissolved organic carbon (DOC), which is a
major element of organic matter. A controversy about the
measurement of DOC concentrations in seawater was
current during the late 1980s and into the early 1990s.
The high temperature (catalytic) combustion (HTC)
method introduced by Sugimura and Suzuki (1988) led
to an argument about the existence of a much larger reservoir of DOC in the ocean than had been estimated by
the conventional method based on wet chemical oxidation (WCO, Menzel and Vaccaro, 1964). This problem
excited many ocean scientists in the world because it had
the potential to drastically change our concept of the oceanic carbon cycle (Williams, 1992). After 1993, the controversy calmed through further examination of the HTC
technique by a few communities (Hedges et al., 1993;
Sharp et al., 1993, 1995) and independent groups (Ogawa
and Ogura, 1992; Tanoue, 1992; Cauwet, 1994). It was
found that the HTC measurement included a potentially
high system blank relative to the DOC level in seawater
(e.g., Williams, 1992; Benner and Strom, 1993). In conclusion, the DOC values obtained from the HTC method
in the early stage were retracted (Suzuki, 1993) and the
concept of “new” DOC that might have represented a huge
oceanic carbon reservoir, missed by the WCO method,
was rejected.
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H. Ogawa and E. Tanoue
Thereafter, the HTC method was reassessed, improved, and reborn as a highly precise method, suitable
for measuring the seawater DOC (Sharp, 1997). In addition, the international distribution of certified reference
materials (CRMs) for DOC analyses, including low carbon water and deep seawater which were distributed by
Dr. Jonathan Sharp in 1997 and Dr. Dennis Hansell since
1999, means that it has become possible to more correctly
compare DOC values measured by different workers.
Consequently, both the accuracy and the precision of DOC
data have been improved. Details of DOC measurement
can be found in the recent review by Sharp (2002).
A typical range of error of the DOC measurement by
the recent HTC technique is 1–2% as a relative value,
i.e., coefficient of variation (c.v.), or 0.5–1 µM as an absolute value, i.e., standard deviation (s.d.) (Qian and
Mopper, 1996; Hansell et al., 1997b; Ogawa et al., 1999).
This is approximately 1/10 the error of the traditional
WCO method. The improved precision of the DOC
method contributed greatly to the resolution of small differences in DOC concentrations in seawater in time and
space, although the previous method had been able to
detect part of them qualitatively. This improvement was
essential for the advancement of oceanic DOC study since
we had to resolve the active but small pool of DOC relative to the major part of DOC that was fairly refractory in
seawater (see below). The introduction of this improved
HTC method supplied quantitative and detailed information for an understanding of the role of DOC in the oceanic carbon cycle.
3.2 Distribution, accumulation and export of DOC in the
upper water column
Since the WCO method was established (Menzel and
Vaccaro, 1964), it has been consistently observed that
elevated concentrations of DOC appear in the upper water column, decreasing with increasing depth, followed
by a low, uniform value in the deep water (e.g., Barber,
1968; Ogura, 1970). This finding suggests that DOC is
supplied by biological production in the surface ocean
and consumed by microbial respiration, and the refractory component resistant to microbial degradation remains
in the deep water (Barber, 1968). In addition, recent observations using the high-precision HTC method were
successful in detecting the detailed distribution and fluctuation of DOC within the surface water column. The
observed changes in the surface DOC appear to be mainly
caused by hydrological transport combined with biological production since they were observed over different
productive sites along with vertical convection or horizontal advection of seawater (Copin-Montégut and Avril,
1993; Carlson et al., 1994; Peltzer and Hayward, 1996;
Hansell and Carlson, 2001a, b). The apparent turnover of
DOC that accumulated and disappeared in the surface
Fig. 1. Typical vertical distributions of DOC in the ocean according to a: conceptual classification of biological reactivity, b: size
distribution and c: chemical composition. At each depth the concentration of bulk DOC is representative of the oceanic value
reported after 1994, mainly including equatorial, subtropical and temperate zones (see Table 1), without the Antarctic and the
Arctic oceans. In a, the semi-labile DOC is expressed as the surface DOC in excess of that in deep water, i.e., the refractory
DOC (=~40 µ M), as proposed by Carlson and Ducklow (1995). The depth profile of fluorescence of humic-like materials (Ex:
320 or 350 nm, Em: 450 nm) simply indicates its relative pattern in typical oceanic sites of the Sargasso Sea and the North
Pacific (Mopper et al., 1991; Chen and Bada, 1992). In b, DOC is separated into the three size classes; very high molecular
weight (VHMW; >10 kDa), high molecular weight (HMW; 1–10 kDa) and low molecular weight (LMW; <1 kDa), obtained
by the ultrafiltration method (Ogawa and Ogura, 1992; Benner et al., 1997). Here, the typical bulk DOC concentrations of the
oceanic water column are the same as shown in a and the mean size distribution at six depths (<20, 100, 150–200, 400, 750–
1000, 4000 m) from the Pacific Ocean, the Atlantic Ocean and Gulf of Mexico are shown (Ogawa and Ogura, 1992; Guo et al.,
1995; Benner et al., 1997; Ogawa, unpublished data). In c, DOC is classified into the three chemical components; total
hydrolyzable amino acids (THAA) determined by HPLC method (Lindroth and Mopper, 1979), after acid hydrolysis with 6N
HCl (Robertson et al., 1987), or a vapor phase hydrolysis (Tsugita et al., 1987), total hydrolyzable neutral sugars (THNS)
determined by HPLC method after acid hydrolysis with 1.2M H2SO4 (Skoog and Benner, 1997), and total carbohydrate by the
MBTH method (MBTH-TCHO), which is a colorimetric technique after hydrolysis with 1.2M H2SO4 (Pakulski and Benner,
1992). The TCHO component other than THNS is illustrated here because THNS should be included in TCHO. The concentrations as carbon in these components were estimated by multiplying the bulk DOC concentrations as shown in a by their
mean yields of DOC, which were obtained from different depths of the Pacific and Atlantic Oceans, i.e., THAA-C/DOC
(Druffel et al., 1992; Ogawa, unpublished data), THNS-C/DOC (Skoog and Benner, 1997) and TCHO-C/DOC (Pakulski and
Benner, 1994). The remainder, given by subtracting (TCHO+THAA)-C from the bulk DOC concentrations, is called the
“uncharacterized” component of DOC here. Thus, note that the bulk DOC concentrations, the size distribution in b and the
chemical composition in c simply indicate their representative values and profiles in the ocean.
water column was occurring on time scales of months to
a year (Carlson et al., 1994). This turnover time is longer
than that of “labile DOC” including free amino acids,
sugars and organic acids which turns over in minutes to
days through rapid bacterial uptake, while it is shorter
than that of “refractory DOC” in the deep ocean, which
is quite resistant to microbial oxidation, with average ages
of hundreds to thousands of years. Therefore, this fraction of DOC is called “semi-labile DOC” as distinguished
from the labile and refractory DOC (Kirchman et al.,
1993; Carlson and Ducklow, 1995; Cherrier et al., 1996).
