Pepsin activity as a function of pH and digestion time on caseins and egg white proteins under static <i>in vitro</i> conditions

Salelles, Lea; Floury, Juliane; Feunteun, Steven Le

doi:10.1039/d1fo02453a

Cited by 50 publications

(29 citation statements)

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“…Although the characteristic bell-shaped curve remained the same for casein substrate 40 as other substrates, we note that the activity change with pH observed in this study on hemoglobin was slightly less than that observed in previous studies for different such as chicken or ostrich substrates; for example, at pH 3 the activity of pepsin on hemoglobin was observed to drop by only 30% in our study (compared to a drop of 60-70% observed in previous studies with other substrates) [45][46][47] . However, we note that these previous studies evaluating pepsin activity as a function of pH employed slightly different methodologies and used different substrates 40 and sources of pepsin such as chicken 45 or ostrich 46 .…”

Section: Pepsin Activity and Solubilitymentioning

confidence: 85%

“…The porcine pepsin (EC 3.4.23.1) activity was determined using the protocol of INRAE-STLO as described in Salelles, et al 40 in 100% H 2 O and 100% D 2 O. In brief, bovine blood hemoglobin (2%, (w/v)) was used as a substrate.…”

Section: Pepsin Activity Assaymentioning

confidence: 99%

“…The activity of porcine pepsin was characterized in the pH range of 1.1 to 5.0 in both 100% H 2 O and 100% D 2 O, using bovine blood hemoglobin as substrate and the INRAE-STLO protocol as described in Salelles, et al 40 . As the porcine pepsin enzyme is known to exhibit maximum activity at pH 2, all measured activities in this study were normalized to the measured activity of pepsin in H 2 O at pH 2 and are provided as a percentage of this value in Table 1.…”

Section: Pepsin Activity and Solubilitymentioning

confidence: 99%

“…We note, however, that the pH activity pro le of pepsin can vary from substrate to substrate as the optimal pH of proteolysis shifts 49,50 . Taking into account the wide variety of edible proteins, Salelles, et al 40 showed that pH activity pro les have differences depending on the substrates. Other substrates, such as wheat gluten or egg white protein, did not produce the bell-shaped curve with maximum activity at pH 2 seen with hemoglobin as a substrate 40,[45][46][47]51 .…”

Section: Pepsin Activity and Solubilitymentioning

confidence: 99%

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Correlating Structure and Activity of Pepsin Enzyme in H2O and D2O for The Study of Gastric Digestion

Bayrak

Han

Greaves

et al. 2022

Preprint

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Pepsin is an aspartic acid protease and the first protease that proteins encounter in the gastrointestinal tract. In vitro digestion assays on proteins including those found in food, or as oral protein and peptide drugs, often involve different solvents or pH conditions where the activity of the digestive enzyme may not be optimal. To improve the accuracy of these assays it is, therefore, important to understand the effect of common experimental conditions, such as solvent, on enzymes such as pepsin. Herein we investigate the activity and structure of porcine pepsin in H2O and D2O at pH values between 1–8. When dissolved in D2O, the enzyme activity of pepsin between pH 1–3 decreased to 60% of the activity in H2O. However, the relative change in activity with pH was similar in D2O and H2O. CD measurements demonstrated that changing the pH and solvent did not influence the secondary structure of the pepsin enzyme. SAXS analysis revealed that structural changes to pepsin did not occur until a pH value between pH 7 and 8, at which point it was denatured and adopted an extended state. Therefore, changes in the pepsin enzymatic activity with pH and solvent change were found to be related to the solubility of pepsin but not to the structure of the protein. In digestion studies, pepsin activity is determined based on its measured activity in H2O at pH 3, regardless of the dynamic pH conditions in digestion or the local environment of the pepsin. This research has therefore significant applicability in improved setups for future digestion and drug bioavailability experiments, as well as future neutron scattering, NMR, and FTIR experiments for enzyme studies in D2O.

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Section: Pepsin Activity and Solubilitymentioning

confidence: 85%

Section: Pepsin Activity Assaymentioning

confidence: 99%

Section: Pepsin Activity and Solubilitymentioning

confidence: 99%

Section: Pepsin Activity and Solubilitymentioning

confidence: 99%

See 2 more Smart Citations

Correlating Structure and Activity of Pepsin Enzyme in H2O and D2O for The Study of Gastric Digestion

Bayrak

Han

Greaves

et al. 2022

Preprint

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“…pathogenicity ( De Sousa-Carvalho et al, 2011 ), it was of interest to analyze also the influence of collagenase on A. castellanii excystment. In addition, pepsin was used in non-optimal conditions, at pH 6.5, which is distant from its optimal pH at 2 and where only a residual activity has been previously observed ( Salelles et al, 2021 ), thus favoring a gentle digestion of the FLA cyst wall. Indeed, at pH 2, no excystment was observed in our conditions after an incubation of up to 18 days at 30°C in the presence of 100 U/mL pepsin, while the amoebae were able to excyst after a treatment in the same conditions at pH 2 without the enzyme, showing a cysticidal effect of pepsin at pH 2 on A. castellanii .…”

Section: Discussionmentioning

confidence: 99%

Stimulation of Acanthamoeba castellanii excystment by enzyme treatment and consequences on trophozoite growth

Fechtali-Moute

Loiseau

Pomel

2022

Front. Cell Dev. Biol.

