Metabolism Clinical and Experimental VOL. XXVIII,
NO. 3
MARCH
1979
-
PRELIMINARY REPORT
Effects
of Insulin Infusion on Plasma Phosphate in Diabetic Patients Michael S. Riley, David S. Schade,
A clinical association between insulin therapy and hypophosphatemia has frequently been made but a dose-response relationship has not been reported. Furthermore, the rapidity by which hypophosphatemia may be induced following an increment in plasma-free insulin concentration is not well defined. Therefore this study compared the effects of different rates of insulin infusion on the changes in plasma phosphate concentration in ketotic, hyparglycemic diabetic man. Sixteen prospective studies were performed in four insulin-dependent ketotic diabetic subjects. Insulin was infused according to one of four different protocols: high dose (1.0 U/kg/hr), low dose (0.1 U/kg/hr), very low dose (0.01 U/kg/hr) and control (saline only). Plasma phosphate, glucose, and free insulin concentrations were measured sequentially during the 60 min infusion periods. We observed that plasma phosphate concentrations declined significantly only with lowdose and high-dose insulin infusions. The magnitude and rapidity of fall of the mean phosphate concentration were greatest with high-dose insulin infusion. Significant hypophosphatemia can be observed within 30 min following the onset of insulin therapy.
is a potentially H YPOPHOSPHATEMIA serious complication of insulin therapy for diabetic ketoacidosis because of the resultant depletion of erythyrocyte 2,3 diphosphoglycerate concentration resulting in impaired tissue oxygen delivery.‘-’ As early as 1939, a direct relationship between insulin administration and a reduction in plasma phosphate concentration was observed and later confirmed by many other investigators.3 A recent clinical study by Piters and coworkers suggested that the incidence of hypophosphatemia may depend upon the dose of insulin administered.4 In order to extend this Metabolism, Vol. 28, No. 3 (March), 1979
and R. Philip Eaton
observation, we examined the relationship between the dose of insulin infused and the rate and magnitude of change in plasma phosphate concentration in hyperglycemic, ketotic diabetic volunteers. MATERIALS
AND
METHODS
The study was conducted utilizing a similar protocol to that described in detail in a previous publication.” Sixteen paired studies were completed in four insulin-dependent diabetic subjects. All subjects were studied between 7 a.m. and 9 a.m after an overnight fast and 24 hr following their last therapeutic insulin injection. Hyperglycemia and hyperketonemia were induced by the administration of 1.0 mg of dexamethasone orally at 8 and 24 hr prior to study (mean plasma bicarbonate = 18 + 2 mEq/liter; mean plasma ketones 2.5 f 0.3 mmol/liter). This protocol was utilized to allow the infusion of insulin for 60 min without the concurrent development of severe hypoglycemia. In all studies, insulin in albuminized saline (or saline alone in the control study) was infused into an indwelling scalp-vein needle located in an antecubital vein, and blood for assay was removed from the contralateral antecubital vein through
From the University of New Mexico School of Medicine, Division of Endocrinology and Metabolism. .4lbuquerque. N.M. Received for publication August 28, 1978. Supported by grants from the Clinical Research Center Program of the Division of Research Resources, National Institutes of Health #RR-O0997-02. The KROC Foundation, and NIAMD #I ROI AM 18681-01. David S. Schade is the recipient of a Research Career Development Award #I KZ4AM00260-01. Address reprint requests to David S. Schade, M.D.. University of New Mexico School of Medicine, Division of Endocrinology and Metabolism, Albuquerque, N.M. 87131. ~1979 by Grune & Stratton, Inc. 002&0495/79/2803-0001$01.00/0 191
