inorganic arsenic in rice using icp-qqqms with and without ...inorganic as (ias) methylated arsenic...
TRANSCRIPT
Inorganic arsenic in rice using ICP-QQQMS with and without chromatography
Jörg Feldmann [email protected]
Trace Element Speciation Laboratory (TESLA) University of Aberdeen
Scotland, UK
Webinar Agilent May 2014
Introduction into arsenic in rice HPLC-ICPMS: method description and results HG-ICPMS: method development and results
Comparison HG-ICPMS/HPLC-ICPMS
Table of content
“…that rice has higher inorganic arsenic concentrations than most other foods, and consequently, diets that rely heavily on
rice may contain the most inorganic arsenic.”
Schoof RA, et al. Fd Chem Toxicol 1999; 37:839-46.
A market basket survey of inorganic arsenic in food.
Arsenic in foodstuff
< 10 ng/g 100 ng/g 1000 ng/g
20,000-200,000
500-1,000
>500,000
1-2
50-90
5-10
220-1,100
1,900
Total Arsenic
Toxicity of arsenic
• Arsenic is a class I carcinogen
– Legislation (WHO, US-EPA, EU,…) : 0.010 mg/L in drinking water
– Food: no regulation, why…
Classes of arsenic species in food commodities
OH
AsOH O
OH
CH3
AsCH3
O
OH
OH
AsOH
OH
CH3
As+CH3
CH3
CH3
Inorganic As (iAs) Methylated arsenic (DMA, Tetra)
CH3
As+
CH3
CH3
CH2
C OH
O
O
CH3 CH3
As
CH3
CH3
O
O OH
OH
Arsenobetaine (AsB) Arsenosugars (AsSugar-OH, -SO4.)
GLU
CYS
GLUCYS
CYS
GLU
AsS S
S
GLY
Arseno-phytochelatins (AsPC3) Arsenolipids (AsHC, AsPL, AsFA)
OAs
CH3
CH3
OH
O
J. Feldmann & EM Krupp: Anal Bioanal Chem (2011), 399, 1735-1741.
Food control: Simplification of Speciation analysis by determining only three fractions of arsenic
1%
1%
18%
78%
2%
98%
Fish Molluscs
Seaweed
69%
1% 1%
1%
28%30%
Rice
toxic (iAs)
non-toxic (AsB)
potentially toxic
arsenic fractions
fat-soluble As
arsenosugars
other organoarsenicals
potentially toxic
2%
85%
5%
1%
7%
13%
5%
50%
13%
23%
9%
45%1 mg/kg
100 mg/kg
3 mg/kg
0.3 mg/kg
Tabloid study with trigger value Risk group: toddler and babies
• Babies have 3 times intake per body weight as
adults mg/kg/d and • Many baby products are based on rice 6 of 17 products could not be exported to
China ( ML: 0.150 mg iAs / kg).
AA Meharg, GX Sun, PN Williams, E Adomako, C Deacon, YG Zhu, J Feldmann, A Raab Environ. Poll. (2008), 152, 746-749
1989 – WHO: PTWI 0.015 mg kg-1 b.w.
1998 – Drinking Water Directive (EU) 98/83/EC (10 µg L-1).
2001 – The Environmental Protection Agency (EPA, U.S.) adopts a new arsenic in drinking water standard (10 µg L-1).
2005 – MLs of iAs and totAs in China for different food commodities (e.g. rice, 0.150 mg kg-1)
2008 – Paper on iAs levels in baby rice published.
2009 – FSA (UK) guidelines recommending no rice milk for young children, similar guidelines given in Denmark.
2009 – EFSA opinion paper on arsenic.
2010 – PTWI of 0.015 mg kg-1 b.w. from 1989 (WHO) was withdrawn by the Joint FAO/WHO committee.
2010 -2011 – Several Proficiency tests by IMEP in food and feed.
2011 – MLs on iAs in food in Australia.
2012 – Proposed draft MLs of iAs in rice by FAO/WHO.
2013 – Rice CRM (ERM - BC211), certified for iAs.
Food regulation – Selected events
…The inorganic forms of arsenic are more toxic as compared to the organic arsenic but so far most of the occurrence data in food collected in the framework of official food control are still reported as total arsenic without differentiating the various arsenic species….
