Isoamylase Pattern Research Articles

Unusual Anodic Migrating Isoamylase Differentiates Selected Malignant from Nonmalignant Ascites

Ascitic fluid from nine patients with nonmalignant ascites and nine patients with malignant ascites was subjected to isoamylase analysis. Unusual isoamylase bands that migrate to the anode were demonstrated in seven of eight patients with ovarian carcinoma and in one patient with gastric carcinoma. In no case of nonmalignant ascites was anodic isoamylase found, despite the presence of normal amylase in all samples. This is the first report of anodic isoamylase from ascitic fluid in gastric carcinoma and the first series comparing isoamylase patterns in malignant and nonmalignant ascites. Anodic isoamylase on electrophoresis of ascitic fluid may prove to be useful as a tumor marker in differentiating selected malignancies from nonmalignant ascites.

Amniotic fluid isoamylase activity in uneventful pregnancies.

A kinetic test (Phadebas) was used to determine the isoamylase activity in 50 serum specimens and 159 samples of amniotic fluid. A highly significant difference between the isoamylase patterns of serum and amniotic fluid was ascertained which strongly supports the view that amylase activity in amniotic fluid is not of maternal origin. During the whole course of gestation the activity of pancreatic isoamylase was constantly low whereas there was an increase of nonpancreatic activity (S-type amylase) from 44 +/- 10 U/l in the 18th week of gestation to 445 +/- 170 U/l in week 39/40. A comparison between the dispersion of the ascertained values and the lecithin-sphingomyelin (L/S) ratio showed a clear overlapping. There was a correlation coefficient of r = 0.645 for 115 amniotic fluid specimens examined in our investigation. The findings show that the estimation of nonpancreatic isoamylase (S-type amylase) activity in amniotic fluid is a valuable indirect parameter for determining fetal maturity and fetal lung maturity. The easy and quick determination method will provide even those hospitals with an index of fetal maturity which are presently not in a position to estimate the L/S ratio, thus being of great assistance in the field of premature deliveries.

Puzzling persistent hyperamylasemia, probably neitherpancreatic nor pathologic

Increased serum amylase levels most commonly signify pancreatic disease. One hundred seventeen consecutive patients were studied because their serum amylase levels were abnormally high for periods ranging from 3 to 48 weeks. In each case, extensive clinical and radiologic evaluation had failed to reveal a reason for the abnormality. The amylase isoenzymes of their sera were separated by polyacrylamide gel electrophoresis, and the fractions were measured by a saccharogenic assay. The findings in the 117 patients showed that 79 percent had non-pancreatic causes for their hyperamylasemia. The biggest single group (64 percent) had a normal distribution of isoamylases, albeit at unusually high concentrations. This phenomenon, which has not been defined previously, is probably a variant of normal in which the homeostatic balance between production and metabolism is set at a high level. Macroamylasemia accounted for 6 percent of the cases and salivary hyperamylasemia for only 9 percent. Three patients had the characteristic isoamylase pattern ("old amylase") associated with pancreatic pseudocysts. Isoamylase fractionation is a cheap, efficient, and effective means of ruling out a pancreatic cause for hyperamylasemia. It is probable that in the majority of cases of persistent hyperamylasemia without obvious clinical cause there will be no disease at all.

Electrophoretic analysis of amylase isozymes in geographical strains of Sitophilus oryzae (L.), S. zeamais Motsch., and S. granarius (L.) (Coleoptera: Curculionidae)

Electrophoretic analysis of adult intestinal tracts indicated that two strains of Sitophilus oryzae (L.) (Japan and Tanzania-90) lacked the major α-amylase isozyme present in 12 other geographical strains. Amylase zymograms of adults of 17 strains of Sitophilus zeamais Motschulsky were not only different from those of S. oryzae but showed significant between-strain variation. Strains of S. zeamais could be arranged into approximately four groups based on isoamylase patterns. There was no apparent correlation between geographical origin or culture diet and isoamylase pattern within these groups. One strain of Sitophilus granarius (L.) (3SG) had an additional minor isozyme not present in five other strains of this species. Although the biological or functional significance of the wide strain to strain variation in isoamylase patterns of S. zeamais has yet to be determined, the qualitative and quantitative differences in amylases between these weevil species indicate these enzymes may be useful chemotaxonomic markers. In addition to differences in amylase composition, there was significant strain variation in weights of wheat-fed weevils from each species.