RILEY, SCHADE. AND
192
Table 1. Plasma Phosphate, Glucose, and Insulin Concentration
EATON
During Insulin Infusion
M,FlUlE5 start
lnfuslon Dose Insulin
Control PO4 $ Glucose Insulin
-10
Basal -1
10
1
20
30
40
50
60
3.5 f 0 2
3.5 f 0.3’
3.5 I 0.3
3.6 f 0.3’
3.7 * 0.2
4.2 f 0.1
4.0 f 0.3
3.3 i 0.4
363 f 42
365 i 48
353 f 46
19 & 4
22 * 5
3.0 f 0.4
3.6 I 0.4
3.7 i 0.3
3.4 f 0.4
3.5 f 0.4
3.6 i 0.3
319 + 34
318 & 35
307 f 37
305 f 37
297 f 30’
20 * 7
28 f 4
28 zt 4
26 f 3
26 f 4
24 f
5
355 f 49
356 zt 48
21 f
21 14
19*4
348 i
50
344 f 49
341 * 49
5
17 + 2
21 54
0.1 pU/kg/hr PO4 Glucose lnsulm
326 i
36
22 i 6
34 f 0.1 298 i
36
26 * 4
3.6 + 0.3 297 * 35 27 & 6
0.1 &N/kg/hr PO4 Glucose Insulin
4.1 f 0.3 311 f
55
4.0 f 0.4 308546
22 * 3
22 z+ 3
4.0 f 0.2
3.5 * 0.4
306zt
51
286 zt 57
59+
17
86 i 26’
3.2 i 0 3 268 f
59’
97 f 35
3.1 f 0.2
3.4 f 03
3.1 * 02’
249 f
57’
233 zt 56t
215 i
124 I
51
119+47*
110 f 32’
58’
1 .O @U/kg/hr 4.5 * 0.4
4.0 f 0.3
Glucose
PO4
383 i 32
4.3 * 0.3
373 f 36
340 i 34’
Insulin
21 zt3
23 rt 5
814 f 257’
3.0 f 0.3’ 281 f
16’
1131 i 316’
2.6 f 0.3t 241 i 25’ 1290 f 438’
2.5 f 0.3t
2.6 zk 0.3’ 189 i
24’
1348 f 231’
2.3 i 0.2’
149 f 27’
137 i- 23’
1356 f 336’
1955 f 640’
‘Compared to basal concentration, = p < 0.05 t Compared to basal concentration, = p < 0.01 $POa Img/dl). Glucose (mg/dl). Insulin WJ/ml).
another indwelling scalp-vein needle. Total volume of saline infused did not exceed 120 ml. On different days the following four infusion regimens were performed in a random sequence on each subject: (1) a control saline infusion; (2) a very-low-dose insulin infusion (0.01 U/kg/hr); (3) a lowdose insulin infusion (0.1 U/kg/hr), and (4) a high-dose insulin infusion (I .O U/kg/hr). Samples were collected for phosphate, glucose, and insulin determinations at IO min intervals before and during the saline or insulin in saline infusion as depicted in Table 1. Glucose was assayed by glucose oxidase6 and insulin by radioimmunoassay’ following precipitation of endogenous antibodies by polyethylene glycol.8 Phosphate concentration was determined spectrophotometrically.9 Variance of the mean is expressed as the standard error, and statistical difference between the basal mean and the post insulin infusion mean was done utilizing Student’s t test.” RESULTS
Plasma Phosphate (Fig. 1, Table 1)
The plasma phosphate concentration at each dose of insulin administered is plotted versus time in Fig. 1. The variance of each mean and the statistical difference are given in Table 1. Plasma phosphate demonstrated a small but statistically significant decline at two observation points (+30 and i-50 minutes) in the
control saline infusion study. Infusion of verylow-dose insulin (0.01 U/kg/hr) resulted in no significant decline in plasma phosphate concentration. During the low-dose insulin infusion (0.1 U/kg/hr), plasma phosphate declined progressively and the final concentration at 60 min was significantly below the initial starting basal concentration (~~0.05). With high-dose insulin infusion (C 1.0 U/kg/hr) plasma phosphate declined rapidly from a basal concentration of 4.5 +_ 0.4 mg/dl to a final value of 2.3 f 0.2 mg/dl, which was approximately 50% below the basal preinfusion concentration. This rapid decline in plasma phosphate concentration was statistically significant at five out of the six observation points (~~0.05). Plasma Glucose (Table 1)
Plasma glucose concentration did not significantly change during the saline control study (p > 0.05). During insulin infusion, plasma glucose declined in a dose-response fashion as has been previously observed.’
