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Chapter 3 Quantitative Analysis of Inorganic Phosphates Using P NMR Spectroscopy 31
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Janice K Gard, David R. Gard, and Clayton F. Callis
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Monsanto Company, 800 North Lindbergh Boulevard, St. Louis, MO 63167
Phosphorus-31 nuclear magnetic resonance ( P NMR) is optimized with respect to accuracy, precision, and analysis time for the characterization of inorganic phosphates. Species determinations in commercial sodium tripolyphosphate are routinely achieved with an accuracy and precision (0.1-0.5%) comparable to that obtained by chromatographic methods as determined in interlaboratory analyses. The method has been completely automated using a robotic sample changer and an algorithm for data analysis. In a demonstration of the precision attainable, the hydrolysis kinetics of tripolyphosphate is obtained with a corre lation coefficient of 0.998. Extension of P NMR to the analysis of higher oligophosphate mixtures (i.e. sodium phosphate glass) has recently been examined using homonuclear 2DJ -resolved spectroscopy to separate the coupling constant and chemical shift information. Semi -quantitative analyses are achieved using curve deconvolution. 31
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The ubiquitous nature and broad importance of phosphates demands exacting analytical methods for their charac terization. Phosphorus-31 nuclear magnetic resonance ( P NMR) has been used as a method for the quantitative analysis of small inorganic phosphates (1-4). Several potential advantages are offered by P NMR including observation of only the phosphorus-containing species, structural information which may complement or aid Retired 3l
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0097H5156/92/0486-0041$06.00/0 © 1992 American Chemical Society In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
PHOSPHORUS CHEMISTRY
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q u a n t i t a t i o n , and q u a n t i t a t i o n w i t h an e l e m e n t a l a s o p p o s e d t o a m o l e c u l a r s t a n d a r d . L i m i t a t i o n s o f p h o s p h o r u s - 3 1 NMR include sensitivity, complexity of spectra for oligophosphates higher than tripolyphosphate, and, o c c a s i o n a l l y , slow r e l a x a t i o n times. L i t t l e appears i n the l i t e r a t u r e , however, c o n c e r n i n g t h e p r e c i s i o n o r a c c u r a c y of p h o s p h a t e a n a l y s i s by P NMR. Recent advances i n instrumentation have revolutionized NMR spectroscopy, p a r t i c u l a r l y w i t h r e s p e c t t o s e n s i t i v i t y and r e s o l u t i o n , and h a s p r o m p t e d t h i s r e e x a m i n a t i o n o f t h e q u a n t i t a t i v e c a p a b i l i t i e s of P NMR. 31
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F u r t h e r e n h a n c e m e n t s o f t h e NMR method h a v e b e e n carried out in our laboratory by optimization of experimental parameters with respect to accuracy, precision, and analysis time u s i n g commercial sodium tripolyphosphate (STP). The NMR technique was then directly compared with chromatographic methods in a c o n t r o l l e d i n t e r l a b o r a t o r y study w i t h t h e use o f L o r e n t z i a n l i n e s h a p e a n a l y s i s t o improve p r e c i s i o n y e t f u r t h e r . The a c c u r a c y and p r e c i s i o n o f t h e NMR method i s i l l u s t r a t e d by the high value of the correlation coefficients in monitoring the kinetics of hydrolysis of sodium tripolyphosphate. With the i n s t a l l a t i o n of a r o b o t i c sample changer and customized robotic software the efficiency of NMR quantitation i s greatly enhanced. Demonstration of the complete automation of NMR quantitative analysis i s herein described, including f u l l d a t a r e d u c t i o n and g e n e r a t i o n o f t h e a n a l y t i c a l r e p o r t . E f f o r t s a r e c u r r e n t l y underway t o e x p a n d t h e u t i l i t y o f P NMR t o q u a l i t a t i v e and q u a n t i t a t i v e a n a l y s i s o f much more c o m p l e x o l i g o p h o s p h a t e m i x t u r e s . A n o v e l a p p l i c a t i o n of homonuclear two-dimensional J-resolved (2DJ) s p e c t r o s c o p y o f sodium polyphosphate g l a s s i s shown t o effectively yield p- p decoupled spectra. Used in c o n j u n c t i o n w i t h L o r e n t z i a n l i n e s h a p e a n a l y s i s and c u r v e deconvolution, semiquantitative analyses of these mixtures has been a c h i e v e d . 3l
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A n a l y t i c a l Methods One-Diaensional Q u a n t i t a t i v e "P NMR Spectroscopy. S p e c t r a were c o l l e c t e d o n a u t o m a t e d V a r i a n XL-200 o r VXR300S Fourier transform NMR spectrometers, operating at p h o s p h o r u s f r e q u e n c i e s o f 80.98 and 121.42 MHz. The p h o s p h a t e s were t y p i c a l l y p r e p a r e d a s 2-5 w e i g h t p e r c e n t i n D 0 w i t h t h e pH m a i n t a i n e d n e a r 9 i n o r d e r t o o p t i m i z e t h e s i g n a l s e p a r a t i o n and t h e l o n g i t u d i n a l r e l a x a t i o n t i m e s (5,6). T v a l u e s were d e t e r m i n e d u s i n g t h e F a s t I n v e r s i o n Recovery Fourier Transform (FIRFT) method (7) and a c q u i s i t i o n p a r a m e t e r s were o p t i m i z e d t o m a x i m i z e the observable magnetization with r e s p e c t t o a n a l y s i s time ( 8 ) . S p e c t r a were a c c u m u l a t e d u s i n g a 20-25 d e g r e e p u l s e w i d t h , 0.50 sec a c q u i s i t i o n time, a 3 t o 5 second repetition 2
1
In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
3.
