Sodium salicylamide: Relative bioavailability and subjective effects The bioavailability of sodium salicylamide (NaSAM) in solution of salicylamide (SAM) tablets was compared in 6 healthy human volunteers. Bioavailability was assessed by plasma level determinations of nonmetabolized salicylamide (free SAM) and salicylamide plus conjugated metabolites (total SAM) for 3 hr following oral doses of 0.65, 1.30, 1.95, and 2.60 gm of salicylamide. The availability of NaSAM was found to be superior to SAM and dose-dependent. Mean peak levels offree SAM and total SAM were higher and were reached earlier after NaSAM liquid than after SAM tablets. Significantly higher mean levels offree SAM were found at the 1.95 and 2.60 gm dose levels after NaSAM administration than after SAM. Mean total SAM concentration was significantly higher after NaSAM at all dosage levels. The sedative effects of salicylamide were assessed with a self-scoring questionnaire. Sedation seemed to increase with increasing dose of both NaSAM and SAM. The sedative response occurred earlier after NaSAM than after SAM. Side effects were minor and transient in nature, occurred at the higher dosage levels, and were predominantly lightheadedness and dizziness. Because NaSAM produces higher drug levels and has a more rapid onset of subjective effects, we conclude that it represents a potentially superior dosage form.
Lawrence Fleckenstein, Pharm.D., Gregory R. Mundy, M.D., Richard A. Horovitz, M.D., and John M. Mazzullo, M.D. Rochester, N. Y.
Department of Pharmacology and Toxicology, University of Rochester School of Medicine and Dentistry
Salicylamide (SAM) is used in many overthe-counter preparations. There are discrepancies between clinical studies in man and experimental studies in animals and its efficacy is disputed. In animals SAM has analgesic,5, 11, 19 antipyretic, 5, 9 and hypnotic 6 activity. Human trials have been less convincing. Double-blind studies have indicated Received for publication July 10, 1975. Accepted for publication Dec. 26, 1975. Reprint requests to: L. Fleckenstein, Pharm.D., Drug Information Service, Alta Bates Hospital, Webster and Colby Streets, Berkeley, Calif. 94705.
poor analgesic activity after 0.6 gm 4 , 18 and favorable analgesia with doses of 2 gm every 4 to 8 hr. 13 For antipyretic efficacy, there are one positive 17 and 2 less favorable trials in the literature. 7, 15 To our knowledge there are no reports of hypnotic activity for salicylamide in humans, but there are 2 positive reports of SAM combined with either acetaminophen or acetanilid. 8, 20 Recently assays have been developed that are sensitive enough to measure plasma levels after normal therapeutic doses; plasma levels were not determined in the human trials cited. 451
452
Fleckenstein et al.
Clinical Pharmacology and Therapeutics
Table I. Data on subjects Age Volunteer (yr)
Subject Subject Subject Subject Subject Subject
I 2 3 4 5 6
25 29 24 23 23 24
Body surWeight face area (kg) (m 2)
58.0 84.1 76.7 90.0 87.0 69.5
1.48 1.90 1.80 1.97 2.00 1.70
Serum creatinine/BUN (mg%)
1.2/11 1.2/13 1.2/18 1.0/16 1.1/16 1.2/13
It has been shown that SAM is eliminated from the body by competing zero- and first-order processes and that the availability of SAM in solution is dose-and dosage form-dependent. 12 It has also been shown that there is a significant "first-pass" effect 1 and the availability of SAM from solid dosage forms is dissolution rate-limited. 3 Since SAM is poorly soluble in water, it is possible that the poor results in some of these human trials are the result of poor bioavailability. In our study, we have investigated the bioavailability of NaSAM, a more water-soluble form of SAM. A further purpose was to detect and quantify subjective central nervous system effects. Methods
Six healthy male volunteers were paid for participation in the study. Subjects were not using medication of any sort for a period of 7 days preceding the study nor during it. Subjects were specifically asked not to take salicylamide, aspirin, or combination preparations containing these drugs during the study. A medical history was taken and a physical examination was performed on each subject before and after the study. The following tests were in the normal range: electrocardiogram, hematocrit, white blood cell count, differential cell count, platelet count, urinalysis, SMA-12, and prothrombin time. Age, weight, body surface area, and serum creatinine and blood urea nitrogen values for each subject are presented in Table I. Design. On the study night each subject received a single oral dose of 8 tablets and 120 ml of liquid at 5 P.M. On a given day, subjects
