K. F. McGeeney

Purification and properties of an α-amylase inhibitor from wheat

Four inhibitors of alpha-amylase (EC 3.2.1.1) were separated from an alcohol extract of wheat by ion-exchange chromatography on DE52-cellulose. One inhibitor, which showed the greatest specificity for human salivary amylase relative to human pancreatic amylase, has been purified by the following steps: (a) alcohol fractionation (60--90%) of water extract (b) ion-exchange chromatography on QAE-Sephadex A-50; (c) re-chromatography on DE52-cellulose and (d) gel filtration on Sephadex G-50. The purified inhibitor is 100 times more specific for human salivary amylase than for human pancreatic amylase. It shows an electrophoretic mobility of 0.2 on disc gel electrophoresis and a molecular weight of about 21 000. This inhibitor contributes about 16% to the total salivary amylase inhibiting power of the wheat extract.

Pancreatic Atrophy: A New Model Using Serial Intra-Peritoneal Injections of L-Arginine

No simple rat model for chronic pancreatitis exists at present. A single dose of arginine has recently been shown to induce acute pancreatitis in rats. This study was designed to assess whether serial injections of arginine would induce reproducible chronic pancreatic damage. Forty rats received an intra-peritoneal injection of 500 mg per 100 g body weight of L-arginine followed by three injections of 250 mg per 100 g over 10 days. The rats were killed 24 h after each injection and at intervals of up to 6 months. Serum amylase levels were increased in the acute phase only. Examination of the pancreas at 24 h showed a severe oedematous pancreatitis. By day 5 there was up to 90% acinar destruction with adipose tissue replacement, although ductal, vascular, and islet cells appeared undamaged. These changes were present 6 months after injection. This is proposed as a new, simple, and reproducible method of inducing chronic pancreatic damage in the rat.

THE IMPORTANCE OF VARYING MOLECULAR SIZE, DIFFERENTIAL HEAT AND UREA INACTIVATION OF PHOSPHATASE IN THE IDENTIFICATION OF DISEASE PATTERNS*

Our interest in alkaline phosphatase studies started in 1964 when we were using gel filtration techniques for many collateral studies. We studied serum in patients with various types of liver disease, and noted that an enzyme was present that was excluded from Sephadex G-200, i.e., it eluted mainly with the macroglobulins.l.2 This suggested an enzyme of unusual size? and prompted our subsequent investigations. During this time, other methods of identifying patterns have been described, and we have used some of them, such as differential therm~stabili ty,~,~ differential urea inactivation68 and phenylalanine inh ib i t i~n .~ Our studies have been devoted mainly to patterns obtained in a variety of diseases involving the hepatobiliary and skeletal systems.

Secretory effect of the venom of the scorpion Tityus trinitatis on rat pancreatic slices.

A linear concentration response relationship was found for amylase release from rat pancreatic slices by venom of the scorpion Tityus trinitatis. Maximum release of amylase was caused by 20 μg venom per ml of medium. Acetylcholine, used as a standard stimulant had a maximum effect at 3 × 10−7 M. Atropine partially blocked both venom and acetylcholine induced release of amylase. Nicotine, hexamethonium and tubocurarine did not affect this stimulated amylase release, nor did they alter the non-stimulated (basal) release of amylase. Physostigmine potentiated the venom stimulated amylase release but had no effect on non-stimulated release. These observations suggest that the venom exerts its secretory effect through a cholinergic mechanism which may involve muscarinic receptors.

Pancreatic and salivary amylase/creatinine clearance ratios in normal subjects and in patients with chronic pancreatitis.