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Acanthamoeba castellanii is a widespread Free-Living Amoeba (FLA) that can cause severe ocular or cerebral infections in immunocompetent and immunocompromised patients, respectively, besides its capacity to transport diverse pathogens. During their life cycle, FLA can alternate between a vegetative form, called a trophozoite, and a latent and resistant form, called a cyst. This resistant form is characterized by the presence of a cell wall containing two layers, namely the ectocyst and the endocyst, mainly composed of cellulose and proteins. In the present work, we aimed to stimulate Acanthamoeba castellanii excystment by treating their cysts with a cellulolytic enzyme, i.e., cellulase, or two proteolytic enzymes, i.e., collagenase and pepsin. While 11 days were necessary to obtain total excystment in the control at 27°C, only 48 h were sufficient at the same temperature to obtain 100% trophozoites in the presence of 25 U/mL cellulase, 50 U/mL collagenase or 100 U/mL pepsin. Additionally, more than 96% amoebae have excysted after only 24 h with 7.5 U/mL cellulase at 30°C. Nevertheless, no effect of the three enzymes was observed on the excystment of Balamuthia mandrillaris and Vermamoeba vermiformis. Surprisingly, A. castellanii trophozoites excysted in the presence of cellulase displayed a markedly shorter doubling time at 7 h, in comparison to the control at 23 h. Likewise, trophozoites doubled their population in 9 h when both cellulose and cellulase were added to the medium, indicating that Acanthamoeba cyst wall degradation products promote their trophozoite proliferation. The analysis of cysts in epifluorescent microscopy using FITC-lectins and in electron microscopy revealed a disorganized endocyst and a reduction of the intercystic space area after cellulase treatment, implying that these cellular events are preliminary to trophozoite release during excystment. Further studies would be necessary to determine the signaling pathways involved during this amoebal differentiation process to identify new therapeutic targets for the development of anti-acanthamoebal drugs.

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Compound probiotics microcapsules improve milk yield and milk quality of dairy cows by regulating intestinal flora

Wu,

Chang,

Zhang

et al. 2024

Food Bioengineering

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To address the issues of probiotic activity loss during storage and feeding, as well as the limited efficacy of single probiotics, a solution was devised by embedding a mixture of Bacillus coagulans SN‐8 (SN‐8) and Saccharomyces boulardii SN‐6 (SN‐6) in a gel. The initial step involved screening the probiotic microcapsules' preparation method and wall material. Using sodium alginate and β‐cyclodextrin as composite wall material and chitosan as the outer coating material allowed for an embedding rate of 82.11% in composite probiotic microcapsules prepared by the air atomization method. Next, in vitro, simulated digestion experiments were conducted to determine the number of viable bacteria and the release rate of the microcapsules. The results showed that compared to the free strain, the mixed probiotic microcapsules retained a survival rate of 67.5% after 3 h of simulated gastric juice exposure and 70.56% after 42 days of storage at 4°C. This demonstrated higher survival rates and storage stability. The prepared probiotic microcapsules were then administered to dairy cows. 16S rDNA gene sequencing showed that consumption of the microcapsules reduced the number of harmful bacteria, such as Paeniclostridium, in the intestinal tract of dairy cows while accelerating the growth of beneficial bacteria, such as Bifidobacterium. In particular, this resulted in a significant improvement in the lactation performance of the cows, with a 4.5% increase in milk fat content, a 92.5% increase in milk protein content, and a 3.5% increase in milk urea nitrogen content (6.75 mL/dL). In conclusion, probiotic microcapsules can effectively regulate intestinal flora, improving milk production, and quality in dairy cows.

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Pepsin activity as a function of pH and digestion time on caseins and egg white proteins under static in vitro conditions

Abstract: The activity of pepsin, the gastric protease, is generally considered to be negligible for pH ≥ 4, based on results obtained with few purified globular proteins. The present study aimed...

Cited by 50 publications

References 50 publications

Correlating Structure and Activity of Pepsin Enzyme in H2O and D2O for The Study of Gastric Digestion

Correlating Structure and Activity of Pepsin Enzyme in H2O and D2O for The Study of Gastric Digestion

Stimulation of Acanthamoeba castellanii excystment by enzyme treatment and consequences on trophozoite growth

Compound probiotics microcapsules improve milk yield and milk quality of dairy cows by regulating intestinal flora

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