INSLILIN AND
PHOSPHATE
193 BASAl
-
SALINE OR tNSULIN
lNFUSlON _7
r-r
j
Fig. 1. Mean plasma phosphate plotted against iime for each concentration of insulin administered. Significance values shown are for final phosphate concentration compared to basal phosphate concentration.
”
-
Plasma Insulin Concentration (Table 1)
Plasma insulin did not significantly change in the control saline study (Table 1). However, with log increments in plasma insulin infusion concentration, approximately log increments in plasma-free insulin concentration were observed. DISCUSSION
The observation that plasma phosphate concentration significantly declined during the treatment of diabetic ketoacidosis was reported by Guest in I 939.3 This observation was followed by the recommendation that administration of phosphate should be employed in the standard treatment of diabetic ketoacidosis.” More recently, this recommendation has gained much support from the improved understanding of the role of phosphate in intermediary metabolism. Recent reports’2,” have stressed the regulatory role that plasma phosphate concentration has on the erythrocyte level of 2,3 diphosphoglycerate. This erythrocyte constituent enhances the release of oxygen from hemoglobin at the tissue level. Thus, the hypophosphatemia that accompanies insulin therapy for diabetic ketoacidosis may decrease tissue oxygen delivery by inhibiting erythrocyte synthesis of 2,3 diphosphoglycer-
ate.“14 Furthermore, other adverse effects of hypophosphatemia have been described. Impaired central nervous system functionI and hepatic dysfunction” may occur. Red blood cell hemolysis,‘6 impaired phagocytosis,” and impaired platelet function’* have also been reported. The hypophosphatemia that accompanies treatment of diabetic ketoacidosis may be due to multiple factors (including insulin administration). However, the recent clinical report by Piters et ah4 comparing the metabolic response in diabetic ketoacidosis to both high- and lowdosage insulin regimens strongly suggests that high-dose insulin therapy may induce severe hypophosphatemia. Our study extended this clinical study to a prospective, randomized paired protocol in which hyperglycemia and hyperketonemia were pharmacologically induced.’ Our results demonstrate that not only is the insulin-induced hypophosphatemia dosedependent, but that significant hypophosphatemia may occur within 30 min following the administration of insulin. ACKNOWLEDGMENT This study was completed with the excellent technical assistance of Enid Pinero MT/ASCP and Jerry Towle MT/ASCP.
REFERENCES 1. Ditzel J, Stand1 E: The problem of tissue oxygenation in diabetes mellitus. Acta Med Stand (Suppl) 578:59-68, 1975 2. Kanter Y, Gerson JR, Bessman AN: 2,3-Diphosphoglycerate, nucleotide phosphate, and organic and inorganic
phosphate during the early phases of diabetic ketoacidosis. Diabetes 26:429-433, 1977 3. Guest GM, Rapoport S: Role of acid soluble phosphorous compounds in red blood cells in experimental rickets, renal insufficiency. pyloric obstruction, gastroenteritis,
194
