THE
MEASUREMENT
PARTICULATE
OF DISSOLVED
ORGANIC
CARBON IN SEAWATER1
AND
David W. Menzel and Ralph E;. Vaccaro
Woods Hole Oceanographic
Jnstitution,
Woods
Hole,
Massachusetts
ABSTRACT
A method is dcscribcd for the rapid dctcrmination
of dissolved organic carbon in seawater
in concentrations
bctwcen 0.1 and 20 mg/liter.
The oxidation is carried out in sealed glass
ampoules using K&Lox as an oxidizing agent after the sample has been freed of inorganic
carbon. The resulting CO2 is passed through a nondispcrsive
infrared analyzer using nitrogen as a carrier and the signal output of the analyzer recorded,
Using appropriate
calibration curves, the carbon content is determined From the height of the peak. Approximately
100 samples can bc analyzed in a single clay with a precision of -I- 0.1 mg/liter using a sample volume of 5 ml.
A companion method is described for the determination
of particulate
carbon by hightemperature
combustion after concentration
of the sample on a glass-fiber filter.
The precision of this method is -c- 10 mg C in a range of O-500 pg C. Approximately
six samples
can be analyzed in an hour.
INTRODUCTCON
have been criticized by Duursma ( 196I ),
and it is generally conceded that the results
so obtained are unreliable because of varying degrees of oxidation. The quantity of
CO, evolved by chromic acid oxidation
has been measured either by acid-base
( Krogh and Keys 1934; Kay 1954) or coulometric titration
( Duursma 1961) with
quantitative
results. Wilson ( 1961) recently proposed wet combustion
using
K&08
as an oxidizing
agent and subsequent detection of the COZ in an infrared
analyzer. Both the Duursma and Wilson
techniques have been found satisfactory,
but each requires lengthy digestion proccdures ( l-3 hr per sample), In addition,
relatively large samples of water (250 ml)
are required which, if collected on shipboard, must be preserved during their return to land-based laboratories.
The method described here overcomes
both of these objections by a) permitting
the oxidation of a large number of samples
simultaneously at the time of collection and
b ) assuring permanent preservation of the
samples. Approximately
100 samples can
be oxidized and analyzed in a normal working day with a precision of +- 0.1 mg/liter
Woods Hole
in a range of 0.1-20 mg/liter.
by National
A companion method is described for
and 544 and
the determination
of particulate
carbon,
the Atomic
after its concentration on a glass-fiber fil138
Accepted methods for the determination
of dissolved carbon in seawater are generally based on wet oxidation, but according to Van Hall, Safranko, and Stenger
( 1963) this is less desirable than direct
combustion, since organic compounds vary
in their susceptibility to oxidation, Therefore, the above authors designed a technique using a high-temperature
combustion in an oxygenated atmosphere. Unfortunately, the lower limit of detection is
2 mg/liter.
The small sample volume
(0.02 ml) required, to avoid the generation of excessive steam pressure within the
combustion unit, precludes the use of their
method for the analysis of seawater samples. Alternative
evaporation
of larger
volumes to dryness, combined with hightemperature combustion, is also undesirable because of the quantity of halides
present and the ease with which organic
compounds can be volatilized during the
evaporation of the acidified sample.
Wet oxidation techniques involving the
titration of unrcduced excess KMn04 ( Korringa and Postma 1957; Gillbricht
1957)
r Contribution
No. 1421 from the
Oceanographic
Institution.
Supported
Scicncc Foundation
Grants No. 889
under Contract
AT( 30-l )3140 with
Energy Commission.
DISSOLVED
AND
PARTICULATE
ORGANIC
CARBON
IN
139
SEAWATER
1.
1.
2.
3.
4.
5.
FIG.
1.
dissolved
NITROGEN SUPPLY
REGULATOR
NEEDLE VALVE
FLOW METER
ASCARITE
TUBE
Schcmntic diagram
organic carbon.
6.
7.
8.
9.
10.
HYPODERMIC
NEEDLE
AMPOULE
RUBBER TUBING
GAS WASHING BOTTLE
Mg(CIO,),
DRYING TUBE
of cquipmcnt
used in the dctcction
ter, by direct combustion at high temperature. In this case, approximately six samples
an hour can bc analyzed with a precision of
-I- 10 pg c.