Assuming that the uniform concentration of the refractory DOC in deep water is extended to the whole water
column, the concentrations of the semi-labile DOC would
be given by the excess of surface DOC over that in deep
water (Fig. 1a). According to recent reports, the concentration of deep water DOC is uniformly about 40 µM,
while DOC in surface waters is 60–80 µM in a large area
of the ocean, except for the Antarctic Ocean where DOC
was restrained in low values (<ca. 60 µM, Table 1). There-
Dissolved Organic Matter in Oceanic Waters
131
Table 1. Oceanic DOC (TOC) concentrations measured by the HTC method after 1994.
Study area
Latitude
DOC (µM)
Literature(c)
Note
Surface(a)
Deep (b )
75°N
48°N
33–60°N
32°N
33–36°N
36°N
1°S–3°N
—
—
50–70
64–71
68–71
70–80
65–97
48 (av.)
45 (av.)
44
44
—
46–48
33–59*
1
1
2
1, 3, 4
5
6, 7
8
Pacific Ocean
equatorial
central equatorial, 140°W
central equatorial, 140°W
central equatorial, 140/135°W
western equatorial, 170°E–150°W
southwest, 170°W
eastern, 103/110°W
northern North
0°
12°S–12°N
12/2°S
12°S–5°N
42°S–0°
67°S–23°N
23/58°N
60–70
63–67
60–80
72–82
65–70
65–90
40–85
—
35–40
39.5 (av.)
36 (av.)
44–45
—
43 (av.)
—
34–39 (av.)
9
10
11
12
13
1, 14
15
1
Indian Ocean & Arabian Sea
central, 80/90°E
Arabian Sea
43°S–5°N
10–20°N
55–80
65–95
43 (av.)
43 (av.)
1, 14
1, 16
TOC
TOC
Antarctic Ocean
Weddell Sea
Ross Sea
Australian sector
Indian sector, 62°E
Antarctic Pacific 170°E/170°W
6°W
71–63°S
77°S
65–56°S
66–49°S
67–42°S
60–47°S
45–60
43–55 (av.)
45–55
52–63
45–75
38–55
40–60
—
40–45
42 (av.)
42 (av.)
34–38
17
4, 18
19
20
1, 14
21
TOC
Arctic Ocean
Greenland
central
central
central
74–81°N
80–85°N
70–85°N
70–85°N
66–80
78 (av.)
63–139
34–107 (av.)
49–54
53 (av.)
—
—
22
23
24
25
Atlantic Ocean
Greenland Sea
13/48°W
northeast, 20°W
Sargasso Sea (Bermuda)
Sargasso Sea (Bermuda)
Middle Atlantic Bight
equatorial
TOC
TOC
TOC
TOC
TOC
TOC
TOC
TOC
(a)
Surface mixed layer or <100 m. The data from coastal areas were excluded.
>1000 m, * exceptionally high values were excluded.
(c)
1. Hansell and Carlson, 1998a; 2. Kähler and Koeve, 2001; 3. Carlson et al., 1994; 4. Carlson et al., 1998; 5. Bates and
Hansell, 1999; 6. Guo et al., 1995; 7. Guo et al., 1996; 8. Thomas et al., 1995; 9. Sharp et al., 1995; 10. Carlson and Ducklow,
1995; 11. Peltzer and Hayward, 1996; 12. Skoog and Benner, 1997; 13. Hansell et al., 1997b; 14. Doval and Hansell, 2000;
15. Hansell and Waterhouse, 1997; 16. Hansell and Peltzer, 1998; 17. Wedborg et al., 1998; 18. Carlson et al., 2000; 19. Ogawa
et al., 1999; 20. Wiebinga and de Baar, 1998; 21. Kähler et al., 1997; 22. Opsahl et al., 1999; 23. Bussmann and Kattner, 2000;
24. Wheeler et al., 1997; 25. Wheeler et al., 1998.
(b)
fore, 20–40 µ M of DOC is estimated as making up the
semi-labile fraction accumulating in the surface waters.
Globally applying the representative DOC distribution
from tropical, subtropical and temperate zones (Fig. 1a
based on the data of literatures given in Table 1), the glo-
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H. Ogawa and E. Tanoue
bal carbon stock of the bulk DOC in the ocean would be
approximately 700 Gt. This is close to the estimate given
by Hansell and Carlson (680 Gt; 1998a). It consists of
650 and 50 Gt for the refractory and the semi-labile DOC,
respectively. The semi-labile DOC is a quantitatively
minor portion (<10% of the bulk DOC) of the carbon stock
in the whole ocean, but it is important for carbon flux.
Hansell and Carlson (1998b) estimated that the global net
production of semi-labile DOC was 17% of global new
production (=1.2 Gt y–1).
The hydrological current in the ocean is accompanied by the transportation of dissolved substances in
seawater. The movement of surface water mass, which is
caused by the seasonal vertical convection or the horizontal advection driven by winds, in particular results in
the export of the semi-labile DOC which is accumulated
due to the vertical stability. Most previous studies on the
export of primary production from the euphotic zone, i.e.,
export production, have focused on the pathway due to
the settling of large particles (POC) using sediment traps
(Eppley and Peterson, 1979). By contrast, it was difficult
to evaluate the role of DOC due to its dilute concentrations in seawater. Quantitative evaluations of the export
production of DOC were keenly pursued in the late 1990s.
Consequently, some studies estimated a substantial portion of the export production in the ocean would be occurring as DOC, comparable to or exceeding that for sinking particles (Copin-Montégut and Avril, 1993; Carlson
et al., 1994; Murray et al., 1994; Peltzer and Hayward,
1996; Libby and Wheeler, 1997; Emerson et al., 1997).
This finding was accepted positively because it was consistent with the hypothesis that DOC export would be
responsible for the carbon imbalance between the export
production by sinking particles and new production based
on nitrate consumption. However, recent studies, especially from the equatorial Pacific, indicated the opposite
case that the vertical flux of DOC was unlikely to account for significant export production, and POC flux
should be more important (Murray et al., 1996; Hansell
et al., 1997a; Hansell and Peltzer, 1998; Doval and
Hansell, 2000). The relative importance of DOC and POC
fluxes appears to depend largely upon both physical and
biological features in the study sites. Not only highly precise measurements of DOC but also more detailed field
observation would therefore lead to more realistic evaluations of the role of DOC export in the oceanic carbon
cycle. The latest knowledge on DOC export in the ocean
is discussed by Hansell and Carlson (2001a, b) and
Hansell et al. (2002).
