APPLICATION NOTE
Liquid Chromatography/
Mass Spectrometry
Authors:
Avinash Dalmia
PerkinElmer, Inc.
Shelton, CT USA
Analysis of Naphthenic Acids in
Filtered Oil Sands Process Water
(OSPW) using LC/TOF with
No Sample Preparation
Introduction
Naphthenic acids (NAs) are
water soluble weak acids,
with the general chemical
formula CnH2n+zO2, where
n is the carbon number,
z is referred to as hydrogen deficiency, is a negative even integer and is the number
of hydrogen atoms that are lost as the number of rings in naphthenic acid increases.
More than one isomer will exist for a given z homolog and the molecular weights
differ by 2 mass units (H2) between z-series and by 14 mass units (CH2) between
n-series. The number of possible naphthenic acids, with the same molecular
composition, are 134 with n ranging from 6 to 30 and the number of saturated rings
ranging from 0 to 6. Fig.1 shows examples of the general structures of naphthenic
acids which have a different number of rings1.
with MS, have been reported in the literature, to minimize
the occurrence of false positives and help in structural
differentiation of NAs based on their characteristic HPLC
retention times10-11. The currently reported LC/MS methods
take anywhere from 30 to 60 minutes. The main objective
of this work was to develop a high resolution ( i.e., mass
resolution ~ 10,000), sensitive, specific and faster LC/TOF
MS method to determine the presence and concentration of
various naphthenic acids in oil sand process water using LC/
TOF MS without sample preparation.
Figure 1. General structure of naphthenic acids.
Introduction
Naphthenic acids occur naturally in crude oils and bitumen
and are the primary toxicants in wastewaters associated with
oil refineries and oil sands extraction. Oil sands are large
natural deposits that contain sand, clay, water and bitumen.
Separating the bitumen from the sand requires an extraction
process that uses large amounts of water. This results in
the large production of oil sand process water containing
naphthenic acids.
Naphthenic acids are toxic to animals and aquatic creatures.
They are persistent in the environment because they are
weakly biodegradable. In addition, they cause corrosion in
refinery units and pipelines. Since toxic action and corrosivity
is determined by the structure of the naphthenic acid,
identifying the type of naphthenic acid and its corresponding
amount in oil sand process water is crucial2,3. In the past,
fourier transform infrared (FTIR) spectroscopy and gas
chromatography (GC) have been used to detect NAs in
tailings pond water due to the ease of adapting the existing
methods used in oil-phase detection of NAs. However, it has
been found that FTIR usually overestimates the concentration
of NAs. This overestimation is thought to be due to the
contribution of the carboxylic acid groups of naturally
occurring organic acids e.g. fulvic acids and humic acids in
the aqueous sample. In the GC method, NAs are usually
derivatized to form esters and detected by flame ionization
detection and mass spectrometry (MS). However, the esters
are usually eluted from GC column as an unresolved hump
and characterization of individual NAs is not possible.
Recently, negative ion mode MS with soft ionization sources
such as electrospray ionization (ESI) and atmospheric pressure
chemical ionization (APCI) have been used successfully
for direct characterization of NAs2-9. Negative ion ESI-MS
has emerged as the method widely used for analysis of
NAs due to its higher sensitivity than APCI-MS. Liquid
infusion MS techniques are rapid; however matrix effects
severely hampers sensitivity and therefore they require time
consuming sample preparation techniques. Furthermore,
unit resolution MS methods are prone to giving false
positive results. Only a few methods which couple HPLC
2
Experimental
All measurement were carried out using the PerkinElmer
Flexar™ FX-15 LC with the PerkinElmer AxION® 2 TOF MS with
Ultraspray 2™ Dual ESI source in negative ion mode. A 15 min
LC gradient method using a PerkinElmer Brownlee™ SPP C8
(2.1 mm x 100 mm, 2.7 µm) at a flow rate of 0.3 ml/min was
developed. The gradient method was 45 % mobile phase B held
isocratically for two minutes, followed by the linear gradients
from 45 % to 67 % B in one minute and 67 % B to 100 % B in
seven minutes and held at 100 % B for five minutes. The mobile
phase A and B composition was 90 %/10 % water/methanol
with 10 mM ammonium acetate and 100 % methanol with 10
mM ammonium acetate, respectively. The column temperature
was kept at 30 °C and the sample injection volume was 50 µL.