ammonium chloride acidosis and diabetic acidosis. Am J Dis Child 58:1072-1084, 1939 4. Piters KM, Kumar D, Pei E, et al: Comparison of continuous and intermittent intravenous insulin therapies for diabetic ketoacidosis. Diabetologia 13:3 17-321, 1977 5. Schade DS, Eaton RP: Dose response to insulin in man: Differential effects on glucose and ketone body regulation. J Clin Endocrinol Metab44:1038-1053, 1977 6. Hagett ASG, Nixon DA: Use of glucose oxidase, peroxidase, and 0-diansidine in determination of blood and urinary glucose. Lancet 2:368-370, 1957 7. Hales CM, Rangle PJ: Immunoassay of insulin and antibody precipitation. Biochem J 88: 137-146, 1963 8. Nakagawa SH, Nakayama H, Sassaki T. et al: A simple method for the determination of serum free insulin levels in insulin-treated patients. Diabetes 22:590-600, 1973 9. Taussky A, Schorr E: A microcolorimetric method for the determination of inorganic phosphorous. J Biol Chem 202:675-685, 1953 10. Armitage P: Statistical Methods in Medical Research. New York, Wiley & Sons, 1971, p 104 Il. Franks M, Berris RF, Kaplan NO, et al: Metabolic studies in diabetic acidosis: The effect of the administration of sodium phosphate. Arch Intern Med 8 I :42-56, 1948 12. Travis SF, Sugerman HJ, Ruberg RL, et al: Alterations of red cell glycolytic intermediates and oxygen trans-
RILEY,
SCHADE,
AND
EATON
port as a consequence of hypophosphatemia in patients receiving intravenous hyperalimentation. N Engl J Med 285763-768, 1971 13. Lichtman MA, Miller DR, Cohen J, et al: Reduced red cell glycolysis, 2, 3-Diphosphoglycerate and adenosine triphosphate concentration and increased hemoglobinoxygen affinity caused by hypophosphatemia. Ann Intern Med 74:562-566, 197 I 14. Alberti KGMM, Darley JH, Emerson PM, et al: 2,3-Diphosphoglycerate and tissue oxygenation in uncontrolled diabetes mellitus. Lancet 2:391-395. 1972 15. Rajan KS, Levison R, Levy CM: Hepatic hypoxia secondary to hypophosphatemia. Clin Res 21:521-525. 1973 16. Jacob HS, Ansden P: Acute hemolytic anexia rigid red cells in hypophosphatemia. N Engl J 285:1446-1450, 1971
with Med
17. Craddock PR, Yawatc Y. VanSanten L: Acquired phagocyte dysfunction: A complication of the hypophosphatemia of parenteral hyperalimentation. N Engl J Med 290:1403-1414. 1974 18. Yawata Y, Hebbel RP, Silvis S, et al: Blood cell abnormalities complicating the hypophosphatemia of hyperalimentation: Erythrocytes and platelet ATP deficiency associated with hemolytic anemia and bleeding in hyperalimented dogs. J Lab Clin Med 84:643-653. 1974
NO. 3
MARCH
1979
-
PRELIMINARY REPORT
Effects
of Insulin Infusion on Plasma Phosphate in Diabetic Patients Michael S. Riley, David S. Schade,
A clinical association between insulin therapy and hypophosphatemia has frequently been made but a dose-response relationship has not been reported. Furthermore, the rapidity by which hypophosphatemia may be induced following an increment in plasma-free insulin concentration is not well defined. Therefore this study compared the effects of different rates of insulin infusion on the changes in plasma phosphate concentration in ketotic, hyparglycemic diabetic man. Sixteen prospective studies were performed in four insulin-dependent ketotic diabetic subjects. Insulin was infused according to one of four different protocols: high dose (1.0 U/kg/hr), low dose (0.1 U/kg/hr), very low dose (0.01 U/kg/hr) and control (saline only). Plasma phosphate, glucose, and free insulin concentrations were measured sequentially during the 60 min infusion periods. We observed that plasma phosphate concentrations declined significantly only with lowdose and high-dose insulin infusions. The magnitude and rapidity of fall of the mean phosphate concentration were greatest with high-dose insulin infusion. Significant hypophosphatemia can be observed within 30 min following the onset of insulin therapy.