Recent studies have also provided the distribution
of DOC in seawater on the global ocean scale. As summarized in Table 1, a number of reports of DOC distributions from a wide variety of regions in the world oceans
were concentrated in a relatively brief period after the
establishment of the improved HTC method (approximately after 1994). Some of these studies measured TOC
instead of DOC when POC concentrations were insignificant, so that the potential contamination of the filtration
procedure could be avoided. A large variation of DOC
was observed in the surface waters (generally within the
range of 45–90 µM) compared with that in the deep waters (35–50 µM), indicating the regional difference in the
accumulation of semi-labile DOC. In general, the surface
concentrations of DOC changed more depending on the
latitudinal than the meridional direction, in the order of
the Arctic (70–100 µM) > subtropical (~80 µ M) > tropical (equatorial) and temperate (60–70 µ M) > subarctic
and subantarctic (50–60 µM) > Antarctic (40–60 µM). It
is necessary to consider that some of the high concentrations of DOC in the Arctic Ocean are exceptional for the
oceanic environments because they might be influenced
by riverine inputs (Bussmann and Kattner, 2000), even
though in situ biological production might also contribute to this (Wheeler et al., 1997). With the exception of
the Arctic Ocean, the hydrological properties appeared
to be a major factor controlling the regional difference in
the surface DOC concentrations in the ocean (Hansell and
Waterhouse, 1997; Doval and Hansell, 2000; Hansell and
Carlson, 2001a, b). In the equatorial area and the high
latitude including the Antarctic Ocean where the deep
water penetrates into the surface by vertical mixing and
upwelling, the DOC concentrations are maintained at a
relatively low level (ca. 40–60 µM). By contrast, elevated
concentrations of DOC (~80 µM) prevail in the subtropical area where the strong vertical stratification restricts
the vertical mixing of the surface water with the deep
water. However, this latitudinal pattern suggests that in
situ biological productivity would not be a unique factor
controlling the DOC concentrations in the surface waters; the subtropical area, a typical oligotrophic ocean,
shows higher concentrations of DOC than the productive
regions including the high latitude and the equatorial area.
In the case of the equator this was explained by the
advection model that some portion of DOC produced in
the equatorial upwelling area is exported toward the north
and south by advection, and accumulates in the surface
waters of the subtropical system (Peltzer and Hayward,
1996; Archer et al., 1997). In the case of the high latitude, high productivity area, however, little is known
about the fate of DOC produced in the euphotic layers.
Downward export, followed by accumulation in the subsurface is a plausible interpretation, but there seems to
be little evidence. Some recent studies have shown consistently that DOC in the surface waters of the Antarctic
Ocean remains at considerably low concentrations, close
to the deep water value (~40 µM, Table 1) (Kähler et al.,
1997; Wiebinga and de Baar, 1998; Carlson et al., 1998,
2000; Wedborg et al., 1998; Ogawa et al., 1999). Deep
vertical convection possibly restricts the accumulation of
DOC within the surface water, despite high productivity
(Ogawa et al., 1999). Carlson et al. (1998) showed that
the biological production in the Antarctic Ocean is preferentially partitioned as POC rather than DOC in the sur-
Dissolved Organic Matter in Oceanic Waters
133
face water, while major portion of the primary production accumulates as DOC in the temperate area of the
Atlantic Ocean. This could also possibly be attributed to
the relatively low concentrations of DOC in the high latitude area. According to Carlson et al.’s findings, the accumulation of the surface DOC enriched with the semilabile DOC in the ocean, may be closely correlated with
the partitioning of produced organic carbon (POC or
DOC), which may be controlled by plankton community
structure and food-web dynamics (Carlson et al., 2000;
Sweeney et al., 2000).
3.3 DOC in deep water
As argued above, the DOC in the deep ocean is the
largest pool of fixed carbon in the ocean (~650 Gt), approximately equivalent to the carbon stock of atmospheric
CO2 (~750 Gt) or terrestrial biomass (~600 Gt) (Hedges,
1992). In addition, this reservoir contains remarkably
unreactive carbon, resistant to microbial oxidation
(Barber, 1968). The apparent average 14C age of 4,000–
6,000 ybp (Williams and Druffel, 1987) is indicative of
this biologically refractory oceanic DOC, which exceeds
the circulation time of the whole ocean (~1,000 yr). Thus,
the refractory DOC appears to make a relatively minor
contribution to the active sites in the global carbon cycle.
On the other hand, it plays a potential role for the sequestration of fixed carbon in the ocean, suggesting its importance in understanding of the mechanism of global
warming. Here, assuming a steady state for the refractory DOC pool in the ocean, its global turnover rate would
be approximately 0.1 Gt y–1 by dividing the stock (~650
Gt) by the average age (4,000–6,000 ybp). Although this
value is much lower than that of the semi-labile DOC
(~1.2 Gt y–1; Hansell and Carlson, 1998b), it is comparable to the global carbon flux of sediment burial in the
ocean (~0.1 Gt y –1; Berner, 1989). However, since this
estimate is based only on the average 14C age, it has been
pointed out that the estimated flux would increase greatly
if the deep DOC is a composite pool of which DOC is
older and younger than the average age (Mantoura and
Woodward, 1983). These authors proposed that a significant contribution of riverine DOC to the oceanic DOC
pool would be consistent with the existence of these different DOC ages. However, a few later studies measuring δ13C or lignin contents of DOM as indicators of the
terrestrial origin demonstrated that most of the oceanic
reservoir of DOC was of marine origin, and the
terregenious source made a minor contribution (MeyersSchulte and Hedges, 1986; Williams and Druffel, 1987;
Opsahl and Benner, 1997).
In the 1990s, deep DOC studies focused on its photochemical degradation (Kieber et al., 1989; Mopper et
al., 1991; Moran and Zepp, 1997; Benner and Biddanda,
1998). These studies showed that the refractory DOC in
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H. Ogawa and E. Tanoue
deep water was photochemically reactive, and degraded
by sunlight into low molecular weight, biologically labile, organic compounds and carbon monoxide. The conclusion was that photochemical degradation is an important pathway for removing refractory DOC in the ocean.
Moreover, the estimated turnover time of the oceanic reservoir of refractory DOC (500–2100 yr), based on the
photochemical removal rate, was less than the apparent
average 14C age of DOC in the deep ocean (Mopper et
al., 1991). Cherrier et al. (1999) measured the natural 14C
in the nucleic acids of marine bacteria from the surface
waters of the North Pacific. Interestingly, their results
gave direct evidence that marine bacteria utilized 14Cdepleted old carbon, i.e., the refractory DOC, probably
through photochemical transformation into biologically
labile DOC (Kieber et al., 1989). On the basis of the bacterial utilization rate of old carbon, the turnover time of
the oceanic reservoir of the refractory DOC is estimated
as ~1,100 yr, which is also significantly shorter than the
average 14C age of the deep DOC. These findings demonstrate that some portion of refractory DOC is more
dynamic than estimated for the bulk pool, and support
previous suggestions about the existence of the different
age components in the refractory DOC pool in the deep
ocean (Mantoura and Woodward, 1983).
Sedimentary organic matter is a possible source of
the older component in the deep DOC. Bauer and Druffel
(1998) found elevated DOC concentrations with a more
14
C-depleted, “older”, signature in the continental slope
and rise waters than those from the adjacent North Atlantic and North Pacific central gyres. This strongly suggests
older DOC inputs from ocean margins to the open ocean
interior, although a recent study pointed out that their interpretation of the DOC age gradient may be wrong
(Hansell et al., 2002). Burdige et al. (1999) estimated a
significant DOC flux from marine sediments (~0.2
Gt C y–1), roughly equivalent to riverine DOC input, using in situ measurements of benthic flux of DOC from
continental margin sediments. This estimate emphasizes
the importance of marine sediment as a source of refractory DOC in the oceans. Detailed profiles of DOC concentrations and 14C contents in the water column above
sediments in continental margin would give a further test
of this hypothesis.
