---
title: "Ticagrelor Pharmacokinetics Unaffected by Gut Microbiota: Human, In Vitro, and PGF Rat Evidence"
id: "plos-one-10-ticagrelor-and-gut-microbiota-absence-of-pharmacokinetic-interaction-in-vitro"
canonical_url: "https://medichelpline.com/clinical-feed/plos-one-10-ticagrelor-and-gut-microbiota-absence-of-pharmacokinetic-interaction-in-vitro"
content_type: "clinical_feed_article"
specialty: "Cardiology"
source_name: "PLOS ONE (Medicine)"
source_url: "https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0359660"
published_at: "2026-10-01T14:00:00.000Z"
evidence_level: "Journal Feed"
license: "CC-BY-NC-4.0 / Informational Use"
---
# Ticagrelor Pharmacokinetics Unaffected by Gut Microbiota: Human, In Vitro, and PGF Rat Evidence
## Provenance & Clinical Metadata
- **Canonical URL:** https://medichelpline.com/clinical-feed/plos-one-10-ticagrelor-and-gut-microbiota-absence-of-pharmacokinetic-interaction-in-vitro
- **Specialty:** [Cardiology](https://medichelpline.com/clinical-feed/cardiology.md)
- **Primary Source:** PLOS ONE (Medicine)
- **Source URL:** [Original Journal Publication](https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0359660)
- **Published At:** 2026-10-01T14:00:00.000Z
- **Evidence Rating:** Journal Feed
## Executive GIST (TL;DR)
- The study evaluated whether **gut microbiota** influence the pharmacokinetics of **ticagrelor** and its active metabolite **AR-C124910XX** using human pharmacokinetic profiling, in vitro co-incubation, and pseudo-germ-free (PGF) rat models. - Twenty-two healthy volunteers received a single oral 90 mg dose of ticagrelor; plasma was sampled up to 48 hours and analyzed by UPLC-MS/MS to determine pharmacokinetic parameters. 16S rRNA sequencing was applied to fecal samples collected before dosing to profile gut microbiota. - Human results showed marked inter-individual variability: approximately a three-fold range in ticagrelor AUC (0–48 and 0–∞) and up to seven-fold variation in Cmax; individuals clustered into Group H (higher AUC, n=12) and Group L (lower AUC, n=10). - Comparison of gut microbiome between Group H and Group L found no significant differences in α-diversity (ACE, Chao, Simpson, Shannon) or β-diversity by PCoA; taxonomic comparisons did not reveal compositional differences linked to pharmacokinetic groups. - In vitro, human and rat fecal microbiota stocks were prepared and co-incubated anaerobically with ticagrelor; quantitative analysis using UPLC-MS/MS indicated that gut microbiota did not directly metabolize ticagrelor in the incubation system. - In vivo, PGF rats (n=8 per group) were produced using a 2-week antibiotic regimen (ampicillin, neomycin, metronidazole, vancomycin) to deplete intestinal flora; validation used fecal DNA quantification. - Pharmacokinetic experiments in rats compared intragastric (18 mg/kg) and intravenous (3 mg/kg) ticagrelor administration between PGF and control animals. Non-compartmental analysis found no significant differences in AUC (0–48), AUC (0–∞), or Cmax for ticagrelor or AR-C124910XX between groups. - Overall conclusion from all approaches: the **gut microbiota** does not impact ticagrelor pharmacokinetics; observed human inter-individual