Pancreatic Exocrine IV. The
Enzyme
Insufficiency
Content of Commercial Pancreatic
Supplements
Frederick W. Pairent, PhD, John M. Howard, MD
Comparative in vitro analyses of 14 commercial pancreatic supplewere performed as to their protease, trypsin, trypsinogen, amylase, and lipase contents. Extreme variation was found in the individual enzyme activities of the various preparations tested. Pancrelipase (Cotazym) had at least double the activity of all enzymes tested in comparison to the next most potent preparation. Two preparations were practically void of amylase activity and had very low lipase activity. Although trypsin and trypsinogen activities were quite variable, protease activity (toward hemoglobin) was the ments
least variable factor tested.
number of
large pancreatic supple¬ A relatively preparations currently replacement therapy pancreatic enzyme
available for oral in exocrine defi¬ enzyme ciency. The selection of one of these supplements by the clinician has been largely a matter of choice, based on sup¬ plement evaluations reported in the scientific journals and on the information given by the drug manufacturers. From the information on drug strength given by the man¬ ufacturers, one might conclude that most of the available preparations are comparable in terms of enzyme content. Previous studies from this laboratory, however, do not confirm this conclusion.'-* Other studies,01* as well as our own, have tended to implicate some of the pancreatic en¬ zyme preparations as superior to others. We have analyzed a number of commercial pancreatic supplements for their proteolytic, amylolytic, and lipolytic activities. On comparison, there is a considerable variation in the enzyme activities of these preparations. The results suggest that some preparations could have little efficacy in the treatment of pancreatic insufficiency. ment
Accepted
are
publication Dec 3, 1974. Department of Biological Chemistry, Hahnemann Medical College, Philadelphia (Dr. Pairent), and the Department of Surgery, Medical College of Ohio, Toledo (Dr. Howard). Reprint requests to Department of Surgery, Medical College of Ohio, Arlington and South Detroit avenues, Toledo, OH 43614 (Dr. Howard). From the
for
MATERIALS AND METHODS Five analyses were carried out on 14 commercial pancreatic supplements. The preparations were obtained through the hospi¬ tal pharmacy at various times over a period of two years. The average tablet weight, or capsule content weight, was obtained by weighing at least 20 tablets, or the contents of at least 20 capsules, on an analytic balance. Enzyme analyses were carried out by pooling and grinding at least 20 tablets, or by pooling and mixing of the contents of at least 20 capsules. An aliquot was accurately weighed and dissolved
in 0.85% sodium chloride. Dilutions of this solution were made with appropriate buffers for each of the enzyme analyses. Protease activity was measured at pH 7.5, using hemoglobin as the substrate according to a modification of the method of Anson.10 Five milliliters of a 1.0 gm/100 ml hemoglobin substrate so¬ lution was preincubated at 37 C. One milliliter of the test solution was added. After exactly ten minutes, 10 ml of a 5% trichloroacetic acid (TCA) solution was added rapidly, with mixing. The solution was filtered after 15 minutes, and the supernatant was analyzed for TCA-soluble peptides using the Folin-Ciocalteau reagent (one-third strength). The activities are expressed as the micromols of TCA-soluble tyrosine equivalents formed per minute per tablet or capsule. Trypsin-trypsinogen analyses were carried out with a modi¬ fication of an ultramicromethod using benzoyl-arginyl-p-nitroanilide as the substrate.11 The product formed by the action of en¬ zymes having arginyl esterase activity is -nitroaniliade, which gives a yellow color in alkaline solution and has a maximum absorbence at 410 µ. Trypsin and trypsinogen analyses are performed simultane¬ ously. For trypsinogen, 10µ1 of the sample is incubated with 5µ1 of a 1 mg/ml solution of enteropeptidase in saline for 30 minutes at 37 C. For trypsin, 5µ1 of saline is added to another 10µ1 aliquot of the sample. One hundred microliters of 0.02M benzoyl-arginyl-pnitroanilide, pH 8.1, TRIS buffer is added to each tube, and incu¬ bation at 37 C is continued. After exactly 15 minutes, 200µ1 of a 0.10N sodium hydroxide solution is added to stop the reaction, the contents are rapidly mixed, and the 410- µ absorbence is read on a
spectrophotometer. The trypsin activity
Downloaded From: http://archsurg.jamanetwork.com/ by a UQ Library User on 06/15/2015
is
expressed
as
the
410- µ absorbence
per
impossible.
minute obtained with the tube where enteropeptidase was not added. Trypsinogen activity is calculated as the 410- µ absorb¬ ence per minute obtained with the tube to which enteropeptidase was added, less the absorbence per minute due to trypsin. Amylase activity was measured by the Somogyi method.12 Lipase activity was measured by the Cherry-Crandall method."