is a potentially H YPOPHOSPHATEMIA serious complication of insulin therapy for diabetic ketoacidosis because of the resultant depletion of erythyrocyte 2,3 diphosphoglycerate concentration resulting in impaired tissue oxygen delivery.‘-’ As early as 1939, a direct relationship between insulin administration and a reduction in plasma phosphate concentration was observed and later confirmed by many other investigators.3 A recent clinical study by Piters and coworkers suggested that the incidence of hypophosphatemia may depend upon the dose of insulin administered.4 In order to extend this Metabolism, Vol. 28, No. 3 (March), 1979
and R. Philip Eaton
observation, we examined the relationship between the dose of insulin infused and the rate and magnitude of change in plasma phosphate concentration in hyperglycemic, ketotic diabetic volunteers. MATERIALS
AND
METHODS
The study was conducted utilizing a similar protocol to that described in detail in a previous publication.” Sixteen paired studies were completed in four insulin-dependent diabetic subjects. All subjects were studied between 7 a.m. and 9 a.m after an overnight fast and 24 hr following their last therapeutic insulin injection. Hyperglycemia and hyperketonemia were induced by the administration of 1.0 mg of dexamethasone orally at 8 and 24 hr prior to study (mean plasma bicarbonate = 18 + 2 mEq/liter; mean plasma ketones 2.5 f 0.3 mmol/liter). This protocol was utilized to allow the infusion of insulin for 60 min without the concurrent development of severe hypoglycemia. In all studies, insulin in albuminized saline (or saline alone in the control study) was infused into an indwelling scalp-vein needle located in an antecubital vein, and blood for assay was removed from the contralateral antecubital vein through
From the University of New Mexico School of Medicine, Division of Endocrinology and Metabolism. .4lbuquerque. N.M. Received for publication August 28, 1978. Supported by grants from the Clinical Research Center Program of the Division of Research Resources, National Institutes of Health #RR-O0997-02. The KROC Foundation, and NIAMD #I ROI AM 18681-01. David S. Schade is the recipient of a Research Career Development Award #I KZ4AM00260-01. Address reprint requests to David S. Schade, M.D.. University of New Mexico School of Medicine, Division of Endocrinology and Metabolism, Albuquerque, N.M. 87131. ~1979 by Grune & Stratton, Inc. 002&0495/79/2803-0001$01.00/0 191
RILEY, SCHADE. AND
192
Table 1. Plasma Phosphate, Glucose, and Insulin Concentration
EATON
During Insulin Infusion
M,FlUlE5 start
lnfuslon Dose Insulin
Control PO4 $ Glucose Insulin
-10
Basal -1
10
1
20
30
40
50
60
3.5 f 0 2
3.5 f 0.3’
3.5 I 0.3
3.6 f 0.3’
3.7 * 0.2
4.2 f 0.1
4.0 f 0.3
3.3 i 0.4
363 f 42
365 i 48
353 f 46
19 & 4
22 * 5
3.0 f 0.4
3.6 I 0.4
3.7 i 0.3
3.4 f 0.4
3.5 f 0.4
3.6 i 0.3
319 + 34
318 & 35
307 f 37
305 f 37
297 f 30’
20 * 7
28 f 4
28 zt 4
26 f 3
26 f 4
24 f
5
355 f 49
356 zt 48
21 f
21 14
19*4
348 i
50
344 f 49
341 * 49
5
17 + 2
21 54
0.1 pU/kg/hr PO4 Glucose lnsulm
326 i
36
22 i 6
34 f 0.1 298 i
36
26 * 4
3.6 + 0.3 297 * 35 27 & 6
0.1 &N/kg/hr PO4 Glucose Insulin
4.1 f 0.3 311 f
55
4.0 f 0.4 308546
22 * 3
22 z+ 3
4.0 f 0.2
3.5 * 0.4
306zt
51
286 zt 57
59+
17
86 i 26’
3.2 i 0 3 268 f
59’
97 f 35
3.1 f 0.2
3.4 f 03
3.1 * 02’
249 f
57’
233 zt 56t
215 i
124 I
51
119+47*
110 f 32’
58’
1 .O @U/kg/hr 4.5 * 0.4
4.0 f 0.3
Glucose
PO4
383 i 32
4.3 * 0.3
373 f 36
340 i 34’
Insulin
21 zt3
23 rt 5
814 f 257’
3.0 f 0.3’ 281 f
16’
1131 i 316’
2.6 f 0.3t 241 i 25’ 1290 f 438’
2.5 f 0.3t
2.6 zk 0.3’ 189 i
24’
1348 f 231’
2.3 i 0.2’
149 f 27’
137 i- 23’
1356 f 336’
1955 f 640’
‘Compared to basal concentration, = p < 0.05 t Compared to basal concentration, = p < 0.01 $POa Img/dl). Glucose (mg/dl). Insulin WJ/ml).