…15 European countries > 100,000 results of As in food - 2/3 < LOD, - only 2 % speciation information
…The highest total arsenic levels were measured in the following food commodities: fish and seafood, … high concentrations in rice grains and rice-based products, …
….. In order to refine risk assessment of inorganic arsenic there is a need to produce speciation data for different food commodities to support dietary exposure assessment and dose-response data for the possible health effects….
As-speciation in rice is simple
OH
OH
As
OH
OH
OH
AsOH
O
OH
OH
AsCH3
O
OH
CH3
AsCH3
O
As(III)
As(V)
MMA
DMA
(CH3)4As+ TETRA (HR Hansen et al. J. Environm. Monit. (2011)
iAs
Introduction into arsenic in rice HPLC-ICPMS: method description and results HG-ICPMS: method development and results
Comparison HG-ICPMS/HPLC-ICPMS
Table of content
Extraction of rice for inorganic arsenic
200 mg rice + 1 % (v/v) HNO3 + 1 % (v/v) H2O2 Microwave oven (Mars5, CEM UK) in Corning tubes, - step 1: 5 min at 50oC, - step 2: 5 min at 75oC, - step 3: 30 min at 95oC. After cooling the samples were centrifuged.
Important for digestion and Oxidation of As(III) As(V)
Agilent 8800 QQQ (ICPMS)
HPLC-ICPMS
• SAX column: PRP X-100; 4.6 * 250 mm was used with 4 g/L (NH4)2CO3 buffer, 1 mL/min at RT; sample volume 50 L.
– Inorganic arsenic (sum of Asiii,Asv)
– Methylated arsenic (mainly DMA)
ICPMS HPLC
0
500
1000
1500
2000
2500
3000
3500
0 200 400 600 800 1000
Inte
nsi
ty
Time (s)
DMA
MMA
As(V)
What is the best method to detect arsenic with an ICPMS?
Arsenic detection in QQQ
As+ ArCl+
Zr+ Co+
O2
As+ AsO+ ArCl+
Zr+ ZrO+ Co+ CoO2
+
AsO+
Zr+
CoO2+
Set to 91u
ICP-MS
Single quadrupole
As+ ArCl+
O2
As+ AsO+ ArCl+
AsO+
Set to 91u
ArCl+, As+.
Set to 75u
Zr+, Co+
ICP-QQQ
Triple quadrupole
removes interfering polyatomics such as ArCl+, but the detecting m/z can still be interfered by newly generated polyatomics or other elements
Removes possible interring elements with Q1 and then removes interfering polyatomics such as ArCl+,
m/z 75: As, 40Ar35Cl
Arsenic detection in QQQ
M1 M2
Cell Q1
Set to M1
Q2
Set to M2
No gas Mode Q1: m/z 75 Q2: m/z 75 75As+
Oxygen mode Q1: m/z 75 Q2: m/z 91 75As16O+
Hydrogen mode Q1: m/z 75 Q2: m/z 75 75As+
Helium mode Q1: m/z 75 Q2: m/z 75 75As+
Ammonia mode Q1: m/z 75 Q2: m/z 91 75As(NH2)+
Q2: m/z 125 75As(NH3)2(NH2)+
O2, He, H2, NH3
ICP-QQQ settings
No gas H2 He O2 H2 + O2 NH3
Gas flow setting
- 2.4 ml/min 3.6 ml/min 30 % 3 mL/min + 30 %
2 mL/min (He) + 28 %
Energy Discrimination
(V)
5.0 5.0 1.0 -7.0 -6.0 -10.0
Wait time offset (ms)
0 0 0 2 2 5
MS-mode MSMS-mode
MS-mode MSMS-mode
MSMS-mode
MSMS-mode
m/z measured (Q2)
35, 75, 103
37, 75, 103 35, 75, 103 51, 75, 91, 103
37, 75, 91, 103
35, 75, 91, 103, 125
m/z 103 was used for the continuous internal standard Rh.
HPLC-ICP-QQQ of a rice extracts using different gas modes
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
0.45
0 2 4 6 8 10 12 14 16
rel. in
tensity (
off
-se
t)
retention time (min)
No gas
H2 6 mL
H2 7 mL
He
O2 (75 75)
O2 (75 91)
As(V) DMA
MMA ?
Check the As species eluting in the void: As(III) or (CH3)4As+
No gas mode or oxygen mode with mass transfer seems best suited
How good is the automatic integration using MassHunter for all gas modes ?