received either active tablets (SAM) with 120 ml of placebo liquid or active liquid (NaSAM in 120 ml) and placebo tablets. * This procedure was repeated once a week for 13 wk. In no case did drug administration take longer than 30 sec. Subjects fasted from 12 A.M. and were not permitted tea or coffee after 10 A.M. on study days. The attempt was made to conduct the study in double-blind fashion but in some instances the subjects recognized a characteristic taste in the NaSAM solution. The dose of NaSAM was gradually increased from 0.65 to 2.60 gm over 13 wk. (Results of the doseranging part of the study will be discussed in a subsequent report.) Only the data for the 0.65, 1.30, 1.95, and 2.60 gm dosage levels of both SAM and NaSAM will be presented in this report. At each dosage level (0.65, 1.30, 1.95, and 2.60 gm) the sequence of drug administration was randomized. Subject 6 was dropped from the study before receiving the 2.60 gm dose and therefore at this dosage a complete crossover was complete for only 5 subjects. Each subject remained on an examining table supine, at 45 0 gatch, during the experimental period. A number 19 G butterfly needle was placed in a forearm vein or the antecubital fossa and kept open with normal saline for the duration of the experimental period (about 3Yz hr). Blood samples were removed through 3-way stopcocks placed in the intravenous line. Less than a liter of normal saline was used over approximately 3Yz hr to keep the line open and to flush it after sampling. Whole blood, 10 ml, was drawn at zero min and at 2, 5, 10, 20, 30, 45, 60, 90, 120, and 180 min after the dose. After the 1.30 gm dose, blood samples were also collected 15 min after dosing. The whole blood was transferred to citrated tubes, centrifuged within 30 min of collection, and the plasma stored at -20 C until assayed. Assay. Free SAM was assayed by minor modification of a fluorometric method described by Barr and Riegelman.2, Plasma, 0.5 ml, was 0
• Sodium salicylamide and placebo preparations were supplied by Vick Divisions Research. Commercially purchased salicylamide tablets, distributed by Harvey Laboratories, Philadelphia, Lot 20001, were used. They had the most rapid disintegration time of several N.F. tablets screened for use.
Sodium salicylamide
Volume 19 Number 4
-
FREE
453
SALICYLAMIDE
--- TOTAL SALICYLAMIDE • o
TIME
0.650 gm. SALICYLAMIDE 0.650 gm. SODIUM SALICYLAMIDE Cn = G)
(min.)
Fig. 1. Mean (± SEM) plasma levels of free and total salicylamide after administration of 0.65 gm of SAM equivalents as SAM tablets and NaSAM solution.
added to 1.0 ml of 0.2 M phosphate buffer (pH 7.0) and extracted twice into 9.0 ml of ethylene dichloride. The mixture was then reextracted into 10 ml of aqueous 0.25 N sodium hydroxide solution. The fluorescence of SAM in basic solution was determined by an AmincoBowman spectrophotofluorometer at maximum activation and emission wavelengths of 332 and 418 nm, respectively. Recovery of SAM from spiked plasma samples was in excess of 95%. Total SAM was determined by hydrolyzing all metabolites to a mixture of salicylic acid and SAM.t6 To the 0.5 ml plasma samples, 0.5 ml water and 2.0 ml of 12.5% v/v sulfuric acid were added in screw-capped centrifuge tubes. Samples were autoclaved at 15 pounds pressure and 120° C for 20 min. Samples were then extracted in the same manner as described for plasma SAM above. The fluorescence of hydrolyzed SAM was determined at an emission wavelength of 400 nm using activation wavelengths of 332 and 304 nm (to include any SAM converted to salicylic acid during hydrolysis). The total SAM concentration was then determined from a standard curve by taking the sum of the two fluorescent measurements corrected with a predosing plasma bank. The standard curve was linear over the range of 0.1 to 50 mcg/m!. The data for drug concentration in plasma were analyzed for significant treatment differences by applying a 3-way analysis of vari-
ance, treating the data as if it were gathered from a split-plot design experiment. Subjective questionnaire. The subjective effects of SAM were evaluated by means of a self-scoring questionnaire similar to that described by Eade and Lasagna. tO On the first page of the questionnaire subjects were asked to discriminate between opposite pairs of subjective responses such as "sleepy" and "awake." They were asked to rate the intensity of responses by circling a number on either side of a central zero point. The subject always had his previous score before him:
o min
15 min 45 min 120 min
3 3 3 3
Sleepy 2 2 2 2
0 0 0 0
I I I I
Awake 2 2 2 2
3 3 3 3
The second page of the questionnare listed 14 side effects. These items could be scored in intensity from 0 to 3 and the subject always had his previous score before him: Lightheadedness
o min
15 min 45 min 120 min
0 0 0 0
2 2 2 2
3 3 3 3
Prior to the study, volunteers were given a set of instructions, and underwent a trial run of the questionnaire. The subjects filled out the questionnaires prior to drug administration and
454
Fleckenstein et al.