The DOC concentrations in the deep ocean have long
been considered to be quite uniform, which is consistent
with the previous concept that the DOC in deep water
would be a kind of residue of microbial process that is
resistant to further degradation (Barber, 1968). Recent
studies with the high precision HTC method, however,
revealed clear signals for DOC concentration gradients
from the global survey of deep ocean DOC (Hansell and
Carlson, 1998a). These authors observed that DOC concentrations in deep waters decreased by 14 µM from the
North Atlantic Ocean, that is the region of deep water
formation, to the terminal of the deep water circulation
in the North Pacific Ocean, suggesting the occurrence of
long-term decomposition of DOC along the global deep
water current. Their findings are basically consistent with
the fact that the apparent 14C age of the deep water DOC
in the Sargasso Sea is ~2,000 yr younger than that in the
north central Pacific (Bauer et al., 1992). On the other
hand, their results also indicated small increases in the
deep water DOC concentrations in the mid-latitudes of
the Southern Hemisphere, suggesting inputs of DOC to
the deep ocean.
Moreover, in the 1990s there were some important
discoveries concerning the photochemical process of
DOM in the ocean. As described above, photochemical
degradation was found to be an important pathway for
the removal of biologically refractory DOC in the ocean.
Interestingly, exposure to the sunlight in the surface water is also likely to induce the production of refractory
DOC from bioavailable substrates (Kieber et al., 1997;
Benner and Biddanda, 1998), implying a broad role for
the photochemical process in the DOC dynamics in the
ocean. A photochemically reactive component in natural
DOC is generally known as a humic-like substance having fluorescent properties inherent in this structure. The
refractory DOC in the deep ocean is representative of this
humic type of fluorescent organic matter. The vertical
distribution of this fluorescence is characterized by a drastic decrease in the upper water column with decreasing
depth (Fig. 1a). The profile displays evidence for the
photodegradation of fluorescence of DOM in the surface
ocean (Mopper et al., 1991; Chen and Bada, 1992). Although it is difficult to directly convert the humic fluorescence to its DOC concentration, the refractory DOC
in surface waters is possibly less than in deep waters. The
semi-labile DOC in surface waters, when it is estimated
on the basis of a uniform concentration of the refractory
DOC in the whole water column, might therefore be underestimated (Fig. 1a). These findings suggest that DOC
in the surface ocean may be involved in more dynamic
carbon cycling than was previously considered.
The mass balance assessment for the oceanic reservoir of DOC indicates that the riverine input (~0.2
Gt y–1; Meybeck, 1982) is roughly equivalent to the turnover of the refractory DOC in the ocean (~0.1 Gt y–1),
but the signature of terrestrial origin remains insignificant in the oceanic pool of DOC (Meyers-Schulte and
Hedges, 1986; Williams and Druffel, 1987; Opsahl and
Benner, 1997). The riverine DOC is known to be typical
of humic-like substances and relatively resistant to microbial degradation, which is similar to the deep ocean
DOC (Mantoura and Woodward, 1983). In addition, recent studies indicate that riverine DOC is generally characterized by enhanced fluorescence and photochemical
reactivity. The above discrepancy (i.e., what is the fate of
riverine DOC?), therefore, is perhaps explained by its
rapid removal through photodegradation prior to entering the oceanic DOC pool (Kieber et al., 1990; Coble,
1996; Opsahl and Benner, 1997).
4. Characterization of DOM in Seawater
4.1 Size distribution
Molecular size spectrum is one of the important characters of DOM. It was found that the results of earlier
reports were not necessarily consistent with each other.
In the 1990s, however, figures about the size distribution
of DOM determined using the ultrafiltration technique
were compiled (see the review by Ogawa, 2000). The significant finding is that the low molecular weight (LMW)
fraction is the major size fraction of DOM throughout
the whole water column in the ocean. The LMW fraction
(less than 1 kDa) accounts for approximately 65–80% of
the bulk DOC, while the high molecular weight (HMW)
fractions are minor portions of DOC (ca. 20–35% for >1
kDa and 2–7% for >10 kDa; Fig. 1b). In addition, it was
found that the HMW fraction was abundant in the surface (ca. 30–35% for >1 kDa and 5–7% for >10 kDa)
compared with the deep water (20–25% for >1 kDa and
2–4% for >10 kDa). Thus, most old DOM in the deep
ocean is LMW-DOM (75–80% of the bulk DOC). This
finding suggests that the HMW-DOM is relatively reactive while the LMW-DOM is the major form of refractory DOM in the ocean. The results of degradation experiments (Amon and Benner, 1994, 1996) and 14C-age
measurements (Guo et al., 1996) of different size classes
of DOM support this concept. However, we have no appropriate explanation for why the LMW-DOM is resistant to microbial oxidation, so that it forms a large organic reservoir in the ocean. This remains one of the biggest questions for the future marine DOM research. Since
the ultrafiltration technique allows us to obtain sufficient
HMW-DOM sample for various chemical analyses, our
knowledge of the chemical nature of the HMW fraction
has been increasing rapidly (see below). However, the
LMW-DOM is the least known fraction of DOM because
of the lack of a method for concentrating and desalting
this fraction. Methodological improvement and further
characterization of the LMW-DOM is needed for better
understanding of the entire DOM nature.
4.2 Elemental composition
Elemental information, the C:N ratio in particular,
is essential to understand the origin and processing of
natural organic matter. The C:N ratio of particulate organic matter (POM) retained on a glass fiber filter has
been directly determined with a multiple elemental
analyzer, typically a CHN analyzer. It is widely known
Dissolved Organic Matter in Oceanic Waters
135
Table 2. The C:N ratio of dissolved organic matter in oceanic seawater.
Study area
Bulk DOM (or TOM indicated by *)
*Sargasso Sea (Burmuda)
North Atlantic (33–60°N)
*Eastern Pacific: equatorial
subtropical
high latitude
C:N atom ratio
Literature
10–14 (surface)
14 ± 2 (surface)
14–15 (deep)
8–12 (surface)
11–14 (surface)
17–18 (surface)
8–12 (surface)
Kähler and Koeve, 2001
Hansell and Waterhouse, 1997
*Equatorial Pacific
7–9 (surface)
Libby and Wheeler, 1997
Southern Ocean
5–8 (surface)
4–5 (deep)
Kähler et al., 1997
Southern Ocean
5–16 (surface)
11–22 (deep)
Ogawa et al., 1999
Ross Sea
8–23 (<150 m)
14–20 (150–600 m)
Carlson et al., 2000
Arctic Ocean
9–25 (surface)
HMW-DOM (>1 kDa) concentrated by ultrafiltration
North Pacific, Equatorial Pacific,
13–18 (surface)
Gulf of Mexico, Sargasso Sea
18–22 (deep)
Wheeler et al., 1998
McCarthy et al., 1993, 1996, 1997, 1998;
Benner et al., 1997
Pacific
16–17 (surface)
17–18 (deep)
Clark et al., 1998
Middle Atlantic Bight
11–16 (surface)
14–24 (deep)
Guo et al., 1996
Humic-like matter collected using XAD resins (type 2 or 8)
North Pacific, Sargasso Sea
39–57
North Pacific, Equatorial Pacific
36–47
that the C:N ratio of POM ranges from approximately 5
to 15 in oceanic waters, and the value in surface waters is
close to that of marine phytoplankton (6–8). However,
fewer studies have focused on the C:N ratio of DOM in
seawater, essentially due to the difficulty of precisely
determining this parameter, which is generally obtained
from separate measurements of DOC and dissolved organic nitrogen (DON). The concentration of DON is indirectly determined by subtracting the concentration of
dissolved inorganic nitrogen (DIN) from total dissolved
nitrogen (TDN). Therefore, the precision of DON data
depends upon measurements of DIN (=(nitrate + nitrite)
136
Bates and Hansell, 1999
Hansell and Carlson, 2001b
H. Ogawa and E. Tanoue
Druffel et al., 1992
Hedges et al., 1992
+ ammonium) and TDN. In particular, this becomes a critical problem in deep water samples in which most TDN
consists of nitrate (up to ca. 45 µM) while DON (~2 µM)
is a relatively minor portion of TDN. If both TDN and
DIN analyses are performed with normal precision (2–
3% of c.v.), it would give an approximately 50% error of
DON value in deep waters. Finally, it would lead to approximately ±10 of the C:N value (as s.d.), even if precise DOC data are used.