0
20
40
60
80
100
120
140
160
180
200
0 20 40 60 80 100 120 140 160 180 200
µg A
s / k
g r
ice m
anual..
µg As / kg rice MassHunter
H2 a
H2 b
He
no gas
O2 (75->75)
O2 (75->91)
Figures of merit No gas H2 He O2 H2 / O2 NH3
LoD (g/kg)
4.1 3.2 6.7 m/z 75: 3.3 m/z 91: 3.2
m/z 75: 6.6 m/z 91: 3.3
m/z 75: 12 m/z 91: 19
m/z 125: 54
BEC 0.15 0.098 0.099 m/z 75: 0.38 m/z 91: 0.10
m/z 75: 0.26 m/z 91: 0.040
m/z 75: 0.12 m/z 91: 0.32 m/z 125: 1.1
S/N 80 21 38 m/z 75: 10 m/z 91: 76
m/z 75: 35 m/z 91: 110
m/z 75: 15 m/z 91: 13 m/z 125: 4
Average RSD (%)
5.2 3.4 1.9 m/z 75: 6.0 m/z 91: 3.6
m/z 75: 2.9 m/z 91: 2.7
m/z 75: 5.8 m/z 91: 5.4
m/z 125: 7.8
Best method is either no gas or O2 or O2/H2 with As on m/z 91.
Optimisation of the gas modes (rice with high DMA)
All gas modes give the same result
0
20
40
60
80
100
120
140
160
180
200
no gas H2 He O2 (75) O2 (91) H2O2 (75) H2O2 (91) NH3 (75) NH3 (91) NH3 (125)
µg
As /
kg
ric
e
DMA
As(V)
NIST-1568 a
Optimisation of the gas modes (rice with high DMA)
0
20
40
60
80
100
120
140
no gas H2 He O2 (75) O2 (91) H2O2
(75)
H2O2
(91)
NH3
(75)
NH3
(91)
NH3
(125)
µg
As /
kg
ric
e
DMA
As(V)
concentration of the arsenic species control material, n=3
All gas modes give the same result.
No significant interference, hence any ICP-MS can be used.
HPLC-ICPMS analysis of CRM rice samples (n=4)
DMA MMA As(V)
BCR-211 (n=4)
119 +/- 12 15 +/-4.0 120 +/- 6.8
BCR-211 (certified)
119 +/- 13 124 +/- 11
NIST-1568a (n=2)
201 +/- 5.8 7.5 +/- 2.3 113 +/- 2.1
Literature 143–185 80–109
Summary of available methods for the analysis of arsenic in rice from various
countries
HPLC-ICPMS / HG-AFS
HPLC-ICPMS HPLC-ICPMS
HPLC-ICPMS
HPLC-ICPMS
ICPMS
Table taken from [Codex Committee on Contaminants in Food – March 2012 Report]
Analytical challenges with new regulatory guideline
Although HPLC-ICPMS can be used for speciation faster and cheaper and provide speciation information
Any intelligent fool can make things bigger, more complex, and more violent.
It takes a touch of genius -and a lot of courage - to move in the opposite direction.
- (Albert Einstein)
… if you are not a genius just look up old papers….
Introducing a selective step for iAs
– Selective volatilisation using hydride generation (NaBH4): • As(=O)(OH)3 (iAs) AsH3 (-55oC)
• (CH3)As(=O)(OH)2 (MMA) (CH3)AsH2 (+2oC)
• (CH3)2As(=O)OH (DMA) (CH3)2AsH (+35oC)
• AsB, As-sugars, As-lipids no volatile product !
…and reactions show pH dependence!
Selective determination of iAs without chromatography
S. Musil, AH Petursdottir, H Gunnlaugsdottir, EM Krupp, J Feldmann
Analytical Chemistry (2014) 86, 993-999
Standard Agilent generator in CF mode
Vmixer = 0.23 ml
vcarrier gas = 0.3 – 0.4 l min-1 ; Makeup gas introduced downstream the GLS
vNaBH4 = 0.5 ml min-1
vsample = 0.5 ml min-1
vHCl = 2.5 ml min-1
2% NaBH4 in 0.1 M NaOH
High concentration of HCl
HG-ICPMS (wet/dry plasma)
WET plasma
DRY plasma
S. Musil, AH Petursdottir, H Gunnlaugsdottir, EM Krupp, J Feldmann
Analytical Chemistry (2014) 86, 993-999
Hyphenation of HG to ICP-MS(MS) – WET plasma conditions
Influence of HCl in the reaction mixture
5M HCl was chosen to eliminate DMA signal satisfactorily!!
