Summary of Study ST002447
This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001578. The data can be accessed directly via it's Project DOI: 10.21228/M8QQ79 This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.
| Study ID | ST002447 |
| Study Title | Bioactive molecule(s) of gut bacteria of Crocodile (Crocodylus palustris) as potential pharmaceuticals |
| Study Summary | Crocodiles thrive in unsanitary conditions, feed on rotten meat, are exposed to heavy metals and are among the very few species to endure the catastrophic Cretaceous-Tertiary extinction event, and yet they can live up to 100 years. We hypothesized that crocodiles have developed mechanisms to achieve such longevity while surviving under stressful conditions. We speculate that their microbial gut flora may produce substances contributing to their “hardiness” and “longevity”. Previously we characterized selected microbial gut bacteria colonizing the gastrointestinal tract of Crocodylus porosus (CP) using 16S rDNA sequencing. Next, bacterial conditioned media containing gut microbial metabolites were prepared. Bioassay-guided testing of selected bacterial conditioned media using LC-TIMS-QTOF MS, revealed the identity of gut microbial metabolites. Among two bacterial conditioned media, i.e., CP27 and 36, the analyses resulted in 141 highly confidently (MS/MS) identified metabolites in both samples. The pairwise comparison of the two samples indicated that 109 metabolites change significantly between them (p <0.05). Among abundant metabolites more prevalent in CP36 there were 2-Methyl-4-nitroimidazole, N-Acetyl-L-tyrosine, Acetaminophen, Trans-Ferulic acid, N, N-Dimethylformamide, Pyrocatechol, Cyclohexanone, 3, 4-Dihydrozphenylglycol, Diphenhydramine, Melatonin, Gamma –terpinene. Whereas in CP27 samples the most abundant metabolites were Carbamazepin, deoxyninosine, Cysteamine, Benzylnicotinate, 3-phenoxypropionic acid, Indole-3-carbinol, Benzaldehyde, Benzocaine, 2-Aminobenzoic acid, 3-Methylindole. Functional enrichment analysis of all identified metabolites with metabolite sets based on drug pathways showed that they were enriched for drug action of top ten pathways associating with enalapril metabolism pathway, diphenhydramine H1-Antihistamine action, enalarpil action pathway, benzocaine action pathway, mepivacaine action pathway, oxybuprocaine action pathways, nifedipine action pathway, propranolol action pathway, acetaminophen metabolism pathway, carbamazepine metabolism pathway. These findings suggest that analyses of crocodile gut bacteria may reveal potential drug leads, intensive future research is needed to realize these expectations. |
| Institute | Sharjah Institute for Medical Research |
| Last Name | Facility |
| First Name | Core |
| Address | M32, SIMR, College of Pharmacy, Health Sciences, University of Sharjah, Sharjah, UAE, Sharjah, 000, United Arab Emirates |
| tims-tof@sharjah.ac.ae | |
| Phone | +971 6 5057656 |
| Submit Date | 2022-12-22 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | d |
| Analysis Type Detail | LC-MS |
| Release Date | 2023-06-22 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Combined analysis:
| Analysis ID | AN003988 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Bruker Elute |
| Column | Hamilton Intensity Solo 2 C18 (100 x 2.1 mm, 1.8 µm, 90 Å) |
| MS Type | ESI |
| MS instrument type | QTOF |
| MS instrument name | Bruker timsTOF |
| Ion Mode | POSITIVE |
| Units | AU |
MS:
| MS ID: | MS003722 |
| Analysis ID: | AN003988 |
| Instrument Name: | Bruker timsTOF |
| Instrument Type: | QTOF |
| MS Type: | ESI |
| MS Comments: | Samples were chromatographically separated by inline reversed-phase chromatography using the Elute HPG 1300 pumps and Elute Autosampler (Bruker, Darmstadt, Germany) with solvent A 0.1% FA in HPLC grade water and solvent B 0.1% FA in ACN. A Hamilton Intensity Solo 2 C18 column (100 mm x 2.1 mm, 1.8 µm beads) was maintained at 35℃ (metabolomics analyses) For metabolomics, 10 µL was injected twice for each sample and eluted using a 30-minute gradient as follows: 1% ACN was held for 2 minutes, ramping to 99% ACN over 15 minutes, held at 99% ACN for 3 minutes before re-equilibrating to 1% ACN for 10 minutes. Flow rates were 250 µL/min for elution and 350 µL/min for re-equilibration. The MS analysis was performed using a TimsTOF (Bruker, Darmstadt, Germany) with Apollo II electrospray ionization (ESI) source. The drying gas was set to flow at 10 L/min and the drying temperature to 220℃ and the nebulizer pressure to 2.2 bar. The capillary voltage was 4500 V and the end plate offset 500V. For proteomics, the scan range was 150 – 2200 m/z, and for metabolomics 20-1300 m/z. For metabolomics the collision energy was set to 20 eV, the cycle time to 0.5 seconds with a relative minimum intensity threshold of 400 counts per thousand and target intensity of 20,000. Sodium formate was injected as an external calibrant in the first 0.3 minutes of each LC-MS/MS run. MetaboScape® 4.0 software was used for metabolite processing and statistical analysis (Bruker Daltonics). The following parameters for molecular feature identification and ‘bucketing’ were set in the T-ReX 2D/3D workflow: For peak detection, a minimum intensity threshold of 1,000 counts is required, as well as a minimum peak duration of 7 spectra, with feature quantification determine using peak area. The file masses were recalibrated based on the external calibrant injected between 0-0.3 min. Only features found in at least three of the twelve samples (per treatment), with retention time between 0.3 and 25 minutes and m/z between 50 and 1,000 were considered and the MS/MS import method was set to be averaged. |
| Ion Mode: | POSITIVE |