Clinical Pharmacology and Therapeutics
-
FREE
SALICYLAMIDE
TOTAL SALICYLAMIDE 1.300 9m. SALICYLAMIDE 1.300 9m. SODIUM SALICYLAMIDE
• "
Cn = 6)
TIME
(min.)
Fig. 2. Mean (± SEM) plasma levels of free and total salicylamide after administration of 1.30 gm of SAM equivalents as SAM tablets and NaSAM solution.
-
FREE
SALICYLAMIDE
--- TOTAL SALICYLAMIDE •
1.950 9m. SALICYLAMIDE
()
1.950 9m. SODIUM SALICYLAMIDE (n =6)
TIME
(min.)
Fig. 3. Mean (± SEM) plasma levels of free and total salicylamide after administration of 1.95 gm of SAM equivalents as SAM tablets and NaSAM solution.
at 15, 45, and 120 min after taking the drug. The subjects were told they would be given an aspirin-like drug. In addition, they were told that the drug might stimulate, depress, or have no effect at all. They were asked to discriminate between these responses. Each sub-
ject was tested in isolation in the examining rooms of a medical clinic. They were asked not to discuss the subjective effects with the other volunteers. Subjects were allowed to study, but could not leave the examining room. In scoring the individual responses, the net
Sodium salicylamide
Volume 19 Number 4
TIME
455
(min.)
Fig. 4. Mean (± SEM) plasma levels of free and total salicylamide after administration of 2.60 gm of SAM equivalents as SAM tablets and NaSAM solution.
score for any item was the sum of the algebraic difference between the O-hr and the 15-, 45-, and 120- min. score. The scoring system allowed for positive or negative scores for the subjective reponses, but only a positive score for responses to side effects. Results
Free SAM. Figs. 1 to 4 show the mean (± SEM) free and total plasma levels, follow-
ing 0.65,1.30, 1.95, and 2.60 gm respectively, of SAM equivalents given as liquid sodium salt or tableted SAM. The peak mean plasma concentration of nonmetabolized (free) SAM was reached earlier and was higher after NaSAM than after SAM. At all dosage levels there is a difference between the treatments due to a treatment-sampling time interaction (p < 0.01); the free SAM concentration in blood after NaSAM is higher than that after SAM tablets at early sampling times (for the IO-min. sample, p < 0.01 at 0.65, 1.30, and 1.95 gm and p < 0.05 at 2.6 gm dose levels). The mean area under the plasma concentration curve (AUC) from NaSAM is greater than from
SAM tablets at the 0.65, 1.95, and 2.60 gm dose levels (p < 0.05, paired t test). The difference in mean AVe at the 1.30 gm dosage level is not significant. In Fig. 5 it can be seen that the systemic availability of free SAM is dose-dependent after NaSAM. As the dose of NaSAM is increased from 0.65 gm to 2.65 gm, the availability of free SAM rises in a nonlinear fashion. The availability of free SAM after SAM is also dose-dependent (not shown). Mean AVe values are lower and have a larger coefficient of variation at every dosage level of SAM than of NaSAM. Total SAM. The treatment: sampling time interaction is significant (p < 0.01) at all dosage levels; the total SAM concentration in plasma after NaSAM liquid is greater than after SAM at all sampling times from 10 through 60 min at all dosage levels (p < 0.05 at 10 and 60 min and p < 0.01 at 15, 30, and 45 min sampling times). The mean AUe for total SAM achieved by the subjects after NaSAM is greater than after SAM at the 0.65 (p < 0.05, paired t test) and 2.60 gm (p < 0.01, paired
456
Fleckenstein et al.
