10 B

Development of rapid and precise
boron isotope analysis
in whole blood
by HR-ICP-MS
Yurie Yamaguchi
Research and Development Office,
Japan Chemical Analysis Center
Objectives
・To develop precise & accurate measurement method of
concentration and isotope ratios of B by HR-ICP-MS.
・To apply this method to actual blood samples.
・To improve accuracy of analysis while shortening an analysis time.
Measurement methods
Required conditions for
10B/11B isotope ratios
①Convenience, rapidity
②High precision
③High sensitivity
HR-ICP-MS
Satisfy the
above
conditions
JCAC
HR-ICP-MS
Ar4＋&10B
are well
separated
Ar4＋
9.98797
Slight
overlapping
of Ar4＋
on 10B
10B
10.01557
Q-ICP-MS
8.99060
11.01294
Optimization of Analytical Conditions
Prevention of pollution from labware
B (ppb)
>2 ppb boron elution measured by
HR-ICP-MS was obtained using
borosilicate glass labware（Fig. a）.
88
Plastic labwares were used to
avoid potential contamination
from borosilicate glass.
22
Int. Intencity［cps］
00
For an accurate determination of B isotope ratios,
it is necessary to clean the sample introduction
units in order to eliminate the memory effect.
Rinsing time
0.1M HNO3 ⇒Ultrapurewater
： 5min
(continuous rinsing) （Fig. b）
speedup↑
Only Ultrapurewater ：14min
：11min
Only 0.1M HNO3
Ultrapure water
in a glass
volumetric flask
44
*106 8
Rinsing solution
(a)
66
Elimination of boron memory effect
＊Rinsing method＊
4
7
6
5
4
3
2
1
0
00
10
20
10
20
Time［hour］
After introduction
25ppb boron solution
30
30
(b)
0.1MHNO3 Ultrapurewater
2.5min
2.5min
0
1
2
3
4
Time［min］
5
5
Selection of internal standard
Y
0.3
R² = 0.9999
0.2
10B/Y
R² = 0.9993
0.1
0.0
0
10
20
30
40
Concentration(ppb)
Intensity ratio
0.8
Sc
0.6
8.0
11B/Y
11B/Be
Be
6.0
R² = 0.9984
4.0
10
R² = 0.9988 B/Be
2.0
0.0
0
10
20
30
40
50
Concentration(ppb)
11B/Sc
R2= 0.9990
0.4
10B/Sc
0.2
Intensity ratio
Intensity ratio
0.4
High correlation coefficient
and accuracy were obtained
using Y standard solution
as internal standard.
R² = 0.9997
0.0
0
10
20
30
Concentration(ppb)
40
Y was chosen as the internal
4
standard for the B determination.
Optimization of Measurement Conditions
6
【Comparison between HR-ICP-MS measuring at Low and Medium resolution】
Resolution Sensitivity
【Low】
【Medium】
★Sensitivity
★Accuracy
◎
○
Accuracy
Spectral Interference
◎
○
△
○
Signal intensity decreases as the resolution increases due to narrow slit.
★Spectral Interference
【L】
【L】
【M】
At Low-resolution measurement, providing
flat-topped peaks and maximum
instrument accuracy.
【M】
40Ar4+ 10
B
40Ar4+
10B
10B
is hindered by 40Ar4+ interference
at m/z = 10 at low resolution.
●The precision was improved when the mass scanning window
was reduced from 20%.
●In this study, isotope measurements for boron determination were
carried out at low resolution(R=300)for accurate measurement.
Measurement of 10B/11B
◆Mass bias correction
●In measuring isotope ratios by ICP-MS, measured values will
deviate from true values due to mass bias effect.
●Therefore, mass bias correction is important for highly accurate
isotope ratios determination.
●A correction factor is calculated as the mean of the measured
isotope ratios of the standard of known isotopic composition,
compared to the measured isotope ratios.
10B/11B
(𝒄𝒆𝒓𝒕𝒊𝒇𝒊𝒆𝒅 𝒗𝒂𝒍𝒖𝒆)
Mass bias factor ＝ 10 11
B/ B(𝒎𝒆𝒂𝒔𝒖𝒓𝒆𝒎𝒆𝒏𝒕 𝒗𝒂𝒍𝒖𝒆)
Katahira (2005),
Nagao (2011)
◆Validation of mass bias factor
1. Calculation of mass bias factor using the standard of known isotopic
composition.