The results of this study indicate that extreme vari¬ ations can be found in the individual enzyme activities of the various preparations tested. One drug, pancrelipase (Cotazym), stands out as the most potent preparation, having double the activity of most enzymes tested in com¬ parison to the next most potent preparation. However, since more than a single tablet or capsule per dosage may be recommended by the manufacturer, several other prep¬ arations shown in the Table can also be considered quite adequate for the treatment of pancreatic insufficiency. Pancreatin (Viokase), pancreatin and pyridoxine hydrochloride (Lipan), and an enzyme mixture containing mainly pancrelipase (Accelerase) can be mentioned in this
RESULTS
Table lists the 14 supplements tested by this laboratory, along with their manufacturer. The mean tablet or cap¬ sule content weight, in milligrams, with the standard de¬ viation of a large number of weighings, is given with the five analyses shown. The enzyme activities are expressed on a "per tablet" or "per capsule" basis so that direct com¬ parison of drug strength can be made. The drug manufac¬ turer's recommendations for dosage varies for each of these products in terms of the number of tablets or cap¬ sules required per meal or per day.
regard.
Two of the preparations shown in the Table appear to be practically void of amylase activity. These same prepa¬ rations also exhibited the lowest lipase activity of all the drugs tested. With the exception of these two drugs, and an enzyme-vitamin complex (Uticon), the ratio of amylase to lipase activities varies within the narrow limits of 8 to 13. This common feature might be expected on the basis of the drugs being prepared from a common source of pan¬ creatic extract. The most variable enzyme system measured in this study was trypsin-trypsinogen. Since these units are com¬ parable, and trypsinogen is rapidly converted to trypsin after ingestion of the drug, there should be little impor¬ tance attached to the presence of trypsin activity in its ac¬ tive or precursor form. It is interesting to note, however, that all but one preparation had at least some tryptic ac¬ tivity present as trypsinogen. This fraction ranged from 5% to 98% of the total tryptic activity of the drug. Assays that include the proenzyme, therefore, are essential in evaluation of trypsin content of a pancreatic supplement.
COMMENT In order to select a good pancreatic enzyme supplement preparation for clinical use, we have analyzed 14 commer¬ cial preparations for their enzyme contents. The stated potencies of these drugs given by the drug manufacturers has, in our experience, proved to be of little value in mak¬ ing the selection. Since the enzyme activities of the crude preparations used in the formulation of pancreatic supple¬ ments (eg, pancreatin) have not been adequately stan¬ dardized, little knowledge is gained in knowing the weight of these preparations incorporated into a tablet or capsule. Even though many manufacturers actually mea¬ sure the enzyme content of their finished product, each company, because of the complexities inherent in enzyme
measurements, has selected different methods, thus mak¬
ing comparative evaluation of their labeling of the drugs
Enzyme Analyses of Pancreatic Supplements Tablet
Drug Accelerase Convertin
Cotazym Dactilase
Enzypan Festal Kanulase
KU-zyme Lipan pancreatin Panteric
Pro-gestive Uticon Viokase
Manufacturer
Form
Organon Ine
Capsule
BF Ascher & Co Ine Organon Ine Lakeside Laboratories Ine Norgine Laboratories Ine Hoechst Pharmaceuticals Ine
Tablet
Dorsey Laboratories Kremers-Urban Co Spirt & Co Ine Eli Lilly & Co Parke Davis & Co Nutrition Control Products Layton Laboratories VioBin Corp
Capsule Tablet Tablet Tablet Tablet
Capsule Tablet Tablet Tablet Tablet Tablet Tablet
372 777 487 583 403 578 801 554 540 352 769 339 872 397
Activity per Tablet
or