GARD ET AL
Quantitative Analysis of Inorganic Phosphates
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d e l a y , g a t e d W a l t z d e c o u p l i n g , c o l l e c t i o n o f 16K data p o i n t s p r i o r t o z e r o - f i l l i n g t o 32K p o i n t s , and a p p l i c a t i o n of a 1.0 Hz exponential line broadening. In the i n t e r l a b o r a t o r y a n a l y s e s 1024 t r a n s i e n t s were g e n e r a l l y c o l l e c t e d r e q u i r i n g about 1 h r . of t o t a l scan time. A spectrum o f commercial sodium t r i p o l y p h o s p h a t e a c q u i r e d u n d e r t h e s e c o n d i t i o n s a t 80.98 MHz i s shown i n F i g u r e 1. N o n - l i n e a r l e a s t s q u a r e s L o r e n z i a n l i n e s h a p e a n a l y s i s was a l s o e x a m i n e d f o r c u r v e f i t t i n g and i n t e g r a t i o n s i n t h e interlaboratory analysis (9).
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R o b o t i c P NMR Assays. S p e c t r a were c o l l e c t e d on a V a r i a n VXR300S s p e c t r o m e t e r equipped with a Varian automatic s a m p l e management s y s t e m ( r o b o t ) w i t h a 50 s a m p l e t r a y u s i n g t h e parameters d e s c r i b e d above. Customized s o f t w a r e was d e v e l o p e d f o r r o b o t c o n t r o l p r o v i d i n g f u l l a u t o m a t e d work-up o f data including the calculation of weight p e r c e n t s and g e n e r a t i o n o f a n a l y t i c a l r e p o r t s . 31
Two-Dimensional S e m i q u a n t i t a t i v e P NMR. S p e c t r a were c o l l e c t e d on a V a r i a n U n i t y 400 F o u r i e r t r a n s f o r m NMR s p e c t r o m e t e r a t 161.90 MHz. The p h o s p h a t e s were p r e p a r e d a t 3-5 w e i g h t p e r c e n t i n w a t e r and a D 0 i n s e r t u s e d f o r l o c k i n g purposes. H o m o n u c l e a r 2 D J - r e s o l v e d s p e c t r a were a c c u m u l a t e d u s i n g an 8K Χ 0.2K d a t a s e t w i t h an a c q u i s i t i o n t i m e i n t h e F d i m e n s i o n o f 0.946 s e c , f o u r s t e a d y s t a t e pulses, 128 transients, and 200 increments i n the F domain. S p e c t r a were a n a l y z e d w i t h z e r o - f i l l i n g t o 16K X 0.5K and a p p l i c a t i o n o f a s i n e b e l l o r s h i f t e d s i n e b e l l w e i g h t i n g f u n c t i o n on a Sun M i c r o s y s t e m s S p a r c 1+ c o m p u t e r . 2
2
A
Chromatography. Separations by ion exchange column c h r o m a t o g r a p h y were p e r f o r m e d a c c o r d i n g t o ASTM methods (10,11). The T e c h n i c o n A u t o A n a l y z e r ( B r a n and L u e b b e ) (12,13) was e m p l o y e d f o r a n a l y s i s o f t h e e l u e n t f r o m ASTM method D 2761 ( 1 1 - 1 3 ) . ASTM method D 501 was f o l l o w e d a s w r i t t e n , u s i n g t h e ammonium m o l y b d a t e c o l o r i m e t r i c a n a l y s i s of the h y d r o l y z e d f r a c t i o n s .
Interlaboratory Tripolyphosphate
Analyses
of
Commercial
Sodium
An interlaboratory s t u d y was conducted to assess the capabilities of standard ion-exchange chromatographic methods and NMR a t 80.9 MHz f o r t h e a n a l y s i s o f sodium t r i p o l y p h o s p h a t e . Three well-mixed, but s e p a r a t e , samples o f c o m m e r c i a l s o d i u m t r i p o l y p h o s p h a t e were s u b m i t t e d t o each l a b o r a t o r y i n t r i p l i c a t e , r e p r e s e n t i n g t h r e e d i f f e r e n t levels of tripolyphosphate. P r o t o c o l was maintained c o n s i s t e n t among m e t h o d s / s i t e s . The r e s u l t s c o m p a r i n g t h e methods a r e p r e s e n t e d i n T a b l e I ( G a r d , D.R.; B u r q u i n , J . C . ; G a r d , J.K.; submitted for publication.). The A u t o A n a l y z e r and t h e NMR analyses were e a c h p e r f o r m e d by two d i f f e r e n t a n a l y s t s ; r e s u l t s by
In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
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p o -
L
Ρ,Ο '3»9
3-
4
2
0
-4
-6
-8
-10
-12
-14
-16
F i g u r e 1. A sample phosphorus-31 NMR spectrum ( a t 80.98 Hz) of a phosphate mixture c o n t a i n i n g p r i m a r i l y sodium t r i p o l y p h o s p h a t e .