2. Measurement of B isotope ratios of the standard of known
concentration.
→Correction using mass bias factor calculated in 1
→Comparison between correction value and natural abundance of B isotope ratios
②Measurement of B isotope ratios
B isotope ratios of NIST-SRM 1643e (10B/11B=0.2484*) was measured
using mass bias correction(n=3).
n= Number of analyses performed on
each sample(average values shown).
NIST-SRM 1643e, standard multi-element solution was used to validate of the isotopic ratio measurements
10B/11B
10B/11B
before correction
0.229
after correction
RSD
(%)
Relative error
(%)
0.247
0.867
－0.75
×1.077
Mass bias factor
*IUPAC Inorganic Chemistry Division, CIAAW(2011)
【Reference values】
Boron Isotope Ratio Analysis on the ELEMENT2（measured by Thermo Fisher Scientific）
★Relative error ：1.31%, ★RSD value ：0.062%（n=5)
●Determined B isotope ratios were in good agreement with the
certified value. (The difference between certified values and
measurement values were better than ±1%.)
●The RSD values obtained for B isotope ratios was better than 1%.
Measurement of B concentration and
10B/11B isotope ratios of BSH and BPA
Difference from
evaluated value(%)
30
Concentration
-5 0
25
15
10
-15
BPA
10
5
15
20
25
30
BSH
-20
>2%
-25
>0.2ppb
-30
-5 0
5
10
15
20
25
30
B (ppb)
100
●：✔
●：×
100
>2.4ppb
>2.9ppb
Isotope ratios
95
>98.8%
90
95
>98.9%
90
85
85
BPA
/
totalB(%)
5
-10
20
0
10B
B (ppb)
0
80
BSH
80
75
75
0
5
10
15
B (ppb)
20
25
30
0
5
10
15
20
B (ppb)
25
30
Determination of B in simulated plasma
Before using whole blood, simulated plasma was used to confirm the effect
of matrix elements concentration on the determination of B.
〈Changes of concentration of B
and matrix elements in plasma〉
3600
～
～
Concentration(ppm)
3200
～
～
150 ～
～
100
40
35
30
25
20
15
10
5
0
×272
×109
～
～ Infusion
Cl
B : 5ppb
Na
Matrix
elements
K
Ca
BNCT
BNCT
■ Plasma
▲ Whole blood
● Erythrocytes
0
〈Methods 〉
Simulated
plasma
Matrix elements were added in
stages while keeping constant the
Mg
concentration of boron as 5ppb.
P
50 100 150 200 250 300 350
Time after onset of infusion(min)
LAAKSO, 2001
～e.g. Na solution～
Na-conc/
B-conc
B-conc
(ppb)
Na-conc
(ppb)
Cl-conc
(ppb)
0
25
50
5
5
5
100 200 300
5
5
5
0
125 250 500 1000 1500
0
193 385 771 1542 2313
B concentration (n=3)
Difference from evaluated value(%)
10
10
10
◆：Na
◆：Cl
5
5.00
0.00
0
0
100
200
300
400
500
Na/B
Cl/B
-5
-5.00
10
10
◆：K
◆：Cl
55
00
0
-10
-10
◆：Na
◆：Cl
55
00
0
100
-5
-5
200
300
Na/B
Cl/B
400
500
-10
-10
-10
-10.00
-5
-5
Difference from natural abundance(%)
10.00
B isotope ratios (n=3)
100
200
300
K /B
Cl/B
400
500
10
10
◆：K
◆：Cl
55
00
0
-5
-5
-10
-10
100
200
300
K /B
Cl/B
400
500
10
◆：Ca
5
0
0
100
200
300
400
500
-5
PP
(P/B=25)
B isotope ratios (n=3)
10
◆：Ca
5
0
0
100
200
300
400
500
-5
-10
-10
Mg
Mg
(Mg/B=25)
Difference from natural abundance(%)
Difference from evaluated value(%)
B concentration (n=3)
Difference from
evaluated value(%)
RSD(%)
Difference from
evaluated value(%)
RSD(%)
0.04
0.16
0.49
1.10
Difference from natural
0.20
abundance(%)
(Mg/B=25)
RSD(%)
1.32
Difference from natural
P
-0.12
P
abundance(%)
(P/B=25)
RSD(%)
0.67
Mg
Mg
Accurate and precise values of B concentration and B
isotope ratios could be obtained despite the presence of
high concentration of matrix elements.