The AxION 2 TOF was scanned over a mass range of 100-650
m/z with a flight voltage of 8000 V and an acquisition rate of
1.5 spectra/s. The ESI source temperature and drying gas flow
rate was 375 °C and 12 L/min, respectively. A commercial
standard mixture of naphthenic acid (Merichem, Houstan Texas
USA) was diluted to a concentration of 50 ppm and analyzed
using the method described above. The filtered oil sand process
water samples were analyzed with LC/TOF MS without any
sample preparation.
Results
A 15 minute gradient LC method coupled with TOF MS was
developed to separate the standard mixture of naphthenic
acids. The method was used to determine the characteristic
retention time of individual NAs and to determine the
presence and amount of NAs in filtered oil sand process
water samples. Figure 2 and Figure 3 show the extracted ion
chromatograms (EICs) for 134 naphthenic acids from the
standard mixture of NAs and a filtered oil sand process
water sample. Using accurate mass measurements of the
NAs, we were able to identify 111 out of possible 134 NAs
in the standard mixture and 54 NAs in the oil sand process
water samples. Figure 4 illustrates the mass and isotope
profile accuracy of one of the NAs (n=14,z=-4) in an oil sand
process water sample. Using lock mass calibration, the
average absolute mass accuracy of the measurements was
2.19 ppm with a standard deviation of 1.5 ppm.
Figure 5 displays the retention time of the naphthenic acids
as a function of n and z in a standard mixture of NAs. For
the same carbon number, the retention time of the NAs
decreases with the increase in number of rings in the
structure. For the same number of rings in the structure,
the retention time increases with the increase in the
number of carbon atoms. Figure 6 and Figure 7
demonstrate the three-dimensional column graph of
percentage intensity of different NAs in the standard
mixture and an oil sand process water sample. Figure 8 and
Figure 9 present the ring type distribution of the NAs in the
standard mixture and the oil sand process water sample.
The data shows a similar ring type distribution of the
standard mixture and the oil sand process water sample.
Both the standard and the samples showed that the
highest amount of a given ring type of NAs had two rings
and this agrees with what has been reported in the
literature2,6. Based on the response relative to the 50 ppm
standard mixture of NAs, Table 1 lists the amount of the
NAs in three oil sand process water samples. The average
measured amount of NA in the oil sand process water
samples was 19.72 mg/L.
Figure 2. EIC of naphthenic acids in a 50 ppm standard mixture of naphthenic acids.
Conclusions
A 15 minute LC/TOF MS method was developed to
determine the presence and amount of different
naphthenic acids in oil sand process water without sample
preparation. The presence of 111 out of possible 134 NAs
was identified in a NAs standard and 54 NAs were
identified in oil sand process water samples by LC/TOF MS.
The identification of NAs was based on their characteristic
retention time and accurate mass measurements using LC/
TOF MS. The absolute mass accuracy of the measurement
was 2.19 ppm using lock mass calibration. The average
measured amount of total naphthenic acid in oil sand
process water was 19.72 mg/L. Using the LC/TOF MS
method developed in this work, we were able to measure
both total and individual amount of naphthenic acids
without any sample preparation or derivatization. In
comparison, the existing methods, such as GC, FTIR and
direct infusion ESI MS can either measure total naphthenic
acids without identifying naphthenic acids individually or
require time consuming sample preparation and
derivatization steps.
Figure 3. EIC of naphthenic acids in an oil sand process water sample.
Figure 4. Mass accuracy and isotope profile of one of the naphthenic acid in an oil
sand process water sample.
Table 1. Total naphthenic acid concentration in oil sand process water samples
measured using LC/TOF MS.
Oil Sand Process
Water Sample
Total Naphthenic Acid/mg/L
120.13
2
19.50
319.54
Figure 5. Retention time of naphthenic acids as a function of n and z in the standard
mixture of naphthenic acids.
3
Acknowledgements
The author would like to thank Emergencies, Operational Analytical Laboratories and Research Support Division
of Environment Canada for providing oil sand process water samples.
References
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4
Figure 6. Relative percentage intensity of different naphthenic acids in a standard
mixture of naphthenic acids.
Figure 7. Relative percentage intensity of different naphthenic acids in an oil sand
process water sample.
Figure 8. Naphthenic acid response as a function of number of naphthenic acid rings
in a standard mixture of naphthenic acids.
Figure 9. Naphthenic acid response as a function of number of naphthenic acid
rings in an oil sand process water sample.
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