-2
-18
-20
ppm
• • • • ( ( • • • l > i > ( | i i i i l i i i i | i i i i | i i i i | i i i i | i i i > | i i i i | i i i i | i i i i | i i i i | i i i i | i i i T | i r T i | i i i i | i i i i ( i i i i | i r r i | r T T i j T i i i | i r T i | i i i i | T i i i | i i T i f r i i r j i i i i ] i
ΡΟ4
Réf.
Ρ, ο 10
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d O S w
O
W
o
3
3. GARD ET A L
Quantitative Analysis ofInorganic Phosphates
each a n a l y s t a r e l i s t e d s e p a r a t e l y i n Table s h a p e a n a l y s i s a n d c u r v e f i t t i n g r o u t i n e was one s e t o f NMR d a t a (row 4 o f T a b l e I ) ; t h e s e p r e s e n t e d i n row 6 o f T a b l e I . IECC d e s i g n a t e s D 501.
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I . The l i n e employed on results are ASTM method
Table I I n t e r l a b o r a t o r y Assay
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% Total
o f Sodium
Tripolyphosphate
P A as-
M e a n / S t a n d a r d D e v i a t i o n o f 9 Samples TripolyPyroOrthophosphate phosphate phosphate
Method AutoAn. AutoAn. IECC P NMR* P NMR* P NMR** 31
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X
0.31/0.09 0.25/0.11 0.55/0.16 0.35/0.11 0.36/0.15 0.51/0.28
7.36/0.73 6.60/0.38 7.66/0.17 7.35/0.48 7.20/0.56 6.68/0.05
92.19/0.78 92.84/0.34 91.03/0.23 91.86/0.51 92.09/0.61 92.33/0.35
Trimetaphosphate 0.14/0.12 0.30/0.03 0.73/0.10 0.44/0.14 0.37/0.18 0.51/0.17
* Separate i n t e g r a t i o n o f t h e t r i p o l y p h o s p h a t e d o u b l e t and the pyrophosphate s i n g l e t ** U s i n g
line
shape a n a l y s i s and c u r v e
fitting
T w o - t a i l e d t e s t s o f t h e n u l l hypothesis a r e used t o i d e n t i f y s i g n i f i c a n t d i f f e r e n c e s among t h e methods ( 1 4 ) . The I E C d e t e r m i n a t i o n s a r e c o n s i s t e n t l y t o o n e e n d o f t h e range o f r e s u l t s . The d e t e r m i n a t i o n o f t r i p o l y p h o s p h a t e by I E C i s l o w e r t h a n f o r t h e o t h e r methods w h i l e t h a t f o r the other species i s higher. H y d r o l y t i c degradation during the IEC a n a l y s i s would account f o r t h i s d i f f e r e n c e s i n c e tripolyphosphate i s t h e s p e c i e s most susceptible t o h y d r o l y s i s . T h i s hypothesis i s independently supported by t h e f a c t t h a t l i t t l e o r no orthophosphate (ahydrolysis p r o d u c t ) c o u l d b e d e t e c t e d b y NMR i n s e p a r a t e a n a l y s e s o f t e t r a s o d i u m pyrophosphate (£0.1%, d e t e c t i o n l i m i t -0.01%), w h i l e s i g n i f i c a n t c o n c e n t r a t i o n s (0.5%) were o b s e r v e d b y I E C . D i f f e r e n c e s i n t h e a c c u r a c y between d i r e c t a b s o r b a n c e measurements f o r ASTM D 501 v s p e a k i n t e g r a t i o n f o r t h e A u t o A n a l y z e r may a l s o b e i m p o r t a n t . S i g n i f i c a n t d i f f e r e n c e s i n the AutoAnalyzer results are -1% d i f f e r e n c e i n t h e d e t e r m i n a t i o n o f t h e major s p e c i e s between t h e two s e t s o f d a t a . This i s mainly t h o u g h t t o r e f l e c t e r r o r i n peak i n t e g r a t i o n ; a l t h o u g h t h e means o f t h e A u t o A n a l y z e r d é t e r m i n a t i o n s f o r p y r o - a n d tripolyphosphate are closer i nthe f i r s t interlaboratory analysis, the standard d e v i a t i o n i s larger. On t h e o t h e r
In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
PHOSPHORUS CHEMISTRY
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h a n d , t h e two s e t s o f r e s u l t s f r o m P NMR using simple e l e c t r o n i c i n t e g r a t i o n a r e i n c l o s e agreement w i t h each other, e s p e c i a l l y c o n s i d e r i n g d i f f e r e n t a n a l y s t s performed t h e a n a l y s e s . R e s u l t s b y P NMR a g r e e w e l l w i t h one s e t o f the AutoAnalyzer r e s u l t s , the only s i g n i f i c a n t d i f f e r e n c e being i n the determination of trimetaphosphate. 31
The p r e c i s i o n f o r the determination of the major p h o s p h a t e s p e c i e s was s i g n i f i c a n t l y increased with the a p p l i c a t i o n o f l i n e s h a p e a n a l y s i s . The t r i p o l y p h o s p h a t e c o n t r i b u t i o n i s , however, w e i g h t e d more a t t h e e x p e n s e o f p y r o p h o s p h a t e u s i n g l i n e s h a p e a n a l y s i s and c u r v e f i t t i n g f o r t h e P NMR, c o m p a r e d t o manual i n t e g r a t i o n s . More accurate determinations are achieved s i n c e curve fitting b e t t e r a c c o u n t s f o r s i g n a l o v e r l a p , b a s e l i n e n o i s e , and contribution of the wings of the Lorentzian peaks, e s p e c i a l l y f o r the lower c o n c e n t r a t i o n s p e c i e s (15,16).