Recently, two approaches have been used to determine carefully the C:N ratio of DOM in seawater. One is
the development of the highly precise method of TDN
measurement combined with the HTC system for DOC
analysis (Hansell, 1993; Kähler et al., 1997; AlvarezSalgado and Miller, 1998; Ogawa et al., 1999). The controversy about the DON (TDN) measurement as well as
DOC was current in the latter 1980s, started by the report
of the Japanese workers that the HTC technique gave
much higher DON concentrations in oceanic seawaters
than the conventional WCO method (Suzuki et al., 1985).
However, after further examination by other workers
(Walsh, 1989; Maita and Yanada, 1990; Hedges et al.,
1993; Koike and Tupas, 1993; Hansell, 1993), their results were found to have little reproducibility, and the
DON values obtained by the HTC method in the early
stage was finally retracted (Suzuki, 1993). Ogawa et al.
(1999) improved a commercial HTC unit of DOC analysis for the simultaneous and highly precise measurement
of DOC and TDN, to which a chemiluminescence detector for nitric monoxide (NO) produced from TDN compounds was connected. They reported a mean error of ±1
for the C:N value with this modified HTC system. However, the high precision HTC method for TDN measurements does not yet compare with DOC for general use.
The other approach is to concentrate DOM in
seawater, followed by the same elemental analysis as used
for POM. The ultrafiltration technique is generally used
for this purpose, which largely reduces the error of C:N
measurements of DOM by the effect of concentration with
desalting, which includes the removal of DIN. The data
obtained by this approach, however, is limited to the
HMW-DOM fraction which is concentrated by ultrafiltration, and it is not applicable to the bulk and the LMWDOM.
The recently reported C:N ratio values of oceanic
DOM are summarized in Table 2. The observed C:N ratios of DOM (mostly >8) were higher than the typical
value of POM, although a few studies reported relatively
lower values (<10). Considering that the source of oceanic DOM is marine organisms that have a lower C:N
ratio (mostly <8), carbon-rich materials would be selectively supplied and/or preserved as DOM. The HMWDOM (>1 kDa) have a relatively higher and uniform value
of C:N, mostly between 13 and 20, compared with the
bulk DOM, suggesting a difference in the elemental composition between the HMW- and the LMW-DOM. The
HMW-DOM consists of relatively carbon-rich materials,
as revealed in recent findings by NMR, which show that
carbohydrates containing no nitrogen are the principal
functional group of the HMW-DOM in oceanic waters
(Benner et al., 1992; Aluwihare et al., 1997). However,
we have little direct evidence for lower C:N ratio of LMWDOM because of difficulty in the precise measurement
of DON in the unconcentrated LMW fraction of DOM in
sweater. Further methodological improvement is needed
to determine the C:N ratio of the LMW-DOM.
Interestingly, the humic-like substance that was collected and concentrated using XAD resins (nonionic
macroporous resins) from acidified seawater samples
showed much higher C:N ratios (36–57, Druffel et al.,
1992; Hedges et al., 1992), indicating that the majority is
considerably carbon rich. This implies the possible occurrence of nitrogen-rich materials in non-humic-like
substance of DOM to compensate for the bulk C:N ratio
(ca. 8–20). It is difficult to specify chemical species of
these substances because they are only operationally defined. However, this finding demonstrates that DOM consists of various fractions that have different C:N ratios.
It appears that there are some regional differences in
the C:N ratio of DOM in the ocean. Ogawa et al. (1999)
and Carlson et al. (2000) showed that the C:N ratios of
the bulk DOM were higher in deep waters than in the
surface ocean, suggesting that nitrogenous compounds
would be selectively decomposed during the downward
transport of DOM. Similarly, relatively higher C:N ratios
of the HMW-DOM were found in deep waters than at the
surface (Guo et al., 1996; Benner et al., 1997). Furthermore, Clark et al. (1998) determined the C:N:P ratio of
the HMW-DOM in waters from different depths in the
Pacific Ocean. Their results showed that the C:P ratio
increased dramatically with depth from 247 at the surface to 539 at 4000m, with significant increases in both
C:N and N:P ratio, suggesting that phosphorus in DOM
would be most preferentially remineralized in the oceanic water column, followed by nitrogen, and carbon was
relatively preserved.
It was observed that the C:N ratios of the bulk DOM
in the surface of the Antarctica Ocean and the subarctic
area of the Atlantic and the South Pacific, were relatively
lower (~10) than those from other oceanic sites including subtropical and temperate areas (Hansell and
Waterhouse, 1997; Ogawa et al., 1999; Carlson et al.,
2000; Kähler and Koeve, 2001). It is possible that the
nutritional condition would control the C:N ratio of DOM
in the surface ocean (Williams, 1995).
4.3 Chemical composition at molecular level
Chemical analyses of marine organic matter at the
molecular level have typically involved degradation (e.g.,
acid or alkaline hydrolyses, oxidation or pyrolysis) for
the release of compounds at monomer level that can be
determined by colorimetry, or identified and quantified
after chromatographic separations. The chemical analysis of DOM is not exceptional. From the results of this
approach, it has long been stated that most DOM in
seawater is uncharacterized at the molecular level
(Williams and Druffel, 1988; Hedges et al., 2000). This
conclusion remains essentially unchanged today. Even
considering the analytical uncertainties and lack of data
as mentioned below, the contribution of the major bio-
Dissolved Organic Matter in Oceanic Waters
137
chemical constituents, i.e., amino acids, carbohydrates and
lipids, may not exceed 30% of bulk DOM. This figure is
in striking contrast to particulate organic matter (POM)
of which more than 80% of organic carbon comprises
these three organic constituents (e.g., Wakeham et al.,
2000).
Amino acids and neutral sugars are generally determined to be major biochemical components that are identifiable at the molecular level. The total concentrations
of amino acids and sugars, as measured by chromatographic techniques after hydrolysis of seawaters, amount
to only 1–3% and 2–4% of DOC, respectively (Fig. 1c).