The clearance of pancreatic and salivary amylase relative to creatinine was measured in 26 control subjects and 22 patients with chronic pancreatitis. Control values for pancreatic amylase clearance (+/- SD) were 2.64 +/- 0.86% compared with 1.54 +/- 0.95% for salivary amylase. In chronic pancreatitis, pancreatic amylase clearance ratios were significantly higher than controls (P less than 0.0005, mean 4.09 +/- 1.63 SD). The difference in clearance rate of salivary amylase did not reach a level of significance when compared with the control group. Twelve of the 22 patients showed pancreatic amylase clearance values above the normal limit of 4.4, while only five were abnormal when the clearance of total amylase was measured. The patients also showed statistically higher (P less than 0.0005) levels of serum salivary amylase when compared with 69 control sera. No such difference was found for the pancreatic component of serum amylase. Comparison of beta2-microglobulin clearance values showed no statistical difference between patients and controls.

α-Amylase and glucoamylase activities of canine serum

Abstract 1. 1. Three methods of α-amylase analysis were compared: (a) a saccharogenic method, (b) an amyloclastic method and (c) the blue starch method. 2. 2. The saccharogenic method yielded falsely elevated valeus for canine serum, i.e. 1·7 times higher than the blue starch or iodometric methods. 3. 3. This excess saccharogenic activity was shown to correlate well with the glucoamylase activity. 4. 4. Gel filtration of canine serum separated α-amylase from glucoamylase. 5. 5. The latter enzyme did not hydrolyse the insoluble blue starch polymer. 6. 6. The blue starch method appears to be specific for α-amylase.

Modified high amylose starch for immobilization of uricase for therapeutic application

Urate oxidase (uricase) was immobilized on carboxymethyl high amylose starch cross‐linked 35 (CM‐HASCL‐35), on aminoethyl high amylose starch cross‐linked 35, as well as on commercial supports, CNBr‐activated Sepharose and diaminodipropylamine agarose. The N‐ethyl‐5‐phenylisoxazolium‐3′‐sulphonate (Woodward reagent K) gave a high binding but totally inhibited the enzyme activity. Best results were obtained with CM‐HASCL‐35 using 1‐ethyl‐3‐(3‐dimethylaminopropyl)carbodi‐imide as a coupling agent. The immobilized enzyme retained 88% of its initial limit rate [Vmax(app)=16 EU/mg for immobilized uricase versus Vmax=18 EU/mg for the free enzyme], with an apparent decrease of affinity for urate substrate [Km(app)=0.17 mM versus Km=0.03 mM for the free enzyme]. The coupling yield was 60% and the modified uricase was found more resistant to proteolysis than the free enzyme. The immobilized uricase retained 25% of its initial activity after 60 min in pancreatic proteolysis medium (pancreatin), whereas the free enzyme retained only 5% of its initial activity. The best immobilization yield was obtained with the polymeric support based on CM‐HASCL‐35 (53%), which gave better results than commercial supports based on agarose.

Exocrine pancreatic response to the venom of the scorpion, Tityus trinitatis

This paper records for the first time the exocrine pancreatic response to scorpion venom, in this case that of Tityus trinitatis, a scorpion endemic in Trinidad. The crude venom injected intravenously into fasting anaesthetised dogs induced a secretion of the exocrine pancreas. The secretion evoked was rich in enzymes.

The effects of glucagon on the exocrine secretion of the perfused canine pancreas

SummaryVOLUME, amylase, lipase and trypsin secretion in the totally isolated canine pancreas were studied. Glucagon does not have an inhibitory effect on the secretin or CCK-Pz evoked exocrine secretion of the isolated gland. In the gland stimulated by large combined doses of secretin and CCK-Pz, glucagon increases enzyme secretion. These findings may indicate that the inhibitory action of glucagon in the intact animal is indirect.

Response of the isolated pancreas to scorpion venom

SummaryTHE venom of the scorpionTityus trinitatis causes secretion of the secretin stimulated isolated canine pancreas. In the absence of secretin, similar doses of venom evoked no response. The implications of these results are discussed. The results with this isolated model show that the action of the venom is directly on the pancreas. Further, these results provide the opportunity to offer a rationale for the pathogenesis of that form of pancreatitis which occurs following the sting of the scorpion.