Clinical Pharmacology and Therapeutics
Table II. Incidence and magnitude of side effects
Lawrence Fleckenstein, Pharm.D., Gregory R. Mundy, M.D., Richard A. Horovitz, M.D., and John M. Mazzullo, M.D. Rochester, N. Y.
Department of Pharmacology and Toxicology, University of Rochester School of Medicine and Dentistry
Salicylamide (SAM) is used in many overthe-counter preparations. There are discrepancies between clinical studies in man and experimental studies in animals and its efficacy is disputed. In animals SAM has analgesic,5, 11, 19 antipyretic, 5, 9 and hypnotic 6 activity. Human trials have been less convincing. Double-blind studies have indicated Received for publication July 10, 1975. Accepted for publication Dec. 26, 1975. Reprint requests to: L. Fleckenstein, Pharm.D., Drug Information Service, Alta Bates Hospital, Webster and Colby Streets, Berkeley, Calif. 94705.
poor analgesic activity after 0.6 gm 4 , 18 and favorable analgesia with doses of 2 gm every 4 to 8 hr. 13 For antipyretic efficacy, there are one positive 17 and 2 less favorable trials in the literature. 7, 15 To our knowledge there are no reports of hypnotic activity for salicylamide in humans, but there are 2 positive reports of SAM combined with either acetaminophen or acetanilid. 8, 20 Recently assays have been developed that are sensitive enough to measure plasma levels after normal therapeutic doses; plasma levels were not determined in the human trials cited. 451
452
Fleckenstein et al.
Clinical Pharmacology and Therapeutics
Table I. Data on subjects Age Volunteer (yr)
Subject Subject Subject Subject Subject Subject
I 2 3 4 5 6
25 29 24 23 23 24
Body surWeight face area (kg) (m 2)
58.0 84.1 76.7 90.0 87.0 69.5
1.48 1.90 1.80 1.97 2.00 1.70
Serum creatinine/BUN (mg%)
1.2/11 1.2/13 1.2/18 1.0/16 1.1/16 1.2/13
It has been shown that SAM is eliminated from the body by competing zero- and first-order processes and that the availability of SAM in solution is dose-and dosage form-dependent. 12 It has also been shown that there is a significant "first-pass" effect 1 and the availability of SAM from solid dosage forms is dissolution rate-limited. 3 Since SAM is poorly soluble in water, it is possible that the poor results in some of these human trials are the result of poor bioavailability. In our study, we have investigated the bioavailability of NaSAM, a more water-soluble form of SAM. A further purpose was to detect and quantify subjective central nervous system effects. Methods
Six healthy male volunteers were paid for participation in the study. Subjects were not using medication of any sort for a period of 7 days preceding the study nor during it. Subjects were specifically asked not to take salicylamide, aspirin, or combination preparations containing these drugs during the study. A medical history was taken and a physical examination was performed on each subject before and after the study. The following tests were in the normal range: electrocardiogram, hematocrit, white blood cell count, differential cell count, platelet count, urinalysis, SMA-12, and prothrombin time. Age, weight, body surface area, and serum creatinine and blood urea nitrogen values for each subject are presented in Table I. Design. On the study night each subject received a single oral dose of 8 tablets and 120 ml of liquid at 5 P.M. On a given day, subjects
received either active tablets (SAM) with 120 ml of placebo liquid or active liquid (NaSAM in 120 ml) and placebo tablets. * This procedure was repeated once a week for 13 wk. In no case did drug administration take longer than 30 sec. Subjects fasted from 12 A.M. and were not permitted tea or coffee after 10 A.M. on study days. The attempt was made to conduct the study in double-blind fashion but in some instances the subjects recognized a characteristic taste in the NaSAM solution. The dose of NaSAM was gradually increased from 0.65 to 2.60 gm over 13 wk. (Results of the doseranging part of the study will be discussed in a subsequent report.) Only the data for the 0.65, 1.30, 1.95, and 2.60 gm dosage levels of both SAM and NaSAM will be presented in this report. At each dosage level (0.65, 1.30, 1.95, and 2.60 gm) the sequence of drug