Capsule Weight, mg ± 20.0 ± 46.4 ± 30.0 ± 14.1 ± 10.9
28.2 ±6.0 ± 8.9 ± 30.2 ±8.4 ± 19.4 ±
± 10.1 ± 20.0
± 7.5
Protease* 70 18 214 39 179 36 19 40 120 26 85 46 70 112
or
Capsule
Trypsinf Trypsinogen^ Amylase§
Lipase'1
10,300 2,210 63,800 17,000 4,630
2,530 1,090 14,300
19,850
2,600 4,630
289 96 11
16,600
60,470 5,810 6,640 15,920 10
36 94
7,400 3,560 2,460
14,200 2,440
887 32 349 324 197
1,770 2,440 3,140
78
1,055 27,510 5,140 17,190 15,900 14,720 25,590
501 830
1,270
2,380 507
1,620 1,220 680
2,230
Micromols of (5%) TCA-soluble tyrosine equivalents per minute, pH 7.5, 37 C (hemoglobin substrate), t Increase in 410- µ absorbence per 15 minutes, pH 8.1, 37 C (benzoly-arginyl-p-nitroanilide substrate). Additional increase in 410- µ absorbence per 15 minutes, pH 8.1, 37 C (benzoyl-arginyl-p-nitroanilide substrate) after 30 minutes, 37 C incubation of sample with enteropeptidase. § Grams of glucose equivalents per 30 minutes, pH 7.2, 37 C (Somogyi units X 1,000). (] Milliliters of 0.050N sodium hydroxide required to neutralize acid formed during 18-hour incubation 37 C (Cherry-Crandall method13). *
Downloaded From: http://archsurg.jamanetwork.com/ by a UQ Library User on 06/15/2015
In distinction to the trypsin and trypsinogen assays, the least variable factor measured was the protease activity of these preparations. There is no correlation between the proteolytic activity toward hemoglobin at pH 7.5 and the trypsin activity of these drugs. This might be an indica¬ tion that chymotrypsin activity of pancreatic supplements must contribute substantially to the digestion of proteins.
These and earlier studies suggest that there may be a substantial variation in enzyme content between various lots of a given product.
This 02723.
investigation
was
supported by
Public Health Service grant AM-
References 1. Giulian BB, Singh LM, Mansfield AO, et al: The treatment of pancreatic exocrine insufficiency: I. In vitro lipolytic activities of pancreatic lipase and 15 commercial pancreatic supplements. Ann
Surg 165:564-570, 1967. 2. Giulan BB, Mitsuoka H, Mansfield AO, et al: The treatment of pancreatic exocrine insufficiency: II. Effects on fat absorption of pancreatic lipase and 15 commercial pancreatic supplements as measured by I131 tagged triolein in the dog. Ann Surg 165:571-579, 1967. 3. Trapnell
JE, Pairent FW, Giulan BB: Techniques for evaluating digestive enzymes, in Siegler PI, Moyer JH III (eds): Pharmacological Techniques in Drug Evaluation. Chicago, Year Book Medical Publishers Inc, 1967, vol 2, p 675. 4. Pairent FW, Trapnell JE, Howard JM: The treatment of pancreatic exocrine insufficiency: III. The effects of pancreatic ductal ligation and oral pancreatic enzyme supplements on fecal excretion in the dog. Ann Surg 170:737-746, 1969. lipid 5. Beck K: The clinical efficacy of some preparations of pancreatic enzymes. Ger Med Mon 8:47-51, 1963. 6. Marks IN, Bank S, Airth EM: Pancreatic replacement ther-
apy in the treatment of pancreatic steatorrhea. Gut 4:217-223, 1963. 7. Jordan PH, Grossman MI: The effect of dosage schedule on the efficacy of substitution therapy. Gastroenterology 36:447-451, 1959. 8. Polocheck AA, Willard RF: Pancreatin as a differential absorption test in patients with steatorrhea. Ann Intern Med 52:1195-1200, 1960. 9. Conte M, Fouet P: Critical study of digestive enzyme substi-
tution therapy. Gastroenterologia 105:30-42, 1966. 10. Anson ML: The estimation of pepsin, trypsin, papain and cathepsin with hemoglobin. J Gen Physiol 22:79-89, 1939. 11. Erlanger BF, Kokowsky N, Cohen W: The preparation and properties of two new chromogenic substrates of trypsin. Arch Biochem Biophys 95:271-278, 1961. 12. Somogyi M: Micromethods for the estimation of diastase. J Biol Chem 125:339-414, 1938. 13. Cherry IS, Crandall LA Jr: The specificity of pancreatic lipase: Its appearance in the blood after pancreatic injury. Am J Physiol 100:266-273, 1932.