MMA sensitivity 45.4 ± 0.2% of iAs(V) sensitivity
DMA sensitivity 2.2 ± 0.05% of iAs(V) sensitivity
0
100000
200000
300000
400000
500000
600000
700000
800000
0 5 10
Sig
nal in
ten
sit
y (
CP
S)
Concentration of HCl for HG generation (M)
Dependence of signal of As species (1 ppb) on concentration of HCl, results corrected to IS 103Rh, signals corrected to blank
iAs(V)
MAs(V)
DMAs(V)
iAs(V) pre-reduced byKI/ascorbicacid and HCl
S. Musil, AH Petursdottir, H Gunnlaugsdottir, EM Krupp, J Feldmann
Analytical Chemistry (2014) 86, 993-999
Rice samples
Speciation for iAs without chromatography
Rice samplea
HG- ICP-MS
HPLC- ICP-MS
Total As
(certified)
iAs
(reported)
JRC 203 92 ± 4
101 ± 5
JRC 165 93 ± 4
103 ± 3
NIST 1568a
100 ± 6
94 ± 5
290 ±
30
94 ± 12
Concentration of iAs in rice samples (μg kg–1) found:
and HG-ICP-MS three times faster than HPLC-ICP-MS
Samples
from
PT (2011)
S. Musil, AH Petursdottir, H Gunnlaugsdottir, EM Krupp, J Feldmann
Analytical Chemistry (2014) 86, 993-999
Oxygen vs no gas mode using HG-ICPMS
AH Petursdottir, N Friedrich, S Musil, A Raab, H Gunnlaugsdottir, EM Krupp, J Feldmann, Analytical Methods (2014) doi:10.1039/C4AY00423J
No gas mode and oxygen mode show the same results! no ArCl interference despite the high HCL concentration. All ICP-MS models can be used for this application.
Can the results be confirmed for rice with different DMA/iAs ratios?
Samples
• 32 rice samples, bought in local stores in Aberdeen, UK.
• 12 rice samples, grown under arsenic exposure.
Extraction
• Microwave extraction, 1% HNO3 and 1% H2O2.
AH Petursdottir, N Friedrich, S Musil, A Raab, H Gunnlaugsdottir, EM Krupp, J Feldmann, Analytical Methods (2014) doi:10.1039/C4AY00423J
HG (iAs conc g kg-1 )
0 100 200 300 400 500
HP
LC
(iA
s c
onc m
g
g k
g-1
)
0
100
200
300
400
500
95% Confidence Band
95% Prediction Band
44 rice samples (n=3)
Slope = 0.97
AH Petursdottir, N Friedrich, S Musil, A Raab, H Gunnlaugsdottir, EM Krupp, J Feldmann, Analytical Methods (2014) doi:10.1039/C4AY00423J
Conclusions
• HPLC-ICP-QQQ is ideally placed for the determination of iAs in rice
• Every gas mode gives acceptable results highlights highest flexibility and 7700 or 7900 can also be used.
• HG-ICPMS is selective enough to determine only iAs
– HPLC not needed!
– Faster determination than using chromatography
– Straight forward data treatment • no peak integration
• easier SOP
Acknowledgement
• Asta Petursdottir
• Helga Gunnlaugsdottir
• Eva M Krupp
• Stanislav Musil
• Nils Friedrich
• Andrea Raab
• Jenny Nelson
and
• Cornelius Brombach for the artwork
Further events
• HPLC-ICP-QQQ use for sulfur speciation in combination with arsenic in garlic
• including the use of ESI-qTOF
75As 75As16O+
32S 32S16O+
watch that space…
BNASS/TraceSpec Aberdeen, Scotland,
31st Aug- 4th Sept. 2014
BNASS 2014 –
17th Biennial Nat. Atomic Spectros. Symp.
TraceSpec 2014 –
14th Workshop on Progress in Trace Metal
Speciation for Environmental Anal. Chem.
http://www.abdn.ac.uk/events/bnass-tracespec-2014/