another indwelling scalp-vein needle. Total volume of saline infused did not exceed 120 ml. On different days the following four infusion regimens were performed in a random sequence on each subject: (1) a control saline infusion; (2) a very-low-dose insulin infusion (0.01 U/kg/hr); (3) a lowdose insulin infusion (0.1 U/kg/hr), and (4) a high-dose insulin infusion (I .O U/kg/hr). Samples were collected for phosphate, glucose, and insulin determinations at IO min intervals before and during the saline or insulin in saline infusion as depicted in Table 1. Glucose was assayed by glucose oxidase6 and insulin by radioimmunoassay’ following precipitation of endogenous antibodies by polyethylene glycol.8 Phosphate concentration was determined spectrophotometrically.9 Variance of the mean is expressed as the standard error, and statistical difference between the basal mean and the post insulin infusion mean was done utilizing Student’s t test.” RESULTS
Plasma Phosphate (Fig. 1, Table 1)
The plasma phosphate concentration at each dose of insulin administered is plotted versus time in Fig. 1. The variance of each mean and the statistical difference are given in Table 1. Plasma phosphate demonstrated a small but statistically significant decline at two observation points (+30 and i-50 minutes) in the
control saline infusion study. Infusion of verylow-dose insulin (0.01 U/kg/hr) resulted in no significant decline in plasma phosphate concentration. During the low-dose insulin infusion (0.1 U/kg/hr), plasma phosphate declined progressively and the final concentration at 60 min was significantly below the initial starting basal concentration (~~0.05). With high-dose insulin infusion (C 1.0 U/kg/hr) plasma phosphate declined rapidly from a basal concentration of 4.5 +_ 0.4 mg/dl to a final value of 2.3 f 0.2 mg/dl, which was approximately 50% below the basal preinfusion concentration. This rapid decline in plasma phosphate concentration was statistically significant at five out of the six observation points (~~0.05). Plasma Glucose (Table 1)
Plasma glucose concentration did not significantly change during the saline control study (p > 0.05). During insulin infusion, plasma glucose declined in a dose-response fashion as has been previously observed.’
INSLILIN AND
PHOSPHATE
193 BASAl
-
SALINE OR tNSULIN
lNFUSlON _7
r-r
j
Fig. 1. Mean plasma phosphate plotted against iime for each concentration of insulin administered. Significance values shown are for final phosphate concentration compared to basal phosphate concentration.
”
-
Plasma Insulin Concentration (Table 1)
Plasma insulin did not significantly change in the control saline study (Table 1). However, with log increments in plasma insulin infusion concentration, approximately log increments in plasma-free insulin concentration were observed. DISCUSSION
The observation that plasma phosphate concentration significantly declined during the treatment of diabetic ketoacidosis was reported by Guest in I 939.3 This observation was followed by the recommendation that administration of phosphate should be employed in the standard treatment of diabetic ketoacidosis.” More recently, this recommendation has gained much support from the improved understanding of the role of phosphate in intermediary metabolism. Recent reports’2,” have stressed the regulatory role that plasma phosphate concentration has on the erythrocyte level of 2,3 diphosphoglycerate. This erythrocyte constituent enhances the release of oxygen from hemoglobin at the tissue level. Thus, the hypophosphatemia that accompanies insulin therapy for diabetic ketoacidosis may decrease tissue oxygen delivery by inhibiting erythrocyte synthesis of 2,3 diphosphoglycer-
ate.“14 Furthermore, other adverse effects of hypophosphatemia have been described. Impaired central nervous system functionI and hepatic dysfunction” may occur. Red blood cell hemolysis,‘6 impaired phagocytosis,” and impaired platelet function’* have also been reported. The hypophosphatemia that accompanies treatment of diabetic ketoacidosis may be due to multiple factors (including insulin administration). However, the recent clinical report by Piters et ah4 comparing the metabolic response in diabetic ketoacidosis to both high- and lowdosage insulin regimens strongly suggests that high-dose insulin therapy may induce severe hypophosphatemia. Our study extended this clinical study to a prospective, randomized paired protocol in which hyperglycemia and hyperketonemia were pharmacologically induced.’ Our results demonstrate that not only is the insulin-induced hypophosphatemia dosedependent, but that significant hypophosphatemia may occur within 30 min following the administration of insulin. ACKNOWLEDGMENT This study was completed with the excellent technical assistance of Enid Pinero MT/ASCP and Jerry Towle MT/ASCP.