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A p p l i c a t i o n o f "P
NMR
to Kinetics 31
The h i g h p r e c i s i o n a n d a c c u r a c y o f P NMR f o r t h e a n a l y s i s of sodium t r i p o l y p h o s p h a t e i n d i c a t e d t h e t e c h n i q u e might a l s o be s u i t a b l e f o r a p p l i c a t i o n t o p r e c i s e k i n e t i c s t u d i e s involving oligophosphates. The h y d r o l y t i c d e g r a d a t i o n o f 0.035 M sodium t r i p o l y p h o s p h a t e was examined in the p r e s e n c e o f v a r i o u s c o n c e n t r a t i o n s o f Me NCl a t 60.3°C. The h y d r o l y s i s was f o l l o w e d o v e r a p p r o x i m a t e l y two h a l f - l i v e s . Even w i t h l a r g e changes i n t h e r e l a t i v e c o n c e n t r a t i o n s o f phosphate s p e c i e s over time, e x c e l l e n t c o r r e l a t i o n f o r p s e u d o - f i r s t order k i n e t i c p l o t s are obtained allowing d i s t i n c t i o n s t o be made among t h e r a t e s u n d e r d i f f e r e n t Me NCl c o n c e n t r a t i o n s ( F i g u r e 2 ) . The h i g h c o r r e l a t i o n f o r t h e s e p l o t s i l l u s t r a t e s t h e combined h i g h a c c u r a c y and p r e c i s i o n o f t h e NMR method ( T a b l e I I ) . Values f o r the r a t e c o n s t a n t s a r e c o n s i s t e n t w i t h t h o s e o f Watanabe e t a l (17) and C r o w t h e r a n d Westman ( 1 8 ) . 4
4
Table II H y d r o l y s i s o f Sodium T r i p o l y p h o s p h a t e w i t h T e t r a m e t h y l a m m o n i u m C h l o r i d e a s F o l l o w e d by (35mM STP,
Me.NCI 0 0.43 1.38
(M}
10
3
k
pH
9.0,
fhr-M
7.56 7.29 6.83
Added Ρ NMR
60.3°C) Correlation coeff. (R) 0.998 0.998 0.998
In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
Quantitative Analysis of Inorganic Phosphates
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3. GARD ETAL
47
In ( [STP]o / [STP]t ) 1.6 3
4
1.2
+ χ
1
M MeNCI k(10 )hr 7.56 0.00 0.43 7.29 1.38 6.83
R 0.998 0.998 0.998
^
^ ^ ^ ^ ^ ^ ^ ^
0.8
0.4
I
ι
600
1000 Hours
l—
1500
2000
F i g u r e 2. P s e u d o - f i r s t order k i n e t i c s p l o t o f t h e h y d r o l y s i s o f 35 mM sodium t r i p o l y p h o s p h a t e (STP) as followed by P NMR (pH 9.0, 60.3°C) i n the presence o f v a r i o u s concentrations o f Me NCl. Me NCl concen t r a t i o n s , observed r a t e constants, and c o r r e l a t i o n c o e f f i c i e n t s (R) are given i n the i n s e t . 31
4
4
American Chemical Society Library 1155 16th St., N.W.