Although a few modifications of the hydrolysis method
were proposed to improve the yields of amino acids
(Robertson et al., 1987; Keil and Kirchman, 1991) and
neutral sugars (Skoog and Benner, 1997), their contribution to the bulk DOC did not change considerably. Lipids
are one of the most well documented organic compounds
in POM and sedimentary organic matter, whereas only a
few analyses have been done on dissolved lipids (e.g.,
Parrish, 1988; Liu et al., 1998; Muhlebach and Weber,
1998; Mannino and Harvey, 1999; Harvey and Mannino,
2001). Dissolved lipids in the Delaware Estuary comprised up to 0.3% and 1.6% of 1–30 kDa and 30 kDa–0.2
µm fractions of HMW-DOC, respectively, and values were
several times lower than those of carbohydrates and amino
acids in these fractions (Mannino and Harvey, 1999, 2000;
Harvey and Mannino, 2001). The major components of
dissolved lipids are fatty acids, with other lipids, including sterols, hydrocarbons etc., representing a minor contribution (Parrish, 1988; Liu et al., 1998; Muhlebach and
Weber, 1998; Mannino and Harvey, 1999). A systematic
survey of dissolved lipids in oceanic water is required,
since lipids are the major biochemical constituent in organisms with proteins and carbohydrates and could play
an important role in the formation of colloids in situ in
seawater.
A modified spectrophotometric determination with
3-methyl-2-benzothiazolinone hydrazone hydrochloride
(MBTH) was proposed for measuring the total carbohydrate (TCHO) concentrations in seawater (Pakulski and
Benner, 1992). The carbon concentrations in TCHO accounted for approximately 15–35% of DOC in the surface and 5–10% in deep waters of various oceanic areas,
indicating that carbohydrates are the most abundant component measurable in marine DOM (Pakulski and Benner,
1994) (Fig. 1c). Although the MBTH method is sensitive
to sugar-derived aldehydes, the method is colorimetric
and does not directly determine the sugar molecules.
These results demonstrate that a significant portion of
DOC occurs as carbohydrate-like compounds that are
identifiable as the functional group (aldehydes) but unidentified by the typical chromatographic technique. In
fact, only 7–20% of TCHO in marine DOM was identi-
138
H. Ogawa and E. Tanoue
fied as neutral sugars, which are major constituents of
carbohydrates in marine organism (Skoog and Benner,
1997).
Total dissolved amino acids (TDAA) that are quantified after acid hydrolysis represent the largest well-defined molecules of DON. Amino acid nitrogen accounts
for approximately 10% or less of bulk DON (Huberten et
al., 1995; Dittmar et al., 2001) and <10% in HMW-DON
(>1 kDa fraction) (McCarthy et al., 1997). The dominant
amino acid in TDAA is glycine, irrespective of the temporal and spatial differences in observations, followed by
alanine, aspartic acid or serine (Hubberten et al., 1994,
1995; Keil and Kirchman, 1999; Dittmar et al., 2001).
Fractionation of DOM using XAD resin into the hydrophobic “humic” and hydrophylic compounds revealed that
glycine and other dominant amino acids in TDAA are
enriched in the hydrophobic fraction (Hubberten et al.,
1994, 1995). Since the concentration of amino acids in
the hydrophilic fraction correlated with chlorophyll, but
no relationship was found between the hydrophobic fraction and chlorophyll, it was concluded that amino acids
are selectively utilized by bacteria and that amino acids
in the hydrophobic fraction occur as a background TDAA,
forming part of recalcitrant DOM (Hubberten et al., 1994,
1995). Enrichment in glycine was also found in fluvial
riverine DOM (Dittmar et al., 2001). The preferential
accumulation of glycine was suggested along the progressive degradation from fresh phytoplankton to deep-sea
sediment (Dauwe et al., 1999).
The occurrence of microbial-derived D-enantiomers
of amino acids has given us an invaluable clue to the contribution of microbes to DOM (McCarthy et al., 1998;
Dittmar et al., 2001). In the HMW-DOM (>1 kDa) fraction from entire water columns in the central Pacific and
the Gulf of Mexico, alanine showed the most pronounced
enrichment, with D/L ratios near 0.5, whereas ratios of
aspartic acid, glutamic acid and serine fell between 0.2
and 0.4, and no other D-amino acid was detected at significant concentrations (McCarthy et al., 1998). Highly
selective patterns of D-amino acid enrichment are a strong
indication that D-amino acids did not derive from abiotic
racemization but represent a constituent of peptidoglycan in the bacterial outer membrane. A systematic survey
of amino acids and their D-enantiomers in the bulk DOM
and POM also demonstrated the major microbial contribution to the bulk DON in seawater in the Arctic Ocean
(Dittmar et al., 2001). Contributions of TDAA nitrogen
to DON were almost constant and accounted for approximately 10% in the water column from the near-shore to
open-sea areas. The D-enantiomers of alanine, aspartic
acid, glutamic acid and serine were determined in the
TDAA, and the proportions of all D-enantiomers increased with depth, with D-alanine comprising 44% of
total alanine in deep water. The similarity of amino acid
composition and high D/L ratios of TDAA, exceeding the
values of bacterioplankton (McCarthy et al., 1998; Amon
et al., 2001), in wide areas of the ocean indicates that
phytoplankton-derived amino acids were replaced by
microbial organic matter during the process of decomposition, and the repeated replacements might shape the
chemical composition of both TDAA and DOM (Meon
and Kirchman, 2001; Amon et al., 2001; Dittmar et al.,
2001).
4.4 Approaches to characterizing DOM at functional
group and biomacromolecular levels
One of the goals of marine organic geochemistry is
to identify organic compounds and to describe the in situ
states of these materials in seawater. For bulk chemical
characterization of DOM, a combination of two techniques, i.e., concentration of DOM with tangential (or
cross) flow ultrafiltration and application of nuclear magnetic resonance (NMR) to the concentrate, has given us
some fundamental new insights into the chemical nature
of DOM. The majority of functional groups in HMW-DOC
(>1 kDa) determined by 13C and 1H NMR spectroscopy
indicate that carbohydrates are the major constituents of
DOM (Benner et al., 1992; Aluwihare et al., 1997), particularly photosynthetically derived acyl oligosaccharides
(Aluwihare et al., 1997). The results of 15N and 31P NMR
analyses of HMW-DOM (>1 kDa) indicated that approximately 70–90% of organic nitrogen is in the form of amide
(McCarthy et al., 1997) and organic phosphorus is dominated by two major classes, phosphorus esters and
phosphonates (Clark et al., 1998; Kolowith (née Clark)
et al., 2001).
The NMR studies clearly demonstrate that most
HMW-DOM (>1 kDa) is identifiable as functional groups
common to biochemical constituents in organisms. A combination of two techniques, i.e., tangential (or cross) flow
ultrafiltration and gel electrophoresis, provided direct
evidence that specific protein molecules occur universally
in HMW-DOM (>10 kDa), and some proteins were identified as bacterial outer membrane proteins, collectively
termed “porin” (Tanoue, 1995, 2000; Tanoue et al., 1995,
1996; Suzuki et al., 1997; Yamada et al., 2000). These
findings, as well as the occurrence of D-enantiomers as
mentioned above, indicate that biomolecules, particularly
bacterial membrane components, survive in DOM. In the
culture experiments of marine bacteria, Ogawa et al.
(2001) found that DOM, including LMW-DOM (<10
kDa), rapidly accumulates (<48 h), indicating that bacterial processes are a possible pathway of production of
even the molecularly uncharacterized component of
DOM.
Interestingly, the spectra of 15N, 31P and 13C NMR,
did not show any significant temporal and spatial variability. Such a stable DOM fraction, a major component
of DOM, has been collectively designated “water humus”.
