1. Dissolved Organic Matter- what is inside the black box?
What is the relationship between organic matter
composition, microbial diversity & production?
How is microbial metabolism coupled to organic matter
structure?
How does DOM affect microbial adaptation & evolution?
What sets the ocean inventory of organic carbon,
and why is so much organic carbon preserved in seawater?
Why do organic nitrogen and phosphorus accumulate
in nutrient limited regions of the surface ocean where they
are most needed, but disappear in the mesopelagic ocean
where they are not needed?

2. DOC sampling is hard to do… Isolation of DOM by ultrafiltration
Separation based on size
1 nm 1 kD
Selects for HMW fraction
about 30-35% TOC
Membrane effects what
Is collected
Some salts collected too!

3. DOC sampling filter (1 nm) sample concentrate (> 1000 MW) pump
filtrate
(< 1000 MW)

4. Ultrafiltration high molecular weight DOM (HMWDOM)
membrane filter
>1000 D
DOM fraction
30-35% DOC
< 1000 D
DOM fraction
65-70% DOC

5. Final product
30-35% of total DOC

6. DOM analysis by Nuclear Magnetic Resonance spectroscopy

7. 13C Nuclear Magnetic Resonance Spectrum
of high molecular weight dissolved organic matter (HMWDOM)
Frequency
Abundance

8. 13C Nuclear Magnetic Resonance Spectrum
of high molecular weight dissolved organic matter (HMWDOM)
Frequency
Different types of carbon
resonate at different
frequencies
Area is proportional to the
amount of carbon
Abundance
ppm

9. 1H, 31P, 15N - NMR spectra of HMWDOM
Clark et al L&O 2001

10. from the Atlantic and Pacific
13CNMR spectra of HMWDOM
from the Atlantic and Pacific
Sargasso Sea
NPSG
3 m
3 m

11. 13CNMR of marine phytoplankton*
Fragilariopsis cylindrus
Phaeocystis antarctica
*Adina Paytan, 2005

12. 13CNMR of plankton tows
Hedges et al GCA 2001

13. 13CNMR of HMWDOM in different
aquatic environments
North Pacific Ocean
Great Salt Lake
Leenher et al., (2004) Biogeochem. 69,
A significant fraction of HMWDOM is the same in all aquatic environments
HMWDOM has a different composition than marine than marine phytoplankton
Suggests selective preservation of a component of POM
Most of the C is carbohydrate (APS) large global inventory
Andrews Creek, RMNP
McKnight et al. (1997) Biogeochem. 62,

14. HMWDOM in the deep sea
Sargasso Sea
NPSG
3 m
3 m
2500 m
1800 m

15. 13C Nuclear Magnetic Resonance Spectrum
of high molecular weight dissolved organic matter

16. 13C Nuclear Magnetic Resonance Spectrum
of high molecular weight dissolved organic matter
COOH
CONH
(10%)

17. 13C Nuclear Magnetic Resonance Spectrum
of high molecular weight dissolved organic matter
OCO
(12%)
COOH
CONH
(10%)

18. 13C Nuclear Magnetic Resonance Spectrum
of high molecular weight dissolved organic matter
OCO
(12%)
HCOH
(55%)
COOH
CONH
(10%)

19. C/N = 15 +/- 3 13C Nuclear Magnetic Resonance Spectrum
of high molecular weight dissolved organic matter
OCO
(12%)
HCOH
(55%)
C/N = 15 +/- 3
COOH
CONH
(10%)
CHx
(10%)

20. C/N = 15 +/-3 13C Nuclear Magnetic Resonance Spectrum
of high molecular weight dissolved organic matter
OCO
(12%)
HCOH
(55%)
C/N = 15 +/-3
HCOH
OCO
= 4-5
COOH
CONH
(10%)
CHx
(10%)

21. HMWDOC, what could it be? From our knowledge of cell biochemicals…
Proteins C/N = 4, CHx(O):CON = 3
Carbohydrates C maybe N HCOH:OCO = 5
Lipids C only CHx:COOH = 18
CHx:COH = 30

22. HMWDOC, what could it be? From our knowledge of cell biochemicals…
Proteins C/N = 4, CHx(O):CON = 3
Carbohydrates C maybe N HCOH:OCO = 5
Lipids C only CHx:COOH = 18
CHx:COH = 30
……..looks to be mostly (50-70%) carbohydrate !

