Assay of gastricsin and individual pepsins in human gastric juice.

Abstract

Aims: To develop and validate an analytical procedure for the quantitation of pepsins and gastricsin in human gastric juice and to assess its potential in a controlled gastric secretory study. Methods: High performance ionexchange chromatography was used to separate human pepsin 1, 3a, 3b, 3c and gastricsin from gastric juice. Computed chromatographic areas for each enzyme were quantified by relation to a known amount of a secondary standard porcine pepsin. The assay procedure was validated by recovery and analytical precision studies. Gastric secretions after pentagastrin and insulin stimulation from 10 patients with portal hypertension were used to assess the potential of the analytical procedure. Results: The assay precision varied from 1.5 to 9*0% within batch and 7 5 to 181% between batch, with about 100% recoveries of porcine pepsin A from human gastric juice over the assay range 0 025-0 5mg/ml. A fourfold increase in combined pepsin and gastricsin concentration was observed following pentagastrin and insulin stimulation. The mean percentage content of pepsins 3a, 3b, 3c, and 1 in non-stimulated gastric juice were 4% 72%, 12% and 1-4%, respectively, and did not change significantly after gastric stimulation. An approximate doubling of the percentage of gastricsin (10% to 20%) relative to the pepsins was observed, however, after both insulin and pentagastrin stimulation. Conclusions: This procedure for quantifying individual human pepsins and gastricsin in gastric juice is simple and reliable. It may be of considerable importance in determining the mechanisms involved in the control and secretion of these digestive enzymes in man, including the effect of anti-ulcer drugs and our understanding of the pathophysiology of peptic ulcer disease. (JClin Pathol 1993;46:254-258) There is a paucity of simple and reliable assays suitable for the quantitation of individual human pepsins in gastric juice, and in particular human pepsin 1 (nomenclature of Etherington and Taylor (Radiometer,

Sensitive Pepsin Immunoassay for Detection of Laryngopharyngeal Reflux

“…The median pepsin concentration of eight human gastric juice samples was 0.4 mg/mL (range 0.09–0.8 mg/mL). These values are within the expected pepsin concentration range of gastric juice 10 …”

“…Inevitably, there may be a matrix or pattern that is not only diagnostic of LPR but also of its severity and it role in causing disease. We believe that the pepsin assay as a spit‐in‐a‐cup test will be found to be the best available screening test for LPR and that tissue sampling tested for markers such as pepsin, carbonic anyhydrase, and E‐cadherin will be diagnostic 8–15 …”

“…Purification of pepsin from gastric juice was achieved by ion-exchange chromatography on a Pharmacia MonoQ 5/50 column (Amersham Biosciences, Piscataway, NJ) developed with a gradient of 0.15 to 0.3 mol/L NaCl in 50.0 mmol/L sodium acetate, pH 4.1, as described previously. 10 The major pepsin A isoform, pepsin 3b, was used for antibody production and immunoassay development. Identity and purity of pepsin 3b was confirmed by N-terminal amino acid sequence analysis and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE).…”

Activity/Stability of Human Pepsin: Implications for Reflux Attributed Laryngeal Disease

“…Fractions containing protein (determined by measuring optical density at 280 nm) were pooled and dialyzed against 50 mmol/L sodium acetate at 4°C to remove all salt before performing the second high‐resolution anion exchange chromatography with a Pharmacia Mono Q, 0.5 × 5 cm, column (GE Health Care, Piscataway, NJ) developed with a linear gradient of 0.09 to 1.0 mol/L NaCl in 50 mmol/L sodium acetate, pH 4.1, over 30 minutes at 0.75 mL/minute. Pepsin 3b was identified by chromatography pattern, isolated, and stored at ‐20°C in 50% glycerol 11 …”

Role of Extra‐Esophageal Reflux in Chronic Otitis Media with Effusion

“…Fractions containing protein (determined by measuring optical density at 280 nm) were pooled and dialyzed against 50 mmol/L sodium acetate at 4°C to remove all salt prior to performing the second high‐resolution anion exchange chromatography with a Pharmacia Mono Q 5/50 column (GE Health Care, Ltd., Piscataway, NJ) developed with a gradient of 0.09 to 1.0 mol/L NaCl in 50 mmol/L sodium acetate, pH 4.1, over 30 minutes at 0.75 mL/min. Pepsin 3b was identified by chromatography pattern, isolated, and stored at −20°C in 50% glycerol 12 …”