The Effect of Temperature and pH on the Stability of Human Pepsin in Stored Gastric Juice: A Method to Prevent Activity Loss

Abstract

de Gara CJ, Burget DW, Sivakumaran T, Hunt RH. The effect of temperature and pH on the stability of human pepsin in stored gastric juice. A method to prevent activity loss. Scand J Gastroenterol 1986, 21, 650-654The mechanisms controlling pepsin secretion are controversial. A contributory factor may be storage-dependent effects. We have studied the effects of temperature, pH, and storage time on human gastric pepsin. Gastric juice samples taken from three healthy volunteers under both basal and post-pentagastrin-stimulated (6 pg/kg subcutaneously) conditions were separated into four aliquots. Each aliquot was titrated to pH 1,4, or 6 or left at ambient pH. Aliquots were then stored at 4°C or frozen at -70°C and stored. On days 1, 3, 7, and 28 aliquots were removed and assayed by the kinetic albumin-bromphenol blue method. In a second experiment we determined the effects of different concentrations of glycerol on the preservation of peptic activity. From these experiments we conclude that pepsin is unstable when stored frozen at low pH but not when stored above pH 2. This pH-dependent stability may explain the variable conclusions other workers report on optimal methods of storing gastric juice. In addition, we have confirmed the suitability of glycerol as a preservative of peptic activity and recommend that gastric juice be stored frozen with 11.5% glycerol.

Characterization of the Human Upper Gastrointestinal Contents Under Conditions Simulating Bioavailability/Bioequivalence Studies

“…Protein Content: Immediately upon aspiration and before storage at j70-C, gastric subsamples in which total protein content was to be measured were titrated to pH 1 to inhibit proteolytic activity of pepsin (27). Similarly, in duodenal aspirates phenylmethylsulfonyl fluoride (PMSF) was added to inhibit trypsin activity by achieving an end concentration of 1 mM (28).…”

“…Pepsin Activity: Immediately upon collection and before storage at j70-C, gastric subsamples in which pepsin activity was to be measured were titrated to pH 6 (27). Pepsin activity was measured by a modification of the method described by Anson (30), and quantification was based on hog pepsin as a standard.…”

Human Gastrointestinal Juices Intended for Use in In Vitro Digestion Models

“…The specific activities of the enzymes in such pooled samples must be analysed, and their stability during storage should be monitored so that the juices are used while the enzymes are still active. Loss of pepsin activity in frozen gastric juice has been reported [20]. However, de Gara et al [20] showed that the activity could be preserved by the addition of glycerol or by increasing the pH outside of the optimal pH range for pepsin.…”

“…Loss of pepsin activity in frozen gastric juice has been reported [20]. However, de Gara et al [20] showed that the activity could be preserved by the addition of glycerol or by increasing the pH outside of the optimal pH range for pepsin. The amylase activity in duodenal juice stored at −20 °C remained largely stable in a study by Muller and Ghale [21] whereas lipase and trypsin activities in hormonally stimulated juice decreased to approximately 50% of the initial activities within 3 weeks.…”

Effectiveness of cricoid pressure in preventing gastric aspiration during rapid sequence intubation in the emergency department: study protocol for a randomised controlled trial

“…Pepsin activity is highly sensitive to pH with maximum activity between pH 1.5-2.5 and irreversible inactivation occurring above pH 7.5 [43]. Pepsin stored in gastric juice samples at 4°C remains active for up to 3 days (evaluated up to pH 6)[44]. The pH of saliva sampled from patients following episodes of gastric regurgitation has been reported to range from 7-8 [45].…”