ANALYSIS
Ol? DISSOLVED
CARUON
The method consists of the wet oxidation of l-5 ml of filtered seawater by potassium pcrsulfate ( K&08)
in a sealed glass
ampoulc (Kimball No. 12011-L) after inorganic forms of carbon have been removed. Following
oxidation, the samples
may be stored indefinitely.
They are subsequently flushed through an infrared CO2
analyzer and the instrument
signal rccorded. Halogens and water vapor are
removed prior to introduction
into the
analyzer.
Appnmtzls
The apparatus used in the detection of
carbon as COP is shown in Fig. 1. This
consists of a source of compressed nitrogen
(which is scrubbed by ascarite to rcmovc
traces of CO2 contamination)
and the flow
11. INFRA RED CO2 ANALYZER
12. AMPLIFIER
lb. RECORDER
of CO2 resulting
from the oxidation
of
adjusted to a rate of exactly 200 ml/min.
Inorganic silicone tubing is preferable to
rubber for connecting the various components. The flow-rate valve terminates with
a hypodermic
needle which is inserted
through the wall of a short section of
tubing (5 mm id.), with the neck of the
ampoule inserted into the lower extremity
of the tube. The upper end of the tubing
terminates at a gas washing cylinder containing acid KI solution (20 g KI in 50
ml 10% HzS04) for the removal of free
chlorine gencrated during the oxidative
digestion.
Next, the nitrogen is passed
through a tube containing Mg( C104)2 for
the removal of water vapor before entering
a Beckman Model 15A infrared analyzer
equipped with 7-inch ( 17.8-cm) detection
cells, The output of the amplifier is recorded on a lo-mv recorder.
Proceclure
1) Five ml of seawater arc injected
through a 2.4-cm glass-fiber filter using a
140
DAVID
mt’ OF SAMPL E (BLANK
100'
1
'
2
'
3
'
4
'
W.
MENZEL
AND
RALPH
F. VACCARO
serted into the solution. The gas flow is
again started and the resulting deflection
on the recorder observed. The height of
this peak is proportional to the amount of
CO2 present in the sample.
DETERMINATION)
5
1
Calibration
mg C/L USING 5 ml SAMPLE
FIG. 2. Calibration
data for the measurement
of dissolved organic carbon and method for the
determination
of blanks by extrapolation.
Millipore
0 2 Filter Corporation
adapter
(XX3002500) into a lo-ml Pyrex ampoule
which has been precombusted in a muffle
furnace at 7OOC. The filters must also be
precombusted at 700C and prerinsed with
5 ml of the sample. 2) Then 0.1 g K2S208
and 0.2 ml 3% H3P04 are introduced into
the ampoule. 3) Nitrogen gas is bubbled
through a $-inch (10.2-cm) cannula into
the ampoule at a rate of 200 ml/min for
3 min to remove all inorganic carbon. 4)
A plug of silicone grease-is placed in thk
end of the ampoule and the ampoulc sealed
in an oxygen-gas flame. This step is extremely critical because no trace of combustion products of the flame must bc permitted to enter the ampoule. The seajing
is best accomplished b; heating the stem
at the midpoint while holding the neck
in a nair 01 forceDs and. wh& it is red
hot, &sting
the a;poule’and
drawing the
glass at the same time so that a seal is
Obtained without producing a capillary .. 5)
has
When a sufficient number of samples
been prepared, the organic carbon is oxidized by autoclaving at l3OC for one-half
hour. 6) After cooling, or at any time
thereafter, the neck of the sealed ampoule
is inserted into the rubber tubing (Fig. 1;
No. 7), and nitrogen is flushed through
the system until the analyzer indicates the
complctc removal of extraneous COB. The
gas flow is then shut off, the tip of the
ampoule is crushed, and the cannula is in2 Registered
tion, Bedford,
trademark, Milliporc
Massachusetts.
