Characterization of Stability of the Young Porcine Intestinal Alkaline Phosphatase as a Candidate Exogenous Biocatalyst
Bibliographic record
Abstract
Intestinal alkaline phosphatase (IAP) is a critical anti‐inflammatory mediator through possessing the ability to catalyze the hydrolytic dephosphorylation of endotoxin lipopolysaccharides (LPS) and other emblematic members of pathogen‐associated‐molecular patterns (PAMPs) such as ATP, thus preventing gut dysbiosis and disorders. Our previous work has demonstrated that early weaning decreases IAP affinity, indicating an increased susceptibility to bowel inflammation and infection in weanling piglets. To determine if the porcine IAP is suitable to serve as an efficacious exogenous feed enzyme biocatalyst, several key stability aspects of the AP need to be thoroughly characterized, including thermostability, i.e., resistance to heating typically at 70 – 80°C for 8 to 10 min during compound feed pelleting, gastric stability and resistance to exocrine pancreatic proteases' hydrolysis. This study was conducted to investigate the stability of the porcine IAP with jejunal samples collected from four 10‐day‐old Yorkshire piglets. The IAP activity was measured in the media containing 1.75 mM P ‐nitrophenyl phosphate at pH 7.4 for 30 min after exposing homogenized porcine jejunal tissue samples (19 – 27 μg) to various stability effectors. Stability of the IAP was assessed through inhibition kinetic analysis by the Eadie‐Hofstee model, including parameter endpoints (parameter estimates ± SE, n = 24) of TC 50 representing the temperature (°C) or time (h) at half the maximal inhibition of the IAP activity; and I max representing the relative maximal level (% of the control) of inhibition in the IAP activity. Thermostability of the porcine IAP was determined by heating the tissue homogenate at the temperature ranging from 22 – 80 °C for 10 min, and was associated with the inhibition kinetics of TC 50 = 24.5 ± 3.2 °C and I max = 80.0 ± 20.0 %. Gastric stability of the porcine IAP was determined by exposing the tissue homogenate to the pH at 3.5 for 0 – 5 h, and was associated with the inhibition kinetics of TC 50 = 0.40 ± 0.06 h and I max = 81.5 ± 3.2 %. Resistance of the porcine IAP to the gastric protease was determined by exposing the tissue homogenate to the 266 U/mL pepsin in the media for 0 – 5 h, and was associated with the inhibition kinetics of TC 50 = 0.46 ± 0.03 h and I max = 92.4 ± 0.8 %. Resistance of the IAP to the exocrine pancreatic proteases was determined by exposing the tissue homogenate to the 66 U/mL trypsin and 59 U/mL chymotrypsin in the media for 0 – 5 h, and was associated with the inhibition kinetics of TC 50 = 0.24 ± 0.11 h and I max = 11.7 ± 1.2 % for the trypsin effect; and of TC 50 = 0.09 ± 0.14 h and I max = 18.54 ± 0.95 % for the chymotrypsin effect, respectively. These results suggest that while the porcine IAP displays a high level of resistance to pancreatic proteases, it is not thermostable, and it is also very susceptible to the deactivation by the gastric acidic pH and pepsin environment. Strategies to improve the gastric acidic pH, protease and heat stability of the porcine IAP are required as a pre‐requisite to use this enzyme as an exogenous feed enzyme. Support or Funding Information NSERC Discovery Program of Canada and Metagen Enzyme Corporation
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.002 | 0.002 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.001 | 0.001 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.001 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".