The humic substances are formed by condensation reactions that depend on intermolecular collisions of degradation products of photosynthetically produced
biopolymers. These condensations include the Maillard
reactions between carbohydrates and amino acids (or proteins) that form “melanoidins”, and such processes are
termed “humification”. Although the occurrence of amino
acids and carbohydrate condensation products has not
been reported in marine environments, the mechanisms
of the Maillard reactions have been investigated in the
field of biochemistry because several diseases are caused
by the Maillard reaction, indicating the reactions progress
on a relatively short time scale.
In the biochemical literature (e.g., review by Njoroge
and Monnier, 1989), glucose and other reducing sugars
react with protein by non-enzymatic, post-translation
modification process called non-enzymatic glycosylation
or glycation, and the products of the reaction are termed
advanced-glycation end-products (AGEs). The compound
N-phenacylthiazolium bromide (PTB) was used to break
down AGEs (Vasan et al., 1996). The results of applications of PTB treatment to geochemical samples indicated
that AGEs certainly formed in ca. 20,000 year old and ca.
4,000 year old sediments (Poiner et al., 1998; Nguyen
and Harvey, 2001) and during the short time scale (3
months) plant litter degradation experiments (Fogel and
Tuross, 1999). “Humification” does occur in the environments on a certain time scale. However it appears that
there is no real evidence that such heteropolycondensation
reactions occur in the water column, with extremely low
levels of organic matter, low temperature and lack of catalytic agents (e.g., Tanoue, 2000). Occurrences of the
pyridine-type N and/or imine-N signals in geochemical
samples are thought to be evidence of AGEs resulted from
the Maillard reaction (Evershed et al., 1997). However,
no pyridine-type N and/or imine-N signals were found in
the above-mentioned 15N NMR spectra of HMW-DOM
(McCarthy et al., 1997). Knicker (2000) found only
amide-N in peptide-like structures with little evidence of
pyridine-type N and/or imine-N signals from the 15N NMR
spectra of the algal residue after two months degradation. Amide and ester linkages and C-P bonds are also
difficult to form via abiological condensation reactions
(e.g., Hedges et al., 2000). The NMR spectra, particularly 15N and 31P NMR spectra (McCarthy et al., 1997;
Clark et al., 1998; Kolowith (née Clark) et al., 2001),
imply that such humification is not a major process for
the formation of stable DOM.
In marine environments, primary production by
phytoplankton is the ultimate source of marine organic
matter; however, growing evidence supports the idea that
the phytoplankton-derived organic matter is decomposed
rapidly by the microbial processes. During the process,
Dissolved Organic Matter in Oceanic Waters
139
certain organic compounds escape from rapid
remineralization and form microbiologically recalcitrant
organic molecules. Thus, degradation processes are more
important than the formation of humic materials in controlling and maintaining the reactive pool of organic matter on the earth’s surface (e.g., see review by Volkman
and Tanoue, 2002). Microbial processes probably control the molecular composition of DOM, not only identified molecules but also the molecularly uncharacterized
fraction, a major part of DOM (Ogawa et al., 2001).
5. Survival of Biomolecules in the Environments
The discussions reported above are persuasive
enough to lead us to believe that some biomolecules, particularly proteins and peptides, survive in marine and
terrestrial DOM. In the sediments, the materials that can
be extracted from sediments with an alkaline solution or
denaturing reagents were subjected to chemical digestion
and cleaved by proteolytic enzymes (Pantoja and Lee,
1999; Nguyen and Harvey, 2001), indicating that
proteinacous materials were preserved. The protein remnants were also identified from the insoluble geopolymers
(humin and kerogen) by thermochemolysis with the tetramethylammonium hydroxide (TMAH)/GC-MS technique (Zang et al., 2000; Mongenot et al., 2001; Knicker
et al., 2001). Direct and indirect evidence consistently
indicates that proteins and proteinacous materials, including peptides and amino acids, are present in detrital organic pool in the environments. Such recognition is an
encouragement to further explain and delineate the processes by which such materials survive and are preserved.
Hypotheses proposed in the fields of chemical oceanography, organic geochemistry and soil chemistry, are described briefly below.
(a) Geopolymerization: The concept is that the monomers
and/or oligomers, particularly amino acids and reduced
sugars, react with each other abiologically (i.e., Maillard
reaction) to form biologically refractory humic substances
and further altered to kerogen. This concept has been considered to be the major process for formation of
uncharacterized DOM; however, recent studies suggest
that this process is not responsible for the formation of
refractory DOM as mentioned above.
(b) Sorption: The concept that organic matter sorbed onto
a mineral surface or inside the micropore of a mineral is
protected from biological degradation.
Simple sorption: It is well known that concentrations of
sedimentary organic matter are related to the grain size,
and in turn to the mineral surface area (e.g., Hedges and
Keil, 1995, and references therein), and that sorption to
solid surfaces has been shown to inhibit bacterial degradation of labile organic matter and proteins (Marshmann
and Marshall, 1981; Taylor et al., 1994a, b; Keil et al.,
1994; Hedges and Keil, 1995; Nagata and Kirchman,
140
H. Ogawa and E. Tanoue
1996).
Mesopore sorption: Based on Mayer’s (1994) study of
the pore sizes of the coastal sediment, a globular macromolecule, e.g., a 67 kDa protein with a Stokes radius of
3.6 nm, or a diameter of 7.2 nm (Creighton, 1993), is capable of diffusing into mesopores of the sediments. Molecules incorporated into the mesopores are protected from
enzymatic as well as bacterial attack because enzymes
cannot access the molecules.
(c) Selective preservation: The concept is that some organic matter is preserved selectively because of its refractory nature. The idea originally came from studies on
kerogens, i.e., the insoluble organic matter of sedimentary rocks that has been thought to be an origin of oil.
Chemical and microscopic examinations suggested that
types of kerogens were principally associated with an
“ultralaminae” microstructure, being constituted with
non-hydrolyzable and diagenetically resistant macromolecules based on a network of long, saturated hydrocarbon chains, termed collectively “algaenans”. The
algaenans are ubiquitous in freshwater green algae (e.g.,
de Leeuw and Largeau, 1993), as cutans in cuticles and
suberans in periderm tissue of higher plants (e.g.,
Collinson et al., 1994), as tegmens in inner seed coats of
freshwater plants (e.g., Hemsley et al., 1993), and as
polycadinenes in resins (van Aarsssen et al., 1990).
Cyanobacteria and bacteria (as bacterans) and marine algae also synthesize algaenans (e.g., Largeau, 1995; Gelin
et al., 1996, 1999).
(d) Encapsulation: The concept is that the proteinaceous
materials are incorporated into a surrounding organic
matrix and are protected from biological attack. The organic matrix could be different in different environments
and time scales. In laboratory experiments, it has been
demonstrated that spiked amino acids or proteins associated with existing natural DOM and in situ biological utilization rates decrease. The results imply that abiotic association with an organic matrix is potentially important
for surviving labile organic compounds in seawater
(Carlson et al., 1985; Keil and Kirchman, 1994).
Membrane encapsulation: Proteolytic digestion indicated
that peptides, particularly phytoplankton membrane components, are preferentially enriched as compared with the
cytosol proteins in estuarine POM (Laursen et al., 1996).