23. Spectral and chemical analyses of HMWDOC
Carbohydrate
50-70% of HMWDOC
200
150
100 50
-O-
-O-
13CNMR
Acid
hydrolysis

24. Spectral and chemical analyses of HMWDOC
Acid hydrolysis followed by
Monosaccharide analyses
yields 7 major neutral sugars
that represent 10-20% of HMWDOC
in surface water
Carbohydrate
50-70% of HMWDOC
200
150
100 50
13CNMR
R F A X Gl M Ga
Acid
hydrolysis

25. NMR and carbohydrate analyses of deep sea HMWDOC
monosaccharide distribution
surface
relative %
deep
relative %

26. Can we use the 14C of pure sugars to determine
If they are part of the reactive or nonreactive fraction of DOC?
reactive DOC
14C=DIC
“nonreactive” DOC
14C = DOC(deep)

27. Radiocarbon analyses of HMWDOC carbohydrates
Sample Hawaii NPSG
DIC ‰ (n=4) 89+7‰*
Glucose 47,
Galactose
Mannose
Xylose 52,
Arabinose ND
Fucose 49,
Rhamnose 40,
Average 56+6‰ ‰
*data from Ellen Druffel

28. Can 14C in sugars constrain the turnover time of
The “semi-reactive” fraction of DOC?
Assume the concentration of reactive DOC is at steady state,and that it produced
each year with a 14C value equal to DIC. Each year a fraction of the sugars
are non-selectively removed and the 14C value for the remainder is calculated.

29. O = -C-N -C-NH2 What else is in HMWDOC ? 15N NMR (90-98%) Amide-N
Amino-N
(2-10%)

30. -C-N- O What else is in HMWDOC ? 15N NMR proteins Amide-N Amino-N
amino sugars
CH3CONH

31. Is a large fraction of HMWDOC and HMWDON
from amino sugars?
15N NMR
1HNMR
Amide-N
Acetate

32. Is a large fraction of HMWDOC and HMWDON
from amino sugars?
15N NMR
1HNMR
Amide-N
Acetate
HNOCCH3
HNOCCH3

33. Is a large fraction of HMWDOC and HMWDON
from amino sugars?
HNOCCH3
Amide-N
Amide-N
Amino-N
HNH
HNH

34. Is a large fraction of HMWDOC and HMWDON
from amino sugars?
15N 1H
CH3CONH
acid
HNH
HNH +
CH3COOH

35. Composition and cycling of HMWDOM
Most of the carbon (75%) is
carbohydrate
Half the N is from aminosugars,
about 10% is from proteins
The sugar/aminosugar fraction
is the “semi-labile: DOC that
accumulates in surface water this may fuel about 50% of
microbial production in surface
waters.
Why does it accumulate?
Surface HMWDOM

36. What is the composition of Non-reactive DOC in the deep ocean
(1800 m Pacific)

37. Non-reactive DOM Semi-reactive DOM Very reactive DOM O -C-N- CH3 O HO
Mostly polysaccharides
Biopolymers…
80-90% of cell C-N-P
Don’t really
Know
Humic substances?
Free amino acids
Simple sugars
Urea…etc.
10-20% of cell C-N-P

38. Non-reactive DOM Semi-reactive DOM Very reactive DOM Humic substances
Proteins
Polysaccharides
Biopolymers…
80-90% of cell C-N-P
Free amino acids
Simple sugars
Urea…etc.
10-20% of cell C-N-P
Concentration = 40 µM
25-40 µM
100 nm-1µM (?)
Inventory = 650 GT C
30-50 GT C
GT C
14C= -400 to -600‰
14C= ‰ (?)
14C= ‰
Annual flux = 0.1 GTC
Moderate to large ?
Large