Downloaded From: http://archsurg.jamanetwork.com/ by a UQ Library User on 06/15/2015
Enzyme
Insufficiency
Content of Commercial Pancreatic
Supplements
Frederick W. Pairent, PhD, John M. Howard, MD
Comparative in vitro analyses of 14 commercial pancreatic supplewere performed as to their protease, trypsin, trypsinogen, amylase, and lipase contents. Extreme variation was found in the individual enzyme activities of the various preparations tested. Pancrelipase (Cotazym) had at least double the activity of all enzymes tested in comparison to the next most potent preparation. Two preparations were practically void of amylase activity and had very low lipase activity. Although trypsin and trypsinogen activities were quite variable, protease activity (toward hemoglobin) was the ments
least variable factor tested.
number of
large pancreatic supple¬ A relatively preparations currently replacement therapy pancreatic enzyme
available for oral in exocrine defi¬ enzyme ciency. The selection of one of these supplements by the clinician has been largely a matter of choice, based on sup¬ plement evaluations reported in the scientific journals and on the information given by the drug manufacturers. From the information on drug strength given by the man¬ ufacturers, one might conclude that most of the available preparations are comparable in terms of enzyme content. Previous studies from this laboratory, however, do not confirm this conclusion.'-* Other studies,01* as well as our own, have tended to implicate some of the pancreatic en¬ zyme preparations as superior to others. We have analyzed a number of commercial pancreatic supplements for their proteolytic, amylolytic, and lipolytic activities. On comparison, there is a considerable variation in the enzyme activities of these preparations. The results suggest that some preparations could have little efficacy in the treatment of pancreatic insufficiency. ment
Accepted
are
publication Dec 3, 1974. Department of Biological Chemistry, Hahnemann Medical College, Philadelphia (Dr. Pairent), and the Department of Surgery, Medical College of Ohio, Toledo (Dr. Howard). Reprint requests to Department of Surgery, Medical College of Ohio, Arlington and South Detroit avenues, Toledo, OH 43614 (Dr. Howard). From the
for
MATERIALS AND METHODS Five analyses were carried out on 14 commercial pancreatic supplements. The preparations were obtained through the hospi¬ tal pharmacy at various times over a period of two years. The average tablet weight, or capsule content weight, was obtained by weighing at least 20 tablets, or the contents of at least 20 capsules, on an analytic balance. Enzyme analyses were carried out by pooling and grinding at least 20 tablets, or by pooling and mixing of the contents of at least 20 capsules. An aliquot was accurately weighed and dissolved
in 0.85% sodium chloride. Dilutions of this solution were made with appropriate buffers for each of the enzyme analyses. Protease activity was measured at pH 7.5, using hemoglobin as the substrate according to a modification of the method of Anson.10 Five milliliters of a 1.0 gm/100 ml hemoglobin substrate so¬ lution was preincubated at 37 C. One milliliter of the test solution was added. After exactly ten minutes, 10 ml of a 5% trichloroacetic acid (TCA) solution was added rapidly, with mixing. The solution was filtered after 15 minutes, and the supernatant was analyzed for TCA-soluble peptides using the Folin-Ciocalteau reagent (one-third strength). The activities are expressed as the micromols of TCA-soluble tyrosine equivalents formed per minute per tablet or capsule. Trypsin-trypsinogen analyses were carried out with a modi¬ fication of an ultramicromethod using benzoyl-arginyl-p-nitroanilide as the substrate.11 The product formed by the action of en¬ zymes having arginyl esterase activity is -nitroaniliade, which gives a yellow color in alkaline solution and has a maximum absorbence at 410 µ. Trypsin and trypsinogen analyses are performed simultane¬ ously. For trypsinogen, 10µ1 of the sample is incubated with 5µ1 of a 1 mg/ml solution of enteropeptidase in saline for 30 minutes at 37 C. For trypsin, 5µ1 of saline is added to another 10µ1 aliquot of the sample. One hundred microliters of 0.02M benzoyl-arginyl-pnitroanilide, pH 8.1, TRIS buffer is added to each tube, and incu¬ bation at 37 C is continued. After exactly 15 minutes, 200µ1 of a 0.10N sodium hydroxide solution is added to stop the reaction, the contents are rapidly mixed, and the 410- µ absorbence is read on a
spectrophotometer. The trypsin activity
Downloaded From: http://archsurg.jamanetwork.com/ by a UQ Library User on 06/15/2015
is
expressed
as
the
410- µ absorbence
per
impossible.
minute obtained with the tube where enteropeptidase was not added. Trypsinogen activity is calculated as the 410- µ absorb¬ ence per minute obtained with the tube to which enteropeptidase was added, less the absorbence per minute due to trypsin. Amylase activity was measured by the Somogyi method.12 Lipase activity was measured by the Cherry-Crandall method."