REFERENCES 1. Ditzel J, Stand1 E: The problem of tissue oxygenation in diabetes mellitus. Acta Med Stand (Suppl) 578:59-68, 1975 2. Kanter Y, Gerson JR, Bessman AN: 2,3-Diphosphoglycerate, nucleotide phosphate, and organic and inorganic
phosphate during the early phases of diabetic ketoacidosis. Diabetes 26:429-433, 1977 3. Guest GM, Rapoport S: Role of acid soluble phosphorous compounds in red blood cells in experimental rickets, renal insufficiency. pyloric obstruction, gastroenteritis,
194
ammonium chloride acidosis and diabetic acidosis. Am J Dis Child 58:1072-1084, 1939 4. Piters KM, Kumar D, Pei E, et al: Comparison of continuous and intermittent intravenous insulin therapies for diabetic ketoacidosis. Diabetologia 13:3 17-321, 1977 5. Schade DS, Eaton RP: Dose response to insulin in man: Differential effects on glucose and ketone body regulation. J Clin Endocrinol Metab44:1038-1053, 1977 6. Hagett ASG, Nixon DA: Use of glucose oxidase, peroxidase, and 0-diansidine in determination of blood and urinary glucose. Lancet 2:368-370, 1957 7. Hales CM, Rangle PJ: Immunoassay of insulin and antibody precipitation. Biochem J 88: 137-146, 1963 8. Nakagawa SH, Nakayama H, Sassaki T. et al: A simple method for the determination of serum free insulin levels in insulin-treated patients. Diabetes 22:590-600, 1973 9. Taussky A, Schorr E: A microcolorimetric method for the determination of inorganic phosphorous. J Biol Chem 202:675-685, 1953 10. Armitage P: Statistical Methods in Medical Research. New York, Wiley & Sons, 1971, p 104 Il. Franks M, Berris RF, Kaplan NO, et al: Metabolic studies in diabetic acidosis: The effect of the administration of sodium phosphate. Arch Intern Med 8 I :42-56, 1948 12. Travis SF, Sugerman HJ, Ruberg RL, et al: Alterations of red cell glycolytic intermediates and oxygen trans-
RILEY,
SCHADE,
AND
EATON
port as a consequence of hypophosphatemia in patients receiving intravenous hyperalimentation. N Engl J Med 285763-768, 1971 13. Lichtman MA, Miller DR, Cohen J, et al: Reduced red cell glycolysis, 2, 3-Diphosphoglycerate and adenosine triphosphate concentration and increased hemoglobinoxygen affinity caused by hypophosphatemia. Ann Intern Med 74:562-566, 197 I 14. Alberti KGMM, Darley JH, Emerson PM, et al: 2,3-Diphosphoglycerate and tissue oxygenation in uncontrolled diabetes mellitus. Lancet 2:391-395. 1972 15. Rajan KS, Levison R, Levy CM: Hepatic hypoxia secondary to hypophosphatemia. Clin Res 21:521-525. 1973 16. Jacob HS, Ansden P: Acute hemolytic anexia rigid red cells in hypophosphatemia. N Engl J 285:1446-1450, 1971
with Med
17. Craddock PR, Yawatc Y. VanSanten L: Acquired phagocyte dysfunction: A complication of the hypophosphatemia of parenteral hyperalimentation. N Engl J Med 290:1403-1414. 1974 18. Yawata Y, Hebbel RP, Silvis S, et al: Blood cell abnormalities complicating the hypophosphatemia of hyperalimentation: Erythrocytes and platelet ATP deficiency associated with hemolytic anemia and bleeding in hyperalimented dogs. J Lab Clin Med 84:643-653. 1974