Isoamylases and Their Thermolability in Serum and Cyst Fluid from Patients with Pancreatic Pseudocysts

The isoamylase pattern in serum and the amylase thermolability have been suggested as screening tests for the development of pancreatic pseudocysts. To study whether serum reflects the contents of pseudocysts, we have investigated the isoamylases and their thermolability in cyst fluid and in serum from 13 patients with pancreatic pseudocysts. No significant correlation was found between the contents in serum and cyst fluid with regard to total amylase and isoamylase P2 and P3 or with regard to the thermolability of total amylase and isoamylase P2 and P3. Thus, serum does not reflect the cyst contents of isoamylases or their thermolability. Therefore these serum amylase determinations can hardly be expected to be useful in distinguishing patients developing pseudocysts among patients with pancreatitis.

Action pattern of serum amylase using p-nitrophenyl-maltoheptaoside as substrate.

Selected serum samples with different isoamylase patterns, as revealed by agarose gel electrophoresis, were analysed by the Phadebas amylase test, and by using p-nitrophenyl-maltoheptaoside as the substrate. The products with the latter substrate were examined by high performance liquid chromatography (HPLC). For some samples, different catalytic activities were registered by the two amylase methods. In these cases, electrophoresis showed a double pancreatic band associated with increased pancreatic amylase catalytic activity, or an abnormal, broad salivary band associated with increased salivary enzyme catalytic activity. The stoichiometric factors for most of these samples, calculated from the amount of p-nitrophenol detected after HPLC, differed from the mean values.

Foregut mucosal defects: An etiology of hyperamylasemia

To evaluate a preliminary correlation of hyperamylasemia to upper gastrointestinal bleeding, total serum amylase and serum isoamylase profiles were determined in 50 patients with upper gastrointestinal bleeding. Etiologies of the bleeding were determined in 46 patients including gastritis or duodenitis in 25, gastric ulcers in 12, duodenal ulcers in 3, Mallory-Weiss tears in 3, gastric carcinoma in 2, and esophageal varices in 1. Gastritis or duodenitis was seen incidentally in 14 more patients. Hyperamylasemia was seen in 38 patients, most commonly being due to a rise of both nonpancreatic and pancreatic isoamylases (18 patients). In 13 patients it was due to an elevation of nonpancreatic amylase alone, and in 7 patients secondary to elevated pancreatic isoamylase alone. Acute pancreatitis raises only the pancreatic component and cannot explain the hyperamylasemia in most of these patients. Hyperamylasemia did not correlate to etiology of the bleeding; gastritis or duodenitis present in the majority of these patients appears to be the unifying factor. Since both nonpancreatic and pancreatic amylases are present in the duodenum and the stomach with pyloric reflux, reabsorption of intraluminal amylase across damaged mucosa is postulated as a mechanism to explain the observed isoamylase patterns. The possibility of decreased amylase clearance as an explanation is unlikely. An alternative central nervous system mechanism might be invoked. It is concluded that hyperamylasemia is a complex event which the use of isoamylase analysis is beginning to elucidate. The hyperamylasemia seen commonly in patients presenting with upper gastrointestinal bleeding does not imply the presence of acute pancreatitis.

A correlation between clinical pancreatitis and isoenzyme patterns of amylase : Weaver DW, Bouwman DL, Walt AJ, Clink D, Resto A, Stephany J. Surgery 1982; 84:576–580

Fifty-seven patients admitted with the clinical diagnosis of acute pancreatitis had isoamylase analysis on their sera to determine the source of their hyperamylasemia. Our objective was to correlate the isoamylase pattern with our clinical observations. Thirty-nine of 57 patients (68%) had pancreatic hyperamylasemia as expected, but 18 of 57 patients (32%) had normal levels of pancreatic amylase. The hyperamylasemia in the latter group was due either to nonpancreatic hyperamylasemia (16 of 57) of macroamylasemia (2 of 57). Consequently, hyperamylasemia associated with abdominal pain, nausea, and vomiting led to the incorrect diagnosis of acute pancreatitis in 32% of the patients. The measurement of isoamylase profiles can be done rapidly and inexpensively. Knowledge that hyperamylasemia is nonpancreatic in origin may have an important influence on treatment, hospitalization, and the extent of laboratory and radiologic investigation.