administration was randomized. Subject 6 was dropped from the study before receiving the 2.60 gm dose and therefore at this dosage a complete crossover was complete for only 5 subjects. Each subject remained on an examining table supine, at 45 0 gatch, during the experimental period. A number 19 G butterfly needle was placed in a forearm vein or the antecubital fossa and kept open with normal saline for the duration of the experimental period (about 3Yz hr). Blood samples were removed through 3-way stopcocks placed in the intravenous line. Less than a liter of normal saline was used over approximately 3Yz hr to keep the line open and to flush it after sampling. Whole blood, 10 ml, was drawn at zero min and at 2, 5, 10, 20, 30, 45, 60, 90, 120, and 180 min after the dose. After the 1.30 gm dose, blood samples were also collected 15 min after dosing. The whole blood was transferred to citrated tubes, centrifuged within 30 min of collection, and the plasma stored at -20 C until assayed. Assay. Free SAM was assayed by minor modification of a fluorometric method described by Barr and Riegelman.2, Plasma, 0.5 ml, was 0
• Sodium salicylamide and placebo preparations were supplied by Vick Divisions Research. Commercially purchased salicylamide tablets, distributed by Harvey Laboratories, Philadelphia, Lot 20001, were used. They had the most rapid disintegration time of several N.F. tablets screened for use.
Sodium salicylamide
Volume 19 Number 4
-
FREE
453
SALICYLAMIDE
--- TOTAL SALICYLAMIDE • o
TIME
0.650 gm. SALICYLAMIDE 0.650 gm. SODIUM SALICYLAMIDE Cn = G)
(min.)
Fig. 1. Mean (± SEM) plasma levels of free and total salicylamide after administration of 0.65 gm of SAM equivalents as SAM tablets and NaSAM solution.
added to 1.0 ml of 0.2 M phosphate buffer (pH 7.0) and extracted twice into 9.0 ml of ethylene dichloride. The mixture was then reextracted into 10 ml of aqueous 0.25 N sodium hydroxide solution. The fluorescence of SAM in basic solution was determined by an AmincoBowman spectrophotofluorometer at maximum activation and emission wavelengths of 332 and 418 nm, respectively. Recovery of SAM from spiked plasma samples was in excess of 95%. Total SAM was determined by hydrolyzing all metabolites to a mixture of salicylic acid and SAM.t6 To the 0.5 ml plasma samples, 0.5 ml water and 2.0 ml of 12.5% v/v sulfuric acid were added in screw-capped centrifuge tubes. Samples were autoclaved at 15 pounds pressure and 120° C for 20 min. Samples were then extracted in the same manner as described for plasma SAM above. The fluorescence of hydrolyzed SAM was determined at an emission wavelength of 400 nm using activation wavelengths of 332 and 304 nm (to include any SAM converted to salicylic acid during hydrolysis). The total SAM concentration was then determined from a standard curve by taking the sum of the two fluorescent measurements corrected with a predosing plasma bank. The standard curve was linear over the range of 0.1 to 50 mcg/m!. The data for drug concentration in plasma were analyzed for significant treatment differences by applying a 3-way analysis of vari-
ance, treating the data as if it were gathered from a split-plot design experiment. Subjective questionnaire. The subjective effects of SAM were evaluated by means of a self-scoring questionnaire similar to that described by Eade and Lasagna. tO On the first page of the questionnaire subjects were asked to discriminate between opposite pairs of subjective responses such as "sleepy" and "awake." They were asked to rate the intensity of responses by circling a number on either side of a central zero point. The subject always had his previous score before him:
o min
15 min 45 min 120 min
3 3 3 3
Sleepy 2 2 2 2
0 0 0 0
I I I I
Awake 2 2 2 2
3 3 3 3
The second page of the questionnare listed 14 side effects. These items could be scored in intensity from 0 to 3 and the subject always had his previous score before him: Lightheadedness
o min
15 min 45 min 120 min
0 0 0 0
2 2 2 2
3 3 3 3
Prior to the study, volunteers were given a set of instructions, and underwent a trial run of the questionnaire. The subjects filled out the questionnaires prior to drug administration and
454
Fleckenstein et al.