The results of this study indicate that extreme vari¬ ations can be found in the individual enzyme activities of the various preparations tested. One drug, pancrelipase (Cotazym), stands out as the most potent preparation, having double the activity of most enzymes tested in com¬ parison to the next most potent preparation. However, since more than a single tablet or capsule per dosage may be recommended by the manufacturer, several other prep¬ arations shown in the Table can also be considered quite adequate for the treatment of pancreatic insufficiency. Pancreatin (Viokase), pancreatin and pyridoxine hydrochloride (Lipan), and an enzyme mixture containing mainly pancrelipase (Accelerase) can be mentioned in this
RESULTS
Table lists the 14 supplements tested by this laboratory, along with their manufacturer. The mean tablet or cap¬ sule content weight, in milligrams, with the standard de¬ viation of a large number of weighings, is given with the five analyses shown. The enzyme activities are expressed on a "per tablet" or "per capsule" basis so that direct com¬ parison of drug strength can be made. The drug manufac¬ turer's recommendations for dosage varies for each of these products in terms of the number of tablets or cap¬ sules required per meal or per day.
regard.
Two of the preparations shown in the Table appear to be practically void of amylase activity. These same prepa¬ rations also exhibited the lowest lipase activity of all the drugs tested. With the exception of these two drugs, and an enzyme-vitamin complex (Uticon), the ratio of amylase to lipase activities varies within the narrow limits of 8 to 13. This common feature might be expected on the basis of the drugs being prepared from a common source of pan¬ creatic extract. The most variable enzyme system measured in this study was trypsin-trypsinogen. Since these units are com¬ parable, and trypsinogen is rapidly converted to trypsin after ingestion of the drug, there should be little impor¬ tance attached to the presence of trypsin activity in its ac¬ tive or precursor form. It is interesting to note, however, that all but one preparation had at least some tryptic ac¬ tivity present as trypsinogen. This fraction ranged from 5% to 98% of the total tryptic activity of the drug. Assays that include the proenzyme, therefore, are essential in evaluation of trypsin content of a pancreatic supplement.
COMMENT In order to select a good pancreatic enzyme supplement preparation for clinical use, we have analyzed 14 commer¬ cial preparations for their enzyme contents. The stated potencies of these drugs given by the drug manufacturers has, in our experience, proved to be of little value in mak¬ ing the selection. Since the enzyme activities of the crude preparations used in the formulation of pancreatic supple¬ ments (eg, pancreatin) have not been adequately stan¬ dardized, little knowledge is gained in knowing the weight of these preparations incorporated into a tablet or capsule. Even though many manufacturers actually mea¬ sure the enzyme content of their finished product, each company, because of the complexities inherent in enzyme
measurements, has selected different methods, thus mak¬
ing comparative evaluation of their labeling of the drugs
Enzyme Analyses of Pancreatic Supplements Tablet
Drug Accelerase Convertin
Cotazym Dactilase
Enzypan Festal Kanulase
KU-zyme Lipan pancreatin Panteric
Pro-gestive Uticon Viokase
Manufacturer
Form
Organon Ine
Capsule
BF Ascher & Co Ine Organon Ine Lakeside Laboratories Ine Norgine Laboratories Ine Hoechst Pharmaceuticals Ine
Tablet
Dorsey Laboratories Kremers-Urban Co Spirt & Co Ine Eli Lilly & Co Parke Davis & Co Nutrition Control Products Layton Laboratories VioBin Corp
Capsule Tablet Tablet Tablet Tablet
Capsule Tablet Tablet Tablet Tablet Tablet Tablet
372 777 487 583 403 578 801 554 540 352 769 339 872 397
Activity per Tablet
or
Capsule Weight, mg ± 20.0 ± 46.4 ± 30.0 ± 14.1 ± 10.9