Filter
Corpora-
and determination
of blanks
The method was initially
calibrated
using dextrose solutions of appropriate conSubsequently, standardizations
centration.
were accomplished by injecting given quantities of CO:! gas directly into the system
with a microsyringe. Fig. 2 shows recorder
values obtained using standard dextrose
solutions corrected for blanks and those
obtained using CO2 gas. It should be noted
that the response of the analyzer is not
linear and carbon concentrations must be
obtained from the appropriate calibration
curve. The curve shown was adjusted so
that 4 mg C/liter gave approximately 100%
deflection. Variable potentiometers on the
amplifier make it possible to adjust the
sensitivity to any desired range. In addition, the degree of response can bc adjusted
by manipulating the volume of the sample
analyzed. If concentrations over 4 mg/liter
are anticipated, this becomes ncccssary bccause more than 2,OPI; C ( absolute) causes
considerable flattening of the curve.
The most satisfactory method for freeing
water of organic carbon was found to be
by double distillation from 1 liter of distilled water containing 1 ml of HsP04 and
10 g of K2S20s. An alternative method for
determining
blanks is to determine the
organic carbon in 1, 2, 3, 4, and 5 ml of
TARLE 1. Relative recovery of organic carbon
CO, using JGS,O, oxidation
on S-ml volumes
enriched seawater samples
~----.-~
__-_ __------~.Compound
Maxichlcd.
~_____
L-cysteine a HCl
l-10 phenanthrolint
Adcnylic acid
Sulfanilamidc
Chitodextrin
Acetic acid
lx leucine
Dextrose
Carbon
~~
11111111
mg/liter
-Minimum
as
of
__hverage
-__--
%
Rccovcry
1.44
1.50
1.44
1.4’7
101
1.76
3.12
2.75
2.86
2.05
1.95
3.00
1.87
3.14
2.70
2.70
2.04
2.04
3.05
1.74
3.00
2.80
3.00
1.96
1.86
3.00
1.80
3.09
2.76
2.90
2.01
1.92
3.01
_____-
102
99
100
101
98
98
100
Y
DISSOLVED
AND
PARTICULATE
ORGANIC
CARBON
IN
141
SEAWATER
1.
1.
6b.
L
OXYGEN SUPPLY
REGULATOR
COLEMANCARBON-HYDROGEN ANALYZER
lt SPIROMETER
Mg(ClO.&
DRYING TUBE
INFRA RED CO2 ANALYZER
(a)
AMPLIFIER
(b)
7. CIRCULATING
PUMP
(a) VACUUM
(b) PRESSURE
8. ASCARITE TUBE
9,lO. SHUT-OFF VALVES
2.
3.
4.
5.
6.
I
FIG. 3. Schematic
in scawatcr.
dingram
of cquipmcnt
usccl in the combustion
any seawater sample. The recorder values
so obtained are then plotted, extrapolated
to zero carbon, and the peak height indicated is then used to determine the carbon
in the blank ( Fig, 2). Generally, values obtained by either method for a S-ml sample
were found to bc equivalent to 0.52-0.54
mg C/liter and occupied 17% of the recorder scale set to read 4 mg C/liter at
100% deflection.
and dctcction
of particulate
present in concentrations
of detection.
ANALYSIS
OF PARTICULATE
below
carbon
the limit
CARBON
Recovery
The method for the determination
of
total particulate carbon consists of concentration of the particulate matter from a
l- to 4-liter sample on a glass-fiber filter,
combustion in an automated furnace at
800C in the presence of CuO, using oxygen
as a carrier, and the detection of the resulting COZ by infrared absorption.
No direct criteria for determining
the
relative recovery of carbon from natural
seawater samples were available. Thcrefore, a series of organic compounds was
dissolved in scawatcr at the concentration
specified in Table 1 and the recovery determined. It is apparent from the data that
100% recovery was realized for a large
variety of compounds,
Refractory longchain and polycyclic
hydrocarbons
may
give lower yields, but, because of their
limited
solubility,
they arc presumably
Fig. 3 is a schematic diagram of the
combustion, circulating, and detection units
used for the determination
of particulate
carbon. The combustion unit consists of
the P. H. Coleman Company’s CarbonHydrogen analyzer with modified absorption tubes. The manufacturer’s
recommendations are followed except that the
CO2 absorption tube normally containing
ascarite is filled with Mg( CIO, ),, and a
connection is made at the outlet of this
142
DAVID W. MENZEL
AND RALPH 17. VACCARO
tube to pump the combustion gases into
a Lucite spirometer graduated for volume.