Determination of B in whole blood
To find optimum digestion method of whole blood sample,
B recovery test using hotplate digestion and microwave digestion
were performed by the following method.
Hotplate digestion
Teflon
Teflon
Microwave digestion
Whole Blood
conc. HNO3
B/BPA/BSH
Teflon
Quartz
Quartz
Hotplate
digestion
HR-ICP-MS
measurement
Microwave digestion
(Ethos TC, Milestone)
■：Hotplate (Digestion procedure×3, Measurement×3)
■：Microwave (Digestion procedure×3, Measurement×3)
Results～closed system～
80
80
60
/ totalB(%)
100
Recovery
40
10B
Recovery(%)
100
60
20
20
0
0
B
BPA
BSH
Teflon
B
BPA
Isotope ratio
40
BSH
Quartz
Recovery ■■
B
BPA
BSH
Teflon
10B
/ totalB isotope ratios ■■
As for B, good agreement with the natural abundance.
As for BPA/BSH, favorable B isotope ratios were obtained.
Comparison between Teflon and Quartz inserts ■
Thermal conductivity ：Teflon << Quartz
Handling
：Teflon > Quartz
Ease of rinsing
: Teflon << Quartz
BPA
Quartz
Microwave digestion
>98% with low RSD expect for BSH solutions.
B
BSH
Results～open system～
There is a possibility of boron analyte loss in the case of open system digestion
due to the chemical property of boron volatility.
Thus, it is necessary to explore relationship between
【Recovery－Sample amount】 and 【Recovery ―Digestion time】
Sample amount
0.05g
0.05g
Heating
time
Teflon
28min.
28min.+20min.(After completion of digestion) 45min.
Quartz
22min.
22min.+20min.(After completion of digestion) 30min.
80
80
Recovery
40
■ ■ ：0.05g
■
：0.1g
20
0
B
BPA
Teflon
BSH
B
BPA
Quartz
BSH
10B
60
/ totalB(%)
100
Recovery(%)
100
60
0.1g
Isotope ratio
40
■ ■ ：0.05g
■
：0.1g
20
0
B
BPA
Teflon
BSH
B
BPA
Quartz
BSH
Results～open system～
Comparison between Teflon and Quartz inserts
Thermal conductivity : Teflon << Quartz
Handling
: Teflon < Quartz
Ease of rinsing
: Teflon << Quartz
Sample amount/
Heating time
― independent
Depends on
sample amount and
heating time.
80
80
Recovery
40
■ ■ ：0.05g
■
：0.1g
20
0
B
BPA
Teflon
BSH
B
BPA
Quartz
BSH
10B
60
/ totalB(%)
100
Recovery(%)
100
60
Isotope ratio
40
■ ■ ：0.05g
■
：0.1g
20
0
B
BPA
Teflon
BSH
B
BPA
Quartz
BSH
【Relationship between recovery and heating time】…Focus on the case of using quartz as
vessel.
100
After completion
100
75
B
(Quartz)
50
25
◆
◇
◆
◆
Recovery(%)
Recovery(%)
of digestion
：0.05g
：0.05g
：0.1g
：0.5g
0
BPA
(Quartz)
50
25
◆
◇
◆
◆
：0.05g
：0.05g
：0.1g
：0.5g
0
0
20
40
60
Heating time(min.)
80
0
20
40
60
Heating time(min.)
80
【The degree of boron Volatilization rate】
100
Recovery(%)
75
B > BPA >>> BSH
75
BSH
(Quartz)
50
25
◆
◇
◆
◆
：0.05g
：0.05g
：0.1g
：0.5g
0
0
20
40
60
Heating time(min.)
80
BSH has a greater binding energy than
that of B4H16N2O11 (Rational formula of B
standard solution) and BPA.
Thus, B-B binding is hard to be dissociated,
and therefore, volatilization of boron is
hardly occurred.
【Relationship between recovery and heating time】…Focus on the case of using quartz as vessel.