Clinical Pharmacology and Therapeutics
-
FREE
SALICYLAMIDE
TOTAL SALICYLAMIDE 1.300 9m. SALICYLAMIDE 1.300 9m. SODIUM SALICYLAMIDE
• "
Cn = 6)
TIME
(min.)
Fig. 2. Mean (± SEM) plasma levels of free and total salicylamide after administration of 1.30 gm of SAM equivalents as SAM tablets and NaSAM solution.
-
FREE
SALICYLAMIDE
--- TOTAL SALICYLAMIDE •
1.950 9m. SALICYLAMIDE
()
1.950 9m. SODIUM SALICYLAMIDE (n =6)
TIME
(min.)
Fig. 3. Mean (± SEM) plasma levels of free and total salicylamide after administration of 1.95 gm of SAM equivalents as SAM tablets and NaSAM solution.
at 15, 45, and 120 min after taking the drug. The subjects were told they would be given an aspirin-like drug. In addition, they were told that the drug might stimulate, depress, or have no effect at all. They were asked to discriminate between these responses. Each sub-
ject was tested in isolation in the examining rooms of a medical clinic. They were asked not to discuss the subjective effects with the other volunteers. Subjects were allowed to study, but could not leave the examining room. In scoring the individual responses, the net
Sodium salicylamide
Volume 19 Number 4
TIME
455
(min.)
Fig. 4. Mean (± SEM) plasma levels of free and total salicylamide after administration of 2.60 gm of SAM equivalents as SAM tablets and NaSAM solution.
score for any item was the sum of the algebraic difference between the O-hr and the 15-, 45-, and 120- min. score. The scoring system allowed for positive or negative scores for the subjective reponses, but only a positive score for responses to side effects. Results
Free SAM. Figs. 1 to 4 show the mean (± SEM) free and total plasma levels, follow-
ing 0.65,1.30, 1.95, and 2.60 gm respectively, of SAM equivalents given as liquid sodium salt or tableted SAM. The peak mean plasma concentration of nonmetabolized (free) SAM was reached earlier and was higher after NaSAM than after SAM. At all dosage levels there is a difference between the treatments due to a treatment-sampling time interaction (p < 0.01); the free SAM concentration in blood after NaSAM is higher than that after SAM tablets at early sampling times (for the IO-min. sample, p < 0.01 at 0.65, 1.30, and 1.95 gm and p < 0.05 at 2.6 gm dose levels). The mean area under the plasma concentration curve (AUC) from NaSAM is greater than from
SAM tablets at the 0.65, 1.95, and 2.60 gm dose levels (p < 0.05, paired t test). The difference in mean AVe at the 1.30 gm dosage level is not significant. In Fig. 5 it can be seen that the systemic availability of free SAM is dose-dependent after NaSAM. As the dose of NaSAM is increased from 0.65 gm to 2.65 gm, the availability of free SAM rises in a nonlinear fashion. The availability of free SAM after SAM is also dose-dependent (not shown). Mean AVe values are lower and have a larger coefficient of variation at every dosage level of SAM than of NaSAM. Total SAM. The treatment: sampling time interaction is significant (p < 0.01) at all dosage levels; the total SAM concentration in plasma after NaSAM liquid is greater than after SAM at all sampling times from 10 through 60 min at all dosage levels (p < 0.05 at 10 and 60 min and p < 0.01 at 15, 30, and 45 min sampling times). The mean AUe for total SAM achieved by the subjects after NaSAM is greater than after SAM at the 0.65 (p < 0.05, paired t test) and 2.60 gm (p < 0.01, paired
456
Fleckenstein et al.
Clinical Pharmacology and Therapeutics
Table II. Incidence and magnitude of side effects