In Phosphorus Chemistry; Walsh, E., et al.; Washington, D.C. ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
PHOSPHORUS CHEMISTRY
48 Robotic
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P NMR
A s s a y s o f Sodium T r i p o l y p h o s p h a t e
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31
A l t h o u g h P NMR y i e l d s r a p i d , r e p r o d u c i b l e q u a n t i f i c a t i o n of o l i g o p h o s p h a t e s , q u a n t i t a t i v e s p e c t r a r e q u i r e l o n g e r accumulations than simple survey spectra. Manual d a t a r e d u c t i o n o f t h e s p e c t r a i s a s u b j e c t i v e p r o c e s s , and c a n be t i m e - c o n s u m i n g , especially when l a r g e numbers o f samples a r e i n v o l v e d . In o r d e r t o improve spectrometer e f f i c i e n c y , t h e p h o s p h a t e a s s a y s a r e now b e i n g p e r f o r m e d on a VXR300S e q u i p p e d w i t h a r o b o t i c s a m p l e changer. Utilizing easy-to-use menu-driven software, laboratory p e r s o n n e l d e s i g n a t e t h e p r e f e r r e d s o l v e n t , d u r a t i o n and t y p e o f shimming ( m a g n e t i c f i e l d o p t i m i z a t i o n ) , a u t o m a t i o n o p e r a t i n g p a r a m e t e r s e t ( e n v i r o n m e n t ) , and p l o t t e r c h o i c e . Initiation of the automation r u n i s s i m p l y begun by c l i c k i n g a s i n g l e menu b u t t o n . The c u s t o m i z e d s o f t w a r e then a u t o m a t i c a l l y accumulates the data, s t o r e s i t i n a common a r e a f o r r o b o t i c r u n s , a n d p e r f o r m s t h e F o u r i e r t r a n s f o r m a n d i n t e g r a t i o n , and c a l c u l a t e s t h e r e l a t i v e d i s t r i b u t i o n o f phosphate s p e c i e s . S p e c t r a comparable t o that i n Figure 1 are generated without f u r t h e r operator i n t e r v e n t i o n . Improvements i n t h e s e n s i t i v i t y a r e o b s e r v e d a t t h e h i g h e r f i e l d (121.4 MHz) o f t h e VXR300S, r e s u l t i n g i n shorter o v e r a l l accumulations. The d a t a r e d u c t i o n s o f t w a r e F o u r i e r t r a n s f o r m s and phases t h e d a t a a f t e r z e r o - f i l l i n g and a p p l i c a t i o n o f t h e d e s i g n a t e d e x p o n e n t i a l f i l t e r f u n c t i o n . The u s u a l c h e m i c a l s h i f t range o f t h e peaks i s noted, as t h e system s e a r c h e s for the primary phosphorus-containing species. A signalt o - n o i s e check i s p e r f o r m e d o v e r t h a t r e g i o n , and peaks g e n e r a t i n g a v a l u e l e s s than t h r e e a r e d e s i g n a t e d as not b e i n g p r e s e n t ( o r a p p r o x i m a t e l y 0.2 w e i g h t p e r c e n t , t h e l o w e r c o n c e n t r a t i o n l i m i t f o r t h e r o u t i n e P a s s a y ) . Once the peaks are found, the spectrum i s automatically integrated. The i n t e g r a l v a l u e s a n d m o l e c u l a r w e i g h t o f each of the s p e c i e s are then used i n the computation of the r e l a t i v e weight percent d i s t r i b u t i o n of the phosphate species. The c o m p u t e r - d e r i v e d d a t a r e d u c t i o n c o m p a r e s q u i t e f a v o r a b l y t o t h a t p e r f o r m e d by human a n a l y s t s a s s e e n in Table I I I . The two methods commonly d i f f e r b y a r e l a t i v e e r r o r o f l e s s t h a n 0.2 w e i g h t p e r c e n t f o r t h e major s p e c i e s . The mean f o r e a c h s p e c i e s i s shown t o i l l u s t r a t e t h e l a c k o f b i a s b e t w e e n t h e two t e c h n i q u e s . The m a i n a d v a n t a g e s o f t h e c o m p u t e r method a r e i t s s p e e d and lack of subjectivity. The automated quantitative analyses are normally performed a t n i g h t , a l l o w i n g access t o t h e s p e c t r o m e t e r d u r i n g t h e d a y , when demand f o r t h e instrument i s high. 31