P-65) - Isoamylase pattern in serum and urine of human, dog, rabbit and rat with special reference to renal isoamylase clearance

P-65) - Isoamylase pattern in serum and urine of human, dog, rabbit and rat with special reference to renal isoamylase clearance

Pancreatitis-like isoamylase pattern in normal persons

On the assumption that a rise in the pancreatic type isoamylases may not necessarily indicate underlying pancreatitis, genetic studies of human serum and urinary amylase isoenzymes have been performed with the use of electrophoresis. Although the preponderant increase in the two principal pancreatic isoamylases Amylase-1 and 2 has been accepted to be a specific index of pancreatic involvement, 1.68% of normal persons had Amylase-2 with an elevated amylase activity (named “Dominant Amylase-2”) up to the same levels as the major isoenzymes. Results of pancreozymin-secretin test and other laboratory findings of these persons with Dominant Amylase-2 were all within normal ranges. Pedigree studies confirmed an autosomal dominant mode of inheritance for this variant. The importance of serial determination and pedigree investigations has been shown to distinguish normal persons having Dominant Amylase-2 from patients with pancreatitis without elevated amylase activity. The existence of an inherited trait of pancreatitis-like isoamylase pattern in healthy individuals must be borne in mind before coming to a conclusion when amylase isoenzymes are used for clinical medicine, though preponderance of the pancreatic type isoenzymes in serum and urine has been revealed to be a characteristic finding in pancreatitis. Knowledge of amylase genetic polymorphism provides a scientific basis for amylase isoenzyme interpretation.

Clinical evaluation of the pancreatitis-like isoamylase pattern in normal persons.

Amylase isoenzyme analysis of serum and urine has been performed in 4001 normal persons and 500 patients with various disease using electrophoresis on thin layer polyacrylamide gel. Although elevation of amylase activity in amylase-1 and 2 has been reported to be the specific findings in patients with pancreatitis, 1.69% of normal persons had an elevated Amylase-2(named "Dominant Amylase-2") up to the same levels as major isoenzymes (Amylase-1 and 3), along with Amylase-1. Pedigree study confirmed an autosomal dominant mode of inheritance for Dominant Amylase-2. Knowledge of the genetic polymorphism is of importance in clinical assessment of amylase isoenzymes in patients having an elevated Amylase-2 suggestive of pancreatitis. Predominance of the pancreatic components in serum and urine has been revealed to be a specific index of pancreatic involvement. However, the existecne of an inherited trait of pancreatitis-like isoamylase pattern in healthy individuals must be borne in mind. On the basis of the present study, it may be concluded that a rise in the pancreatic type isoenzymes may not necessarily indicate underlying pancreatitis, especially in the absence of elevated amylase and lipase levels.

The nature and mode of the changes in the electrophoretic mobility of amylase isozymes

Amylase isozyme analysis of serum, urine and other body fluids has been used in clinical diagnosis, in that identification of specific isozymes may permit identification of diseased organs. Contrary to what may be expected, the occurrence of multiforms of α-amylase and the changes in the electrophoretic mobility of amylase isozyme in serum and urine have been found. The present investigation was undertaken in an attempt to make clear the nature and the mode of the changes in the electrophoretic mobility of amylase isozyme, using electrophoresis on a thin layer polyacrylamide gel.Serum and urinary amylase isozymes of normal adult persons can be separated into two major isozymes (band-1 and -3) with high amylase activity and two or three minor ones with low amylase activity. Band-1 has the same electrophoretic mobility as the pancreatic isoamylase and band-3 is the same as the salivary isoamylase. The proportions of amylase activity of the individual isozymes can be altered, however, by prolonged storage in the cold or incubation at 37°C. The relative amylase activity of band-1 and -3 reduced and that of band-2 and -5 increased. After 5 to 7 days at 37°C, new bands of higher anodic mobilities come into being, the amylase activity of which increases in the course of time Thus, band-2, -4, -6 and -8 have appeared and increased in activity at the expense of band-1. Band-5, -7 and -9, not visible in the same amount of fresh sample have become to be seen at the expense of band-3.Neither bacterial contamination during prolonged storage, nor the addition of protease and trypsin had any effect on the changes of the banding pattern. Therefore, these factors could not be implicated as a cause of the changes in isozyme proportionality. On the other hand, trypsin-inhibitor and trasylol (Kallikrein-trypsin inhibitor) could not inhibit the transformation of isozymes into more anionic forms.Storage at alkaline pH and high temperature accelerated quantitative changes in the isozyme pattern. Addition of 0.1 mole CaCl2 and the storage below -18°C could retard the transformation of isoamylases.Column chromatographic investigation revealed that the molecular weights had not been significantly altered during storage or incubation at 37°C.These observations showed the difficulty of the interpretation of the isoamylase pattern, because the number of bands, as well as their relative amylase activities, may vary if the amylase activity, reaction time and conditions of the sample are not taken into account. Thus, artifacts formed during collection, storage or electrophoretic procedures and possible hereditary variants must be considered before coming to a conclusion of the pathological variants of the isoamylase pattern.