The spirometer is filled with distilled water
containing 5 ml of HCl/liter
which has
been sparged with pure oxygen or nitrogen
Eor one-half hour to remove tracts of CO,
and to prevent subsequent CO2 transfer
over the water-gas interface. On the recorder side of the spirometer, two tubes
are inserted in the gas path, one containing
Mg( C1O4)2 to remove water vapor and the
other containing ascaritc to remove COa
from the system after each analysis. This
tube is fitted with a bypass so that gases
can be circulated without interception during the analysis. The circulation of gases
is achieved with a Neptune Dynapump
Model 2A. The detection unit employed
is the same described above for dissolved
carbon.
Procedure
1) One to 4 liters of seawater are filtered
through a precombusted
( 700C ) 2.$-cm
glass-fiber filter at 12 cm (Hg) vacuum.
2) Filters are dried and preserved in a
vacuum desiccator over silica gel. 3) A filter
is placed in a combustion tube packed according to the directions of the manufacturer of the combustion unit and combusted at 8OOC. 4) The combustion gases
are passed into a spirometer at a flow rate
of 200 ml/min and diluted to 1 liter with
COa-free oxygen. 5) The gas flow is shut
off, the ascarite bypass line is shut off (see
Fig. 3), and the air-circulating
pump is
turned on. After equilibrium
is reached,
indicated by a steady amplified output, the
pump is shut off and the value recorded.
6) To rid the system of COZ in preparation
of the next sample, valve 9 (Fig. 3) is
closed and valve 10 is opened, excess gas
is expelled from the spirometer, and the
circulating pump is turned on so that air
is forced through the ascarite. When the
recorder returns to zero, the next sample
can be introduced.
Standnrdixatiolz and determination
of blanks
Standardization is achieved by injecting
appropriate volumes of COZ gas into the
spirometer, diluting to 1 liter with 02, and
following the procedure described above
(steps 5 and 6). 0 ne ml of COa in 1 liter
of 02 is equivalent to 536 pg C for this
purpose, and the technique we have followed is to set the sensitivity gain on the
amplifier to obtain 100% deflection at this
concentration.
As with the dissolved carbon, this deflection may be set at any point
by adjustment of the sensitivity controls of
the analyzer. When analyses are made on
oceanic water poor in carbon, the scale is
set so that 0.5 ml COz yields 100% deflection, giving a working range of O-268 pg
C and increasing the sensitivity by a factor
of two. A detailed calibration curve must
be obtained using various quantities of CO2
since the response of the recorder is not
linear. Once the shape of the curve is
determined, however, only a single-point
reference calibration is necessary.
Blanks are obtained by combusting a
glass-fiber filter which has been previously
combusted at 700C in a muffle furnace and
rinsed with a small volume of filtered scacarbon
water. Generally the equivalent
measured in these filters is between 15 and
20 E.Lg*
REPERENCES
organic car~UURSMA, E. K. 1961. Dissolved
and phosphorus
in the sea.
bon, nitrogen
N&h. J. Sea Research, 1: 1-141.
1957. Ein Verfahrcn
zum oxyGJLLBRICI-IT, M.
daticvcn Nachweis von organischcr
Substanz
im Sccwnsser. Hclgolaendcr
Wiss. Mccresuntersuch., 6: 76-83.
zur chcmKAY, H. 1954. Einc Micromethoclc
ischcn Bestimmung des organisch Kohlenstoffs
Kicl. Mccresforsch.,
10: 26im Mccrwasscr.
36.
KORRINGA, P., AND I-I. POSITMA. 1957. Investigations into the fertility
of the Gulf of Naples
and acljaccnt salt lakes, with special reference
to shellfish
cultivation.
Pubbl. Staz. Zool.
Napoli, 29 : 229-284.
KROGH, A., AND A. Keys.
1934. Methods for the
determination
of dissolved
organic
carbon
Biol. Bull., 67:
and nitrogen in sea water.
132-144.
VAN HALL,~. E., J.~AFRANKO,ANL) V.A. STENGER
1963. Rapid combustion method for the determination
of organic substances in aqueous
solutions.
Anal. Chem., 35: 315-319.
WILSON, R. F. 196 1. Measurcmcn t of organic
carbon in scn water.
Limnol. Occanog., 6:
259-261.