After completion
of digestion
75
B
(Quartz)
50
25
◆
◇
◆
◆
100
Recovery(%)
Recovery(%)
100
：0.05g
：0.05g
：0.1g
：0.5g
0
75
BPA
(Quartz)
50
25
◆
◇
◆
◆
：0.05g
：0.05g
：0.1g
：0.5g
0
0
20
40
60
Heating time(min.)
80
Sample amount Boron loss
0
20
40
60
Heating time(min.)
80
Sample amount
Heating time
Boron loss
◆
0.05g
-2.8%
◇
0.05g
40min.
-6.8%
◆
0.1g
-9.1%
◆
0.1g
30min.
-9.1%
◆
0.5g
-34.2%
Boron analyte loss occurs
Volatilization rate of boron is higher
with the process of blood
during digestion than after digestion.
digestion.
It is therefore necessary to pay attention to the volatilization of boron
using hotplate digestion in open system
【Methods comparison】
Digestion methods
Hotplate digestion
Microwave digestion
System
open
closed
Preparation time
(Teflon/Quartz)
~1min./~1min.
~3min./－
~6min./~6min.
Digestion time
(0.05g, Teflon)
~28min.
~15min.
~25min.
Cooling time
(Teflon/ Quartz)
7min./3min.
10min./－
20min./20min.
Portability
◎
△
Handling
△
◎
Safety
△
○
○
Contamination
×
◎
Vaporization
△
◎
Homogeneity of
digestion
×
○
Reliability of
digestion
△
○
【Methods comparison】
Digestion methods
Hotplate digestion
Microwave digestion
System
open
closed
Preparation time
(Teflon/Quartz)
~1min./~1min.
Digestion time
(0.05g, Teflon)
Cooling time
(Teflon/ Quartz)
Portability
~3min./－
~6min./~6min.
【Hotplate-open】
✔Complicated operation
~28min.
~15min.
~25min.
is not necessary.
×Boron
analyte
loss may occur
7min./3min.
10min./－
20min./20min.
when it takes a long time
to complete the digestion.
◎
△
△
Handling
◎
Safety
△
○
○
Contamination
×
◎
Vaporization
△
◎
Homogeneity of
digestion
×
○
Reliability of
digestion
△
○
【Methods comparison】
【Hotplate-closed】
Digestion methods
Hotplate digestion
✔Enable to carry out digestion
System
open
within
the shortest time.
Preparation
~1min./~1min.
×There
is atime
possibility
that the ~3min./－
(Teflon/Quartz)
decomposition efficiency
Digestion time
~28min.
~15min.
depends
on a place for
(0.05g, Teflon)
installation.
Cooling time
(Teflon/ Quartz)
7min./3min.
10min./－
Microwave digestion
closed
~6min./~6min.
~25min.
20min./20min.
Portability
◎
△
Handling
△
◎
Safety
△
○
○
Contamination
×
◎
Vaporization
△
◎
Homogeneity of
digestion
×
○
Reliability of
digestion
△
○
【Methods comparison】
Digestion methods
Hotplate digestion
Microwave digestion
System
open
closed
Preparation time
(Teflon/Quartz)
~1min./~1min.
Contamination
×
◎
Vaporization
△
◎
~3min./－
~6min./~6min.
【Microwave-closed】
Digestion✔With
time microwave
~28min. digestion,
~15min.
~25min.
(0.05g, Teflon)
the samples are enclosed,
Cooling time
7min./3min.
10min./－
20min./20min.
so cross contamination
and
(Teflon/ Quartz)
loss of volatiles are eliminated.
◎
△
Portability
×Rapidity is required for the
△
◎
Handling
clinical BNCT, thus it is necessary
△
○
○
Safety to further shorten
the cooling
time.
Homogeneity of
digestion
×
○
Reliability of
digestion
△
○
Conclusions
●This work has clearly shown that precise and
accurate B ratios can be measured by HR-ICP-MS.
●Analytical method has been successfully applied to
①the determination of B in BPA expect for BSH.
②the determination of B in blood sample.
● Digestion method in closed system was applied to the
determination of B in whole blood samples.
● It is more necessary to develop more rapid and efficient
method suitable for clinical applications
by the following methods.
【Hotplate-close】 To improve the efficiency of digestion.
【Hotplate-open】 To expand a range of sample amount.
【Microwave】
To further shorten the cooling time.
Acknowledgement
This work was supported by JSPS KAKENHI
Grant Number 26870866.

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