In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
3. GARD ET A L
49
Quantitative Analysis of Inorganic Phosphates
Table I I I M a n u a l v s . R o b o t i c (ASM) D a t a R e d u c t i o n C o m p a r i s o n o f STP A s s a y s R e l a t i v e Wt % a s Sodium
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Sample No,
Orthophosphate manual ASM
A Β C D Ε F Mean
Pyrophosphate manual ASM
Salts
TripolyptlQSPhate manual ASM
TrimetaphQSPhate manual ASM
0.35 0.17 0.24 0.11 0.14 0.23
0.39 0.15 0.21 0.08 0.12 0.18
8.63 6.62 4.29 6.82 6.31 5.41
8.62 6.61 4.35 6.72 6.16 5.36
89.39 90.99 93.56 91.05 91.60 92.76
89.48 91.18 93.68 91.31 91.84 93.01
1.63 2.22 1.91 2.03 1.95 1.60
1.51 2.05 1.76 1.90 1.87 1.45
0.21
0.19
6.35
6.30
91.56
91.75
1.89
1.76
Hcmonuclear Two-Dimensional
J-Resolved Spectroscopy
Due t o t h e c o m p l e x i t y o f t h e o n e - d i m e n s i o n a l s p e c t r a o f o l i g o p h o s p h a t e g l a s s e s , s p e c i e s d e t e r m i n a t i o n by c u r v e f i t t i n g has been s u c c e s s f u l l y a p p l i e d o n l y t o r e l a t i v e l y pure samples o f i n d i v i d u a l o l i g o p h o s p h a t e s p e c i e s ( 1 9 ) . In an e f f o r t t o r e s o l v e t h e o v e r l a p p i n g m u l t i p l e t s i n c o m p l e x s p e c t r a , t w o - d i m e n s i o n a l NMR i s b e i n g e x a m i n e d . A o n e - d i m e n s i o n a l P spectrum o f a sodium phosphate g l a s s w i t h a n a v e r a g e c h a i n l e n g t h _ o f 4.11 i s shown i n F i g u r e 3. (The a v e r a g e c h a i n l e n g t h , n, i s d e t e r m i n e d b y e l e m e n t a l analysis.) A s c a n be o b s e r v e d , a l a r g e number o f c l o s e l y s p a c e d r e s o n a n c e s a r e o b t a i n e d . An e x p a n s i o n o f t h e " e n d s " r e g i o n i s a l s o s e e n i n t h e i n s e t o f F i g u r e 3. Although some o f t h e p e a k s a r e s e p a r a t e d a t h i g h f i e l d , a s s i g n m e n t s , and t h e r e f o r e q u a n t i t a t i o n , o f e a c h o f t h e v a r y i n g c h a i n l e n g t h r e s o n a n c e s , i s c o m p l i c a t e d by t h e p r e s e n c e o f PP c o u p l i n g . Removing t h i s c o u p l i n g w o u l d g r e a t l y s i m p l i f y the spectrum. S i m p l e h o m o n u c l e a r d e c o u p l i n g w o u l d be i n s u f f i c i e n t , as t h e a b i l i t y t o simultaneously observe a l l p h o s p h o r u s - c o n t a i n i n g s p e c i e s would t h e n be l o s t . 31
31
3l
31
P h o m o n u c l e a r 2DJ e x p e r i m e n t s were p e r f o r m e d o n t h e g l a s s t o e f f e c t i v e l y d e c o u p l e t h e s p e c t r u m . A 2DJ s p e c t r u m o f t h e p h o s p h a t e e n d s r e g i o n i s shown i n F i g u r e 4. The e n d s r e g i o n was u s e d f o r t h e p h o s p h a t e s p e c i e s d e t e r m i n a t i o n because o f i t s r e l a t i v e s i m p l i c i t y and f i r s t o r d e r nature i n comparison w i t h t h e middles r e g i o n . The p h o s p h o r u s c h e m i c a l s h i f t s a r e s e e n on one a x i s , a n d t h e P- P c o u p l i n g c o n s t a n t s on t h e o t h e r . P a i r s ( o r more) o f c o n t o u r s a t one c h e m i c a l s h i f t a r e t h e p e a k s f o r one ( c h a i n l e n g t h ) s p e c i e s , s e p a r a t e d by t h e c o u p l i n g c o n s t a n t . A v e r t i c a l p r o j e c t i o n o f t h e spectrum onto t h e chemical s h i f t a x i s c l e a r l y d e l i n e a t e s the newly-separated resonances, 31
31
In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
"ends"
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-E -4 -6 -8 -10 -12 -14 -16 -18 F i g u r e 3. B a s e s p e c t r u m - 161.9 MHz P NMR s p e c t r u m o f s o d i u m p h o s p h a t e g l a s s w i t h n= 4.1. Inset- Expansion o f t h e "ends" r e g i o n . Each s p e c i e s i s d e s i g n a t e d by t h e number o f p h o s p h o r u s atoms i n t h e m o l e c u l e .
-88
I I 1 1 I 1 I I I I I I I 1 I I I I I I I I I I I I I I I I I I I I I I I I I I I I I I I I I I I I I Γ I I I I I I I I I I 1 I I I I' I I I
-22
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ortho
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3. GARD ET A L
Quantitative Analysis of Inorganic Phosphates
51
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r e s u l t i n g i n a phosphorus-phosphorus decoupled spectrum as shown a t t h e t o p o f F i g u r e 4. A monotonie i n c r e a s e i n chemical s h i f t i s seen with i n c r e a s i n g phosphorus c h a i n length species(20). I n an e f f o r t t o q u a n t i f y t h e amounts o f t h e v a r i o u s c h a i n l e n g t h s , t h i s p r o j e c t i o n was s u b j e c t e d t o s p e c t r a l curve-fitting. I n F i g u r e 5 i s seen t h e r e s u l t s o f t h i s a n a l y s i s , w i t h t h e e x p e r i m e n t a l spectrum a t t h e bottom, and t h e c o m p u t e r - g e n e r a t e d f i t above i t . The d e t e r m i n a t i o n o f t h e phosphate s p e c i e s by i n t e g r a t i o n o f t h e s i g n a l a r e a s o f t h e f i t t e d spectrum i s l i s t e d i n T a b l e I V . The average c h a i n l e n g t h d e t e r m i n e d b y 2DJ NMR i s 4.13, w e l l w i t h i n experimental error o f t h e 4.11 v a l u e determined by elemental a n a l y s i s . A c o m p a r i s o n i s a l s o made i n T a b l e I V o f t h e 2DJ r e s u l t s w i t h t h a t p r e v i o u s l y f o u n d f o r a s o d i u m p h o s p h a t e g l a s s d e t e r m i n e d b y p a p e r c h r o m a t o g r a p h y t o be n= 4.0 ( 2 1 ) . T h e r e s u l t s a r e c u r r e n t l y c o n s i d e r e d t o be s e m i - q u a n t i t a t i v e , a s t h e 2DJ e x p e r i m e n t c a n g i v e r i s e t o a r t i f a c t s ; t r u e i n t e n s i t i e s d o n o t a l w a y s r e s u l t when p r o j e c t i o n s o n t o t h e c h e m i c a l s h i f t a x i s a r e made b e c a u s e of poor l i n e s h a p e s . More a c c u r a t e q u a n t i f i c a t i o n c o u l d p o s s i b l y be a c h i e v e d by u s i n g volume i n t e g r a l s o r by u t i l i z i n g t h e methods o f Xu e t a l ( 2 2 ) t o h e l p e l i m i n a t e these shortcomings.
Composition
T a b l e IV o f Sodium P h o s p h a t e
% Total
Length,
* Ref.
5
Ρ NMR
1.2 9.4 13.8 24.9 17.5 19.7 7.8 2.8 2.9
Glass = 4.0*
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0 6.6 28.2 27.4 16.9 9.4 5.7 2.7 1.8 1.2
η =
Elemental A n a l y s i s 2DJ P NMR 31
2
31·
Ortho Pyro Tripoly Tetrapoly Pentapoly Hexapoly Heptapoly Octapoly Nonapoly Higher Average Chain
P 0 as2DJ
Phosphate Species
Glass
4.11 4.13
21
In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
PHOSPHORUS CHEMISTRY
52
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F i g u r e 4. p- p homonuclear 2DJ-resolved NMR spectrum of the same g l a s s whose one-dimensional spectrum i s shown i n F i g u r e 3. The c h a r a c t e r i s t i c J c o u p l i n g p a t t e r n i s observed f o r each s p e c i e s . A projection onto the chemical s h i f t a x i s i s shown, above. The p r o j e c t i o n f o r each s p e c i e s i s designated by the number of phosphorus atoms i n the molecule. The l i n e s h a p e of the octaphosphate was not d i s t i n c t enough t o allow f o r independent f i t t i n g .
In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
GARD ET AL
53
Quantitative Analysis of Inorganic Phosphates
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PHOSPHORUS CHEMISTRY
54 Conclusions
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P h o s p h o r u s - 3 1 NMR p r o v i d e s an a c c u r a t e and p r e c i s e method f o r t h e a n a l y s i s o f s o l u b l e p h o s p h a t e s and t h e i r m i x t u r e s . The a c c u r a c y and p r e c i s i o n a r e c o m p a r a b l e t o accepted c h r o m a t o g r a p h i c m e t h o d s . The r e s u l t s s i g n i f y t h a t t h e NMR method might also be successfully applied to less s t r a i g h t f o r w a r d s y s t e m s w i t h h i g h a c c u r a c y and p r e c i s i o n and w i t h t h e a d d i t i o n a l a d v a n t a g e s c h a r a c t e r i s t i c o f NMR w h i c h were e n u m e r a t e d e a r l i e r i n t h e a r t i c l e . The most s i g n i f i c a n t s o u r c e o f e r r o r i n t h e NMR method i s i n i n t e g r a t i o n of the s i g n a l areas, e s p e c i a l l y f o r resonances w i t h c h e m i c a l s h i f t s l y i n g c l o s e t o one a n o t h e r . This s o u r c e o f e r r o r i s a l l e v i a t e d i n l a r g e measure by e m p l o y i n g c u r v e d e c o n v o l u t i o n and l i n e s h a p e a n a l y s i s . Application of quantitative robotic analysis g r e a t l y improves the e f f i c i e n c y , w h i l e r e l i a b l y p r o d u c i n g r e s u l t s e q u i v a l e n t t o t h o s e o b t a i n e d b y manual s e l e c t i o n o f t h e i n t e g r a l r e g i o n s . L i t t l e more t h a n c l i c k i n g a s i n g l e menu b u t t o n i s r e q u i r e d t o i n i t i a t e an a u t o m a t i o n run. By removing much of the analyst subjectivity, more r e p r o d u c i b l e and r e l i a b l e a s s a y s a r e o b t a i n e d . By e x t e n d i n g t h e a p p l i c a t i o n o f t h e h o m o n u c l e a r 2DJ experiment t o the realm of oligophosphate assays, twod i m e n s i o n a l NMR i s s e e n t o be a p r o m i s i n g e x t e n s i o n f o r unravelling complicated oligophosphate s p e c t r a and in quantitative analysis. Coupled w i t h the use o f l i n e s h a p e a n a l y s i s and c u r v e f i t t i n g , r e a s o n a b l e s e m i q u a n t i t a t i v e results are obtained that compare well with other established a n a l y t i c a l techniques. Acknowledgments NMR l i n e shape a n a l y s i s s o f t w a r e d e v e l o p e d f o r use at M o n s a n t o was k i n d l y s u p p l i e d by Mr. N.G. H o f f m a n , R e s e a r c h C o m p u t i n g C o n s o r t i u m , M o n s a n t o Co., and P r o f . A . J . Duben, Southeast M i s s o u r i State U n i v e r s i t y . The a u t h o r s w o u l d also like t o t h a n k Mr. Brad Herman o f t h e Research C o m p u t i n g C o n s o r t i u m , M o n s a n t o Co., f o r h i s d e v e l o p m e n t o f t h e a u t o m a t i c s a m p l e management s o f t w a r e , Dr. W i l l i a m W i s e o f t h e P h y s i c a l S c i e n c e s C e n t e r ( P S C ) , M o n s a n t o Co., f o r his many valuable discussions on homonuclear 2DJ s p e c t r o s c o p y , and Mr. J o h n B u r q u i n ( M o n s a n t o PSC) f o r h i s assistance i n the development o f the robotic assay. A p p r e c i a t i o n i s a l s o e x p r e s s e d t o t h e many a n a l y s t s who p a r t i c i p a t e d i n the study.
In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
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3. GARD ET AL
55 Quantitative Analysis of Inorganic Phosphates
Literature Cited 1. Colson, J.G.; Marr, D.H. Anal. Chem. 1973, 45, 370. 2. Gurley, T.W.; Ritchey, W.H. Anal. Chem., 1975, 47, 1444. 3. Sojka, S.A.; Wolfe, R.A. Anal. Chem., 1978, 50, 585. 4. Stanislawski, D.A.; Van Wazer, J.R. Anal. Chem., 1980, 52, 96. 5. Ruben, I.B. Anal. Lett., 1984, 17, 1259. 6. Glonek, T.; Wang, P.J.; Van Wazer, J.R. J . Amer. Chem. Soc., 1976, 98, 7968. 7. Hanssum, H. J . Magn. Reson., 1981, 45, 461. 8. Becker, E.D.; Ferretti, J.A.; Gambhir, P.N. Anal. Chem., 1979, 51, 1413. 9. Press, W.H.; Flannery, Β.Ρ.; Teukolski, S.A.; Vetterling, W.T. Numerical Recipes: The Art of Scientific Computing, Cambridge Univ. Press, Cambridge, MA, 1986; Chapter 14. 10. ASTM Method D 501, Standard Test Methods of Sampling and Chemical Analysis of Alkaline Detergents, Annual Book of ASTM Standards, American Society for Testing and Materials, Philadelphia, 1989. 11. ASTM Method D 2761, Standard Test Method for Analysis of Sodium Triphosphate by the Simplified Ion Exchange Method, Annual Book of ASTM Standards, American Society for Testing and Materials, Philadelphia, 1989. 12. Lundgren, D.P.; Loeb, N.P. Anal. Chem., 1961, 33, 366. 13. Condensed Phosphates by Column Separation, Industrial Method No.317-74I. Bran and Luebbe, Technicon Industrial Systems, Elmsford, NY. 14. Longley-Cook, L.H. Statistical Problems and How to Solve Them, Barnes & Noble, New York, N.Y., 1970; Chapter 14. 15. Sotak, C.H.; Dumoulin, C.L.; Levy, G.C. Anal. Chem., 1983, 55, 782. 16. Weiss, G.H.; Ferretti, J.A. J . Magn. Reson., 1983, 55, 397. 17. Watanabe, M.; Matsuura, M.; Yamada, T. Bull. Chem. Soc. Japan, 1981, 54, 738. 18. Crowther, J.P.; Westman, A.E.R. Can. J . Chem., 1954, 32, 42. 19. MacDonald, J.C.; Mazurek, M. J . Magn. Reson., 1987, 72, 48. 20. Glonek, T.; Costello, A.J.R.; Myers, T.C.; Van Wazer, J.R. J . Phys. Chem. 1975, 79, 1214. 21. Van Wazer, J.R. Phosphorus and Its Compounds, Interscience Publishers, New York, 1958, Vol. 1, Chap. 12. 22. Xu, P.; Wu, X-L.; Freeman, R. J . Magn. Reson., 1991, 95, 132. RECEIVED December 10, 1991
In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