28.2 ±6.0 ± 8.9 ± 30.2 ±8.4 ± 19.4 ±
± 10.1 ± 20.0
± 7.5
Protease* 70 18 214 39 179 36 19 40 120 26 85 46 70 112
or
Capsule
Trypsinf Trypsinogen^ Amylase§
Lipase'1
10,300 2,210 63,800 17,000 4,630
2,530 1,090 14,300
19,850
2,600 4,630
289 96 11
16,600
60,470 5,810 6,640 15,920 10
36 94
7,400 3,560 2,460
14,200 2,440
887 32 349 324 197
1,770 2,440 3,140
78
1,055 27,510 5,140 17,190 15,900 14,720 25,590
501 830
1,270
2,380 507
1,620 1,220 680
2,230
Micromols of (5%) TCA-soluble tyrosine equivalents per minute, pH 7.5, 37 C (hemoglobin substrate), t Increase in 410- µ absorbence per 15 minutes, pH 8.1, 37 C (benzoly-arginyl-p-nitroanilide substrate). Additional increase in 410- µ absorbence per 15 minutes, pH 8.1, 37 C (benzoyl-arginyl-p-nitroanilide substrate) after 30 minutes, 37 C incubation of sample with enteropeptidase. § Grams of glucose equivalents per 30 minutes, pH 7.2, 37 C (Somogyi units X 1,000). (] Milliliters of 0.050N sodium hydroxide required to neutralize acid formed during 18-hour incubation 37 C (Cherry-Crandall method13). *
Downloaded From: http://archsurg.jamanetwork.com/ by a UQ Library User on 06/15/2015
In distinction to the trypsin and trypsinogen assays, the least variable factor measured was the protease activity of these preparations. There is no correlation between the proteolytic activity toward hemoglobin at pH 7.5 and the trypsin activity of these drugs. This might be an indica¬ tion that chymotrypsin activity of pancreatic supplements must contribute substantially to the digestion of proteins.
These and earlier studies suggest that there may be a substantial variation in enzyme content between various lots of a given product.
This 02723.
investigation
was
supported by
Public Health Service grant AM-
References 1. Giulian BB, Singh LM, Mansfield AO, et al: The treatment of pancreatic exocrine insufficiency: I. In vitro lipolytic activities of pancreatic lipase and 15 commercial pancreatic supplements. Ann
Surg 165:564-570, 1967. 2. Giulan BB, Mitsuoka H, Mansfield AO, et al: The treatment of pancreatic exocrine insufficiency: II. Effects on fat absorption of pancreatic lipase and 15 commercial pancreatic supplements as measured by I131 tagged triolein in the dog. Ann Surg 165:571-579, 1967. 3. Trapnell
JE, Pairent FW, Giulan BB: Techniques for evaluating digestive enzymes, in Siegler PI, Moyer JH III (eds): Pharmacological Techniques in Drug Evaluation. Chicago, Year Book Medical Publishers Inc, 1967, vol 2, p 675. 4. Pairent FW, Trapnell JE, Howard JM: The treatment of pancreatic exocrine insufficiency: III. The effects of pancreatic ductal ligation and oral pancreatic enzyme supplements on fecal excretion in the dog. Ann Surg 170:737-746, 1969. lipid 5. Beck K: The clinical efficacy of some preparations of pancreatic enzymes. Ger Med Mon 8:47-51, 1963. 6. Marks IN, Bank S, Airth EM: Pancreatic replacement ther-
apy in the treatment of pancreatic steatorrhea. Gut 4:217-223, 1963. 7. Jordan PH, Grossman MI: The effect of dosage schedule on the efficacy of substitution therapy. Gastroenterology 36:447-451, 1959. 8. Polocheck AA, Willard RF: Pancreatin as a differential absorption test in patients with steatorrhea. Ann Intern Med 52:1195-1200, 1960. 9. Conte M, Fouet P: Critical study of digestive enzyme substi-
tution therapy. Gastroenterologia 105:30-42, 1966. 10. Anson ML: The estimation of pepsin, trypsin, papain and cathepsin with hemoglobin. J Gen Physiol 22:79-89, 1939. 11. Erlanger BF, Kokowsky N, Cohen W: The preparation and properties of two new chromogenic substrates of trypsin. Arch Biochem Biophys 95:271-278, 1961. 12. Somogyi M: Micromethods for the estimation of diastase. J Biol Chem 125:339-414, 1938. 13. Cherry IS, Crandall LA Jr: The specificity of pancreatic lipase: Its appearance in the blood after pancreatic injury. Am J Physiol 100:266-273, 1932.
Downloaded From: http://archsurg.jamanetwork.com/ by a UQ Library User on 06/15/2015