Serum isoamylase pattern in obstructive pancreatic disease.

On agarose gel electrophoresis the pancreatic serum isoamylases can be separated into a main fraction and more anodally migrating minor fractions. In acute pancreatic hyperamylasemia as in normal subjects the activity of the anodally migrating pancreatic amylase does not exceed 20% of the activity of the main fraction. In patients with pancreatic main duct occlusion or stenosis with or without calculi or cysts, the anodally migrating pancreatic amylase increases proportionally more than does the main fraction, and can even be stronger than the main fraction. The majority of these patients also had abnormally increased total pancreatic serum amylase activity.

Relation of electrophoretic pattern of amylase isoenzymes to severity of pancreatic disease

Serum and urinary amylase isoenzymes were studied in an attempt to evaluate the usefulness of isoamylase analysis in the diagnosis of pancreatic diseases. The amylase in human serum and urine was separated into 4 to 5 distinct isoenzymes, which have mobilities consistent with those of the pancreatic isoamylases (Amylase-1, -2, -4 and -6) and the salivary isoamylases (Amylase-3, -5 and -7). The normal isoamylase pattern was Amylase-1, -2, -3 and -5. Amylase-4 and -6 could not be found in normal serum and urine. Amylase activity in Amylase-1 and/or -2 increased remarkably, having the normal isoamylase pattern, in the patients with elevated serum amylase activity after stimulation with pancreozymin-secretin or after endoscopie retrograde cholangiopancreatography. However, in the patients with acute pancreatitis, salivary type isoamylases disappeared and only the pancreatic type components were identified. It seems that the appearance of Amylase-4 and disappearance of the salivary components indicate severe pancreatic inflammation and that the apparent specificity of this isoenzyme pattern affords a mean for conforming the diagnosis of acute pancreatitis. However, the occurrence of an inherited trait in the pancreatic isoamylases of healthy individuals suggests that a rise in the pancreatic components may not necessarily indicate pancreatic disorders.

Amylase in the lung

Although elevated amylase levels in serum, pleural fluid, and extracts of tumor tissue in primary lung cancer have been reported, electrophoretic and column-chromatographic studies have not revealed the ectopic production of amylase but have merely shown an increase of amylase activity of chiefly the salivary type in these materials. The present study was designed to make clear the nature of the amylase or amylase-like substance in the serum, pleural fluid and tumor extracts, and to determine whether amylase might be produced ectopically in tumor tissues. Our data not only confirmed that the hyperamylasemia in some cases of primary lung cancer was due to an increase in salivary type isoamylases, but also showed that the same isoamylase pattern occurs in serum, pleural fluids, and diseased lung tissue of patients with pneumonia. However, the elution pattern of amylase in these materials in column-chromatography on Sephadex G-75 Superfine was different from that of salivary amylase. On the basis of our observations, it seems reasonable to conclude that the salivary type hyperamylasemia in some cases of primary lung cancer may be due to an increase in the amylase contained in normal lung tissues, resulting from activation and release into the blood stream by some inflammatory process. However, ectopic production of amylase was demonstrated in one particular case of primary lung cancer in which a high amylase content and a peculiar isoamylase were found both in the primary and metastatic lesions.