The occurrences of bacterial outer membrane components
in HMW-DOM (e.g., Tanoue et al., 1995; McCarthy et
al., 1998) suggest that the concept could be one of the
possible mechanisms for survival of biomacromolecules
in DOM.
Liposome encapsulation: Nagata and Kirchman (1992)
found that one type of flagellate fecal pellets, consisting
of phospholipids, enzymes and other biochemically labile substances, form liposome-like colloidal particles.
They proposed that liposome-like organic complexes pro-
duced by flagellates could provide a microenvironment
for geochemical modification of organic matter to proceed (see also review by Nagata and Kirchman, 1997).
Network encapsulation: The concept has come from studies of sedimentary organic matter. Proteinacous materials are incorporated within the above-mentioned refractory aliphatic macromolecular matrix and are sterically
protected from bacterial hydrolysis (Knicher and Hatcher,
1997). The proteinacous materials extracted from the sediment were readily degradable by chemical and enzymatic
procedures.
(e) Aggregation: Nguyen and Harvey (2001) showed in
laboratory experiments that proteolytic digestable
peptides were preserved in diatom detritus by the formation of large aggregates. They mentioned that non-covalent interactions, the conformation alteration and the
hydrophobicity of the original polypeptides could be important for the preservation of protein and/or peptides.
Every hypothesis itemized above is applicable to the
respective natural environments. Considering the conditions of the water column in the marine environments,
the hypotheses of encapsulation or aggregation seem to
be probable mechanisms for the survival of biomolecules.
The majority of DOM has a molecular mass of <1 kDa.
D-enantiomers of amino acids are also detected in the
LMW-DOM, indicating that the hypotheses of membrane
and network encapsulations may not be applicable to the
entire DOM. At present, liposome encapsulation or aggregation is a likely mechanism for survival of
biomolecules, assuming that such a macrostructure is
formed reversibly from LMW-DOM, although the boundary conditions for association-dissociation equilibrium are
unknown.
So far, the sources of the biomolecules that we have
identified are very specific. For example, the source of
the porin protein identified from DOM (e.g., Tanoue et
al., 1995) was derived not from the bulk bacterial population but from a specific species of Gram-negative bacterium. Biosynthesis of phosphonates has been reported
from a few groups of organisms (e.g., Clark et al., 1999),
although an actual source of phosphonates in DOM is
unknown. The above hypotheses are based on the same
idea, i.e., how biomolecules are protected from biological attack. However, the hypothesis does not explain why
selective biomolecules actually survive in DOM. From
this point of view, the physicochemical properties of the
biomolecules or the process by which the biomolecules
are transferred from source organisms to DOM might be
other factors determining the survival of the biomolecules
in DOM. In fact, synergism among different factors, protection mechanisms, physicochemical properties or transfer processes, might control the survival of the
biomolecules in the sea.
6. Concluding Remarks
The amount of information on oceanic DOM has increased dramatically in the last decade, supported by
progress in the chemical approaches, including both the
improvement of conventional methods and the introduction of new techniques. We are now looking at various
aspects of DOM from different viewpoints, such as the
bulk properties (i.e., DOC), size distribution, elemental,
isotopic, molecular and functional group levels. In addition, we know that both biological and physicochemical
processes contribute significantly to DOM dynamics in
the ocean, including its production, transformation and
decomposition. Some findings have had a great individual
impact upon our concepts. However, the combination of
different types of information allows us to better face
various aspects of the oceanic DOM and the new problems that are challenges for future. From this point of
view, four topics are given here although they are closely
related with one another.
6.1 Semi-labile DOC
The high precision HTC method has been successful in detecting the detailed distribution and fluctuation
of the semi-labile DOC in the ocean, and in quantifying
the role of DOC in export production (Copin-Montégut
and Avril, 1993; Carlson et al., 1994; Carlson and
Ducklow, 1995; Peltzer and Hayward, 1996; Hansell and
Carlson, 1998a, b, 2001a, b; Hansell et al., 2002). This
semi-labile DOC, however, is simply a conceptual category of DOC based on vertical distributions of DOC (see
Fig. 1a), but is supported by no evidence from chemical
characterization. Moreover, little is known about the
mechanisms of production and decomposition of the semilabile DOC in the ocean. Thus, a future challenge should
focus on its qualitative characterization as well as the
quantitative evaluation.
6.2 LMW-DOM
Most of the oceanic DOM consists of the LMW (<1
kDa) components (Ogawa, 2000; see Fig. 1b). This is
consistent with other information demonstrating that
LMW-DOM is responsible for the major nature of bulk
DOM in the ocean, which is biologically refractory and
molecularly uncharacterized (Amon and Benner, 1994,
1996; Guo et al., 1996). However, there is little knowledge about its chemical features and processes of production, transformation and decomposition. In particular, the reason why these small molecules are apparently
resistant to microbial utilization is the key to understanding them; what mechanism allows part of the biologically
fixed carbon in the ocean to remain in seawater for a long
period and to develop a huge carbon reservoir of DOM?
Dissolved Organic Matter in Oceanic Waters
141
6.3 Molecularly uncharacterized DOM
The detailed chemical characterization of HMWDOM clearly indicates its biogenic signature, which is
incompatible with the previous concept of “humic substance” or “gelbstoffe” for a molecularly uncharacterized
DOM in seawater. Recent results from NMR studies show
that a major part of HMW-DOM is identifiable as functional groups common to biopolymers (Benner et al.,
1992; McCarthy et al., 1997; Aluwihare et al., 1997; Clark
et al., 1998), but it still remains uncharacterized at the
molecular level by chromatographic techniques (see Fig.
1c). What are the carbonaceous compounds that consist
of functional groups of carbohydrates but are unidentified as neutral sugars? What are the nitrogenous compounds that have an amide functional group but are unidentified as amino acids? This is an important issue to
resolve in the future, and should be a key to understanding the transformations of biopolymers into DOM.
6.4 Biologically refractory DOM
Recent findings of photochemical degradation
(Kieber et al., 1989; Mopper et al., 1991), 14C content in
bacterial nucleic acid (Cherrier et al., 1999), DOC gradient in the deep ocean (Hansell and Carlson, 1998a), bacterial contribution to the production of DOM (Tanoue et
al., 1995, 1996; McCarthy et al., 1998; Ogawa et al.,
2001) give us a novel view that biologically refractory
DOM in the ocean might exist as a more dynamic pool
than was previously considered from the apparent average 14C-age of the bulk DOM (Williams and Druffel,
1987; Bauer et al., 1992). Most of our previous concepts
of oceanic DOM dynamics have been based upon the average 14C-age of bulk DOM. However, the above findings enable us to speculate on possible pathway that marine bacteria recycle the old carbon of DOM in the ocean.
Further evidence is necessary for us to learn the real age
spectrum and the real cycle of refractory DOM, and to
lead to a better understanding of the role of DOM in the
oceanic carbon cycling. Our future challenge should be
focused on, in particular, the detailed characterization of
photoreactive substances in DOM and the bacterial
processing of DOM, including production, transformation and remineralization.
Acknowledgements
We are grateful to Dr. D. A. Hansell and two anonymous reviewers for their careful reading and valuable
comments for the revision of this paper.
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