JOURNAL
OF APPLIED PHYSIOLOGY 1976. Printed
Vol. 40, No. 1, January
in U.S.A.
Cerebrospinal fluid sampling technique and Astrup pH and Pcoz values DONALD G. DAVIES Department of Physiology,
Texas Tech University
SchooZ of Medicine,
DAVIES, DONALD G. Cerebrospinal fluid sampling technique and Astrup pH and PCO~values. J. Appl. Physiol. 40(l): 123125. 1976. -The pH and Pco2 values measured by the Astrup technique were compared in cerebrospinal fluid (CSF) obtained using two different sampling techniques: 1) a direct or in vivo technique and.2) the widely accepted syringe sampling technique. In 65 pairs of measurements in 9 dogs it was found that the pH was always overestimated and the Pco2 always underestimated in the syringe sample when compared to the in vivo sample. The equations describing the relationships are as follows: 1) pH (syringe = 0.995 pH (in vivo) + 0.084 and2) Pco2 (syringe) = 0.873 PcoZ (in vivo) + 0.2. The amount by which the syringe sample underestimated the true Pco2 value increased with the absolute Pco2 value, consistent with the possibility of there being a diffusional loss of CO, during the transfer of CSF from the syringe to the pH electrode (Pco, (in vivo) - Pco2 (syringe) = 2.4, 4.9, 7.5, and 10.0 mmHg at in vivo Pco& of 20, 40, 60, and 80 mmHg). This study indicates that the technique used for sampling CSF is crucial to the expected accuracy of the results and that the number of transfers of CSF during the sampling and measurement procedures should be minimized in order to obtain reliable results.
acid-base balance; measurement
Astrup
technique;
pH measurement;
Lubbock,
Texas 79409
When CSF was sampled by the conventional technique, two glass syringes (5 ml) were connected to a metal stopcock, one positioned vertically and one horizontally. The stopcock was then connected in series to a second stopcock which was located in the cisternal needle. Between 0.5 and 0.6 ml of CSF was slowly drawn into the horizontal syringe. Both syringes and the stopcock were removed from the second stopcock and the CSF and the air bubbles in the dead space were flushed into the vertical syringe. This procedure completely filled the dead space of the sample syringe with CSF. After this was repeated three additional times, the syringe was removed and immediately capped. The pH was measured at 37°C within 30 s with a Radiometer-Astrup glass microelectrode. The electrode was calibrated both before and after each pH determination with Radiometer precision buffer mixtures of KH,PO, and Na,HPO,, 2H,O with pH values of 7.383 and 6.841, respectively. All values were temperature corrected by using the factor of Mitchell et al. (4) of 0.0037 units/C. Measurements were repeated until two consecutive readings agreed within 0.005 pH units. The second sampling technique will be referred to as the direct or in vivo technique. In this case, the CSF was sampled directly from the animal by placing the polyethylene capillary of the glass microelectrode into the end of the indwelling Riley needle. This type of needle is unique because the shaft extends out beyond the hub and allows for an airtight seal to be made between the needle and the polyethylene capillary of the Astrup microelectrode. The CSF was allowed to flow into the glass electrode under its own hydrostatic pressure. The pH was measured within 2 s after sampling. This procedure was repeated without rinsing the electrode with distilled water or air until two consecutive readings agreed within 0.005 pH units. CSF PcoZ was estimated by the Astrup equilibration technique (5-7). The Pcop was calculated for both the syringe sample and the in vivo sample using the pH values obtained with each method. The accuracy of this technique for measuring CSF Pco2 was verified previously (1, 3, 8). During the measurement, 0.2 ml of CSF was equilibrated for 3 min at 37°C with Scholander analyzed gases containing 4.08% COZ-balance O2 and 9.53% COz-balance 0,. The pH of each sample was measured repeatedly until two consecutive readings agreed within 0.005 units.
PCO~
THE MOST WIDELY ACCEPTED TECHNIQUE for sampling cerebrospinal fluid (CSF) involves the use of a glass or plastic syringe. It is well known that CSF is essentially . unbuffered and the strong possibility exists that pH could be overestimated due to the loss of molecular CO, during the transfer of CSF from the syringe to the pH electrode. This overestimation of CSF pH results in an underestimation of CSF PCO~ when measured by the Astrup equilibration technique since the calculated Pco2 value is dependent on the measured pH. In this study, a sampling technique was developed whereby CSF was sampled directly from an indwelling needle in the cisterna magna. This method eliminates the requirement for a syringe. The acidbase values obtained with this technique were compared to those obtained with the syringe technique. In all cases, lower pH values and higher Pco2 values were obtained with the direct sampling technique.
RESULTS METHODS
Figure 1 shows the syringe pH values plotted against the in vivo pH values. The equation for the line of best fit through the points is: pH (syringe) = 0.995 pH (in vivo) + 0.084. It is evident that the pH is consistently overestimated when the CSF sample is obtained by the syringe technique. Table 1 shows syringe pH values calculated for different in vivo pH values from the equation for the line of best fit. Table 1 indicates that the CSF pH is overestimated by approximately 0.05 units by the syringe technique.
CSF was sampled in nine mongrel dogs, anesthetized with sodium pentobarbital (25 mg/kg). They were paralyzed with succinylcholine and mechanically ventilated. Catheters were placed in the femoral artery for blood pressure measurement and in the femoral vein for administration of the succinylcholine and pentobarbital. A 19-gauge Riley arterial puncture needle was inserted in the cisterna magna and secured with a stereotaxic apparatus. 123
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124
D. G. DAVIES Pco,
PH (syringe)
(syringe)
I 7.7
80
7.6
entity
60 50 40 30 20
I
71
I
72
L
1
73
74 pH
1
75
I
1
7.6
7.7
FIG. 1. In vivo pH values plotted against syringe pH values. Equation for line of best fit through the points as follows: pH (syringe) = 0.995 pH (in viva) + 0.084. Line of identity is shown.
TABLE
1. In vivo and syringe pH values In Vivo
Syringe
7.000 7.200 7.400 7.600 7.800
7.049 7.248 7.447 7.646 7.845
IO
(In VIVO)
AtSyringe
It is well known that it is extremely difficult to accurately estimate CSF pH and Pco~. This is primarily the result of the low-protein concentration and therefore, the low buffering capacity of CSF (2). Even a slight loss of molecular CO, during the sampling or measurement procedures will result in drastic increases in the pH value. The higher pH values obtained from the syringe samples in the present study are most likely the results of such a loss of CO, and not faulty pH measurement since consecutive readings could be obtained which agreed within 0.005 pH units. In addition the observations that the in vivo Pcoz was higher and Pco2 difference between the syringe sample and the in vivo sample increased with the absolute Pco2 are both consistent with their being a loss of CO, during the transfer of the CSF to the pH electrode. When the direct sampling technique was used in this study, the CSF was never
50
60
70
80
90
100
(In VIVO)
2. In vivo and syringe PCOB values (mmHg) In Vivo
0.049 0.048 0.047 0.046 0.045
DISCUSSION
40
FIG. 2. In vivo Pcoz values plotted against syringe Pco,, values. Equation for line of best fit through the points is as follows: PCO~ (syringe) = 0.873 Pco2 (in vivo) + 0.2. Line of identity ” is shown.
TABLE
The data for the Astrup Pcoz measurements are presented in Fig. 2 where the syringe Pcoz values are plotted against the in vivo Pco2 values. The equation for the line of best fit is: Pco2 (syringe) = 0.873 Pco2 (in vivo) + 0.2. Table 2 shows the syringe PCO~ values calculated for various in vivo Pcoz values from the equation for the line of best fit. Figure 2 and Table 2 both indicate that, although there is fairly good agreement between the two different Pco2 values at low Pco&, the degree by which the Pcoz is underestimated in the syringe sample increases markedly as the absolute Pco2 increases.
30
Pco,
- In Vivo)
Syringe pH values for different in vivo values calculated from the following equation: pH (syringe) = 0,995 pH (in vivo) + 0.084. Differences between the syringe and in vivo pH values are also shown.
20
Syringe
20 30 40 50 60 70 80 Syringe Pco2 values the following equation:
17.6 26.4 35.1 43.9 52.6 61.3 70.0
A(In Vivo
- Syringe)
2.3 3.6 4.9 6.2 7.4 8.7 10.0
for different PcoB (syringe)
Differences between the in vivo shown.
in vivo values calculated from = 0.873 Pcoz (in vivo) + 0.2. and syringe PCO, values are also
exposed to room air. This procedure prevented the loss of molecular CO, and probably accounts for the lower pH values and higher Pcoz values obtained with the in vivo method. Therefore, the data indicate that the in vivo technique provides a better estimate of CSF Pco~. CaZculation of CSF Pco2 from pH anti: total CO, values. The chance that the syringe pH could be overestimated is also important with respect to an alternative method for estimating CSF Pco2 where it is calculated from the measured pH and total CO2 values. In this case, the calculated Pcoz would be less than the true Pcoz if the syringe pH were higher than the true pH. Correction factors. The existence of the high correlation coefficients between the syringe and the in vivo values should allow one to apply correction factors to previously published measurements in order to estimate the actual in vivo Pco&. These correction factors could be derived from the equations for the lines of best fit for the pH and Pco2 values obtained in this study. The author thanks Mr. Kenneth Corder for his excellent technical assistance during these studies. This study was supported by a grant from the American Heart Association, Texas Affiliate, Inc. Received
for publication
20 June
1975.
Downloaded from www.physiology.org/journal/jappl by ${individualUser.givenNames} ${individualUser.surname} (192.236.036.029) on August 12, 2018. Copyright © 1976 American Physiological Society. All rights reserved.
CSF
pH
AND
Pco2
VALUES
125
REFERENCES 1. DAVIES, D. G., R. S. FITZGERALD, AND G. H. GURTNER. Acid-base relationships between CSF and blood during acute metabolic acidosis. J. Appl. Physiol. 34: 243-248, 1974. 2. DAVSON, H. Physiology of the Cerebrospinul FZuid. Boston, Mass. : Little, Brown, 1967, p. 272-279. 3. LEE, J. E., R. CHU, J. B. POSNER, AND F. PLUM. Buffering capacity of cerebrospinal fluid in experimental acidosis. Am. J. Physiol. 217: 1035-1038, 1969. 4. MITCHELL, R. A., D. A. HERBERT, AND C. T. CARMEN. Acid-base constants and temperature coeffkients for cerebrospinal fluid. J. Appl. Physiol. 20: 27-30, 1965.
5. SIGGAARD-ANDERSEN, 0. The pH-log Pcoz blood acid-base nomogram. Stand. J. Clin. Lab Invest. 14: 598-604, 1962. 6. SIGGAARD-ANDERSEN, O., AND K. ENGLE. A new acid-base nomogram. &and. J. Clin. Lab Invest. 12: 177-186, 1960. 7. SIGGAARD-ANDERSEN, O., K. ENGLE, K. JORGENSEN, AND P. AsTRUP. A micromethod for determination of pH, carbon dioxide, base excess and standard bicarbonate in capillary blood. Stand. J. Clin. Lab Invest. 12: 172-176, 1960. 8. VAN HEIJST, A. N. P., B. F. VISSER, AND A. H. J. MASS. A micromethod for the determination of pH and Pco2 in human cerebrospinal fluid. Clin. Chim. Actu 6: 589490, 1961.
Downloaded from www.physiology.org/journal/jappl by ${individualUser.givenNames} ${individualUser.surname} (192.236.036.029) on August 12, 2018. Copyright © 1976 American Physiological Society. All rights reserved.
OF APPLIED PHYSIOLOGY 1976. Printed
Vol. 40, No. 1, January
in U.S.A.
Cerebrospinal fluid sampling technique and Astrup pH and Pcoz values DONALD G. DAVIES Department of Physiology,
Texas Tech University
SchooZ of Medicine,
DAVIES, DONALD G. Cerebrospinal fluid sampling technique and Astrup pH and PCO~values. J. Appl. Physiol. 40(l): 123125. 1976. -The pH and Pco2 values measured by the Astrup technique were compared in cerebrospinal fluid (CSF) obtained using two different sampling techniques: 1) a direct or in vivo technique and.2) the widely accepted syringe sampling technique. In 65 pairs of measurements in 9 dogs it was found that the pH was always overestimated and the Pco2 always underestimated in the syringe sample when compared to the in vivo sample. The equations describing the relationships are as follows: 1) pH (syringe = 0.995 pH (in vivo) + 0.084 and2) Pco2 (syringe) = 0.873 PcoZ (in vivo) + 0.2. The amount by which the syringe sample underestimated the true Pco2 value increased with the absolute Pco2 value, consistent with the possibility of there being a diffusional loss of CO, during the transfer of CSF from the syringe to the pH electrode (Pco, (in vivo) - Pco2 (syringe) = 2.4, 4.9, 7.5, and 10.0 mmHg at in vivo Pco& of 20, 40, 60, and 80 mmHg). This study indicates that the technique used for sampling CSF is crucial to the expected accuracy of the results and that the number of transfers of CSF during the sampling and measurement procedures should be minimized in order to obtain reliable results.
acid-base balance; measurement
Astrup
technique;
pH measurement;
Lubbock,
Texas 79409
When CSF was sampled by the conventional technique, two glass syringes (5 ml) were connected to a metal stopcock, one positioned vertically and one horizontally. The stopcock was then connected in series to a second stopcock which was located in the cisternal needle. Between 0.5 and 0.6 ml of CSF was slowly drawn into the horizontal syringe. Both syringes and the stopcock were removed from the second stopcock and the CSF and the air bubbles in the dead space were flushed into the vertical syringe. This procedure completely filled the dead space of the sample syringe with CSF. After this was repeated three additional times, the syringe was removed and immediately capped. The pH was measured at 37°C within 30 s with a Radiometer-Astrup glass microelectrode. The electrode was calibrated both before and after each pH determination with Radiometer precision buffer mixtures of KH,PO, and Na,HPO,, 2H,O with pH values of 7.383 and 6.841, respectively. All values were temperature corrected by using the factor of Mitchell et al. (4) of 0.0037 units/C. Measurements were repeated until two consecutive readings agreed within 0.005 pH units. The second sampling technique will be referred to as the direct or in vivo technique. In this case, the CSF was sampled directly from the animal by placing the polyethylene capillary of the glass microelectrode into the end of the indwelling Riley needle. This type of needle is unique because the shaft extends out beyond the hub and allows for an airtight seal to be made between the needle and the polyethylene capillary of the Astrup microelectrode. The CSF was allowed to flow into the glass electrode under its own hydrostatic pressure. The pH was measured within 2 s after sampling. This procedure was repeated without rinsing the electrode with distilled water or air until two consecutive readings agreed within 0.005 pH units. CSF PcoZ was estimated by the Astrup equilibration technique (5-7). The Pcop was calculated for both the syringe sample and the in vivo sample using the pH values obtained with each method. The accuracy of this technique for measuring CSF Pco2 was verified previously (1, 3, 8). During the measurement, 0.2 ml of CSF was equilibrated for 3 min at 37°C with Scholander analyzed gases containing 4.08% COZ-balance O2 and 9.53% COz-balance 0,. The pH of each sample was measured repeatedly until two consecutive readings agreed within 0.005 units.
PCO~
THE MOST WIDELY ACCEPTED TECHNIQUE for sampling cerebrospinal fluid (CSF) involves the use of a glass or plastic syringe. It is well known that CSF is essentially . unbuffered and the strong possibility exists that pH could be overestimated due to the loss of molecular CO, during the transfer of CSF from the syringe to the pH electrode. This overestimation of CSF pH results in an underestimation of CSF PCO~ when measured by the Astrup equilibration technique since the calculated Pco2 value is dependent on the measured pH. In this study, a sampling technique was developed whereby CSF was sampled directly from an indwelling needle in the cisterna magna. This method eliminates the requirement for a syringe. The acidbase values obtained with this technique were compared to those obtained with the syringe technique. In all cases, lower pH values and higher Pco2 values were obtained with the direct sampling technique.
RESULTS METHODS
Figure 1 shows the syringe pH values plotted against the in vivo pH values. The equation for the line of best fit through the points is: pH (syringe) = 0.995 pH (in vivo) + 0.084. It is evident that the pH is consistently overestimated when the CSF sample is obtained by the syringe technique. Table 1 shows syringe pH values calculated for different in vivo pH values from the equation for the line of best fit. Table 1 indicates that the CSF pH is overestimated by approximately 0.05 units by the syringe technique.
CSF was sampled in nine mongrel dogs, anesthetized with sodium pentobarbital (25 mg/kg). They were paralyzed with succinylcholine and mechanically ventilated. Catheters were placed in the femoral artery for blood pressure measurement and in the femoral vein for administration of the succinylcholine and pentobarbital. A 19-gauge Riley arterial puncture needle was inserted in the cisterna magna and secured with a stereotaxic apparatus. 123
Downloaded from www.physiology.org/journal/jappl by ${individualUser.givenNames} ${individualUser.surname} (192.236.036.029) on August 12, 2018. Copyright © 1976 American Physiological Society. All rights reserved.
124
D. G. DAVIES Pco,
PH (syringe)
(syringe)
I 7.7
80
7.6
entity
60 50 40 30 20
I
71
I
72
L
1
73
74 pH
1
75
I
1
7.6
7.7
FIG. 1. In vivo pH values plotted against syringe pH values. Equation for line of best fit through the points as follows: pH (syringe) = 0.995 pH (in viva) + 0.084. Line of identity is shown.
TABLE
1. In vivo and syringe pH values In Vivo
Syringe
7.000 7.200 7.400 7.600 7.800
7.049 7.248 7.447 7.646 7.845
IO
(In VIVO)
AtSyringe
It is well known that it is extremely difficult to accurately estimate CSF pH and Pco~. This is primarily the result of the low-protein concentration and therefore, the low buffering capacity of CSF (2). Even a slight loss of molecular CO, during the sampling or measurement procedures will result in drastic increases in the pH value. The higher pH values obtained from the syringe samples in the present study are most likely the results of such a loss of CO, and not faulty pH measurement since consecutive readings could be obtained which agreed within 0.005 pH units. In addition the observations that the in vivo Pcoz was higher and Pco2 difference between the syringe sample and the in vivo sample increased with the absolute Pco2 are both consistent with their being a loss of CO, during the transfer of the CSF to the pH electrode. When the direct sampling technique was used in this study, the CSF was never
50
60
70
80
90
100
(In VIVO)
2. In vivo and syringe PCOB values (mmHg) In Vivo
0.049 0.048 0.047 0.046 0.045
DISCUSSION
40
FIG. 2. In vivo Pcoz values plotted against syringe Pco,, values. Equation for line of best fit through the points is as follows: PCO~ (syringe) = 0.873 Pco2 (in vivo) + 0.2. Line of identity ” is shown.
TABLE
The data for the Astrup Pcoz measurements are presented in Fig. 2 where the syringe Pcoz values are plotted against the in vivo Pco2 values. The equation for the line of best fit is: Pco2 (syringe) = 0.873 Pco2 (in vivo) + 0.2. Table 2 shows the syringe PCO~ values calculated for various in vivo Pcoz values from the equation for the line of best fit. Figure 2 and Table 2 both indicate that, although there is fairly good agreement between the two different Pco2 values at low Pco&, the degree by which the Pcoz is underestimated in the syringe sample increases markedly as the absolute Pco2 increases.
30
Pco,
- In Vivo)
Syringe pH values for different in vivo values calculated from the following equation: pH (syringe) = 0,995 pH (in vivo) + 0.084. Differences between the syringe and in vivo pH values are also shown.
20
Syringe
20 30 40 50 60 70 80 Syringe Pco2 values the following equation:
17.6 26.4 35.1 43.9 52.6 61.3 70.0
A(In Vivo
- Syringe)
2.3 3.6 4.9 6.2 7.4 8.7 10.0
for different PcoB (syringe)
Differences between the in vivo shown.
in vivo values calculated from = 0.873 Pcoz (in vivo) + 0.2. and syringe PCO, values are also
exposed to room air. This procedure prevented the loss of molecular CO, and probably accounts for the lower pH values and higher Pcoz values obtained with the in vivo method. Therefore, the data indicate that the in vivo technique provides a better estimate of CSF Pco~. CaZculation of CSF Pco2 from pH anti: total CO, values. The chance that the syringe pH could be overestimated is also important with respect to an alternative method for estimating CSF Pco2 where it is calculated from the measured pH and total CO2 values. In this case, the calculated Pcoz would be less than the true Pcoz if the syringe pH were higher than the true pH. Correction factors. The existence of the high correlation coefficients between the syringe and the in vivo values should allow one to apply correction factors to previously published measurements in order to estimate the actual in vivo Pco&. These correction factors could be derived from the equations for the lines of best fit for the pH and Pco2 values obtained in this study. The author thanks Mr. Kenneth Corder for his excellent technical assistance during these studies. This study was supported by a grant from the American Heart Association, Texas Affiliate, Inc. Received
for publication
20 June
1975.
Downloaded from www.physiology.org/journal/jappl by ${individualUser.givenNames} ${individualUser.surname} (192.236.036.029) on August 12, 2018. Copyright © 1976 American Physiological Society. All rights reserved.
CSF
pH
AND
Pco2
VALUES
125
REFERENCES 1. DAVIES, D. G., R. S. FITZGERALD, AND G. H. GURTNER. Acid-base relationships between CSF and blood during acute metabolic acidosis. J. Appl. Physiol. 34: 243-248, 1974. 2. DAVSON, H. Physiology of the Cerebrospinul FZuid. Boston, Mass. : Little, Brown, 1967, p. 272-279. 3. LEE, J. E., R. CHU, J. B. POSNER, AND F. PLUM. Buffering capacity of cerebrospinal fluid in experimental acidosis. Am. J. Physiol. 217: 1035-1038, 1969. 4. MITCHELL, R. A., D. A. HERBERT, AND C. T. CARMEN. Acid-base constants and temperature coeffkients for cerebrospinal fluid. J. Appl. Physiol. 20: 27-30, 1965.
5. SIGGAARD-ANDERSEN, 0. The pH-log Pcoz blood acid-base nomogram. Stand. J. Clin. Lab Invest. 14: 598-604, 1962. 6. SIGGAARD-ANDERSEN, O., AND K. ENGLE. A new acid-base nomogram. &and. J. Clin. Lab Invest. 12: 177-186, 1960. 7. SIGGAARD-ANDERSEN, O., K. ENGLE, K. JORGENSEN, AND P. AsTRUP. A micromethod for determination of pH, carbon dioxide, base excess and standard bicarbonate in capillary blood. Stand. J. Clin. Lab Invest. 12: 172-176, 1960. 8. VAN HEIJST, A. N. P., B. F. VISSER, AND A. H. J. MASS. A micromethod for the determination of pH and Pco2 in human cerebrospinal fluid. Clin. Chim. Actu 6: 589490, 1961.
Downloaded from www.physiology.org/journal/jappl by ${individualUser.givenNames} ${individualUser.surname} (192.236.036.029) on August 12, 2018. Copyright © 1976 American Physiological Society. All rights reserved.