A Cause of Hyperamylasemia Associated with Chronic Liver Disease

The cause of hyperamylasemia associated with chronic liver disease is unclear. In an attempt to identify the tissue of origin of hyperamylasemia in 3 patients with chornic active hepatitis their serum was isoelectrically focused. The isoamylase patterns obtained were compared to those of pancreatic and salivary amylase. The apparent salivary gland origin of the excessive blood amylase in the patients studied was substantiated by radiological demonstration of parotid sialoectasia in one patient and histological evidence of sialoadenitis in another. Further evidence was the coincident isoelectric points of the predominant isoamylase in the sera of the liver disease patients and of patients with parotid inflammatory disease. Hyperamylasemia associated with chronic liver disease may be of salivary gland origin and as such forms part of the spectrum of extrahepatic manifestations of chronic active hepatitis.

The characteristics of amylase activity and the isoamylase pattern in serum and urine of infants and children

Determination of amylase activity and isoamylase patterns were performed in serum and urine of normal newborns, infants and children of different ages. In the serum of newborn infants measurable amounts of amylase were present. The activity increased with the age and reached the normal adult level by approximately 8 months of age. Isoamylase analysis revealed that the low level of serum amylase in infants was mainly due to deficiency of the pancreatic-type isoamylase. The absence of the pancreatic isoamylase in newborns and young infants is a physiological and developmental phenomenon. Great caution is therefore necessary when amylase isoenzymes are used in the diagnosis of abnormal pancreatic function and such results have always to be interpreted in relation to the age of the child.

Studies on serum amylase in normal man and in acute pancreatitis.

Amylase isoenzymes in serum, urine, saliva, jejunal juice, and pancreatic tissue were separated by isoelectric focusing. Isoamylase patterns obtained indicated that the majority of amylase activity in normal serum is of salivary gland origin. Pancreatic amylase is characteristically predominant in acute pancreatitis. The increased renal clearance of amylase in acute pancreatitis may be partly due to the increased proportion of the smaller molecular weight pancreatic amylase. However, a demonstrated increase in the renal clearance of salivary amylase in acute pancreatitis suggests a renal cause also. Autopsy pancreas samples devoid of TAME (p-tosyl arginine methyl ester) esterase activity (e.g. trypsin and plasma enzymes such as thrombin and plasmin) had isoenzyme patterns different to those samples with free proteolytic activity. Incubation of TAME esterase free pancreas with trypsin caused conversion of the former isoamylase pattern to one with the predominant isoenzymes focusing coincident with the predominant peak in serum from acute pancreatitis, jejunal aspirate, and TAME esterase positive autopsy pancreas. Such conversion suggests that pancreatic amylase is synthesized in a form different from that found in the intestinal lumen and serum.

Serum amylase isozyme changes in chronic pancreatitis and their clinical significance.

The diagnostic usefulness of serum amylase isozyme measurement was investigated in 17 cases of definite chronic pancreatitis and 13 cases of suspected chronic pancreatitis, a total of 30 cases. Six types of serum isoamylase patterns were distinguished according to the character of the two main amylase activities at the fraction of fast-gamma globulin and pre-gamma globulin. The amylase activity of the fast-gamma fraction rises in acute exacerbation, and falls in cases of chronic calcifying pancreatitis and in cases with a marked decrease in pancreatic exocrine function. Especially, a fall in fast-gamma amylase activity excellently reflects a decrease in pancreatic enzyme production. The measurement of amylase activity at the fast-gamma fraction therefore appears to be a useful method for assessing pancreatic damage. The significance of a frequent rise in pre-gamma amylase activity in chronic pancreatitis remains unknown.

Antidepressive Pharmakotherapie als m�gliche Ursache von Speicheldr�sensch�digungen

The side-effects of many psychopharmacons, esp. antidepressants, namely inhibited salivation accompanied by dryness of the mouth, prompted us to investigate the effects on rat salivary glands of an Amitriptyline (Laroxyl®) treatment. In both the parotid and submaxillary gland, medication of different length resulted in proteodyschylic changes as well as alterations in the isoamylase patterns, and histochemically in decreased enzymatic activities. Especially the changes observed in the submaxillary gland are similar to the disturbed glandular function apparent after antihypertensive treatment, e.g., with Clonidine (Catapresan®) which we interpreted as a central neurogenic sialoadenosis. Therefore, besides exhibiting an atropine-like effect, antidepressants possibly also affect sympathetic centres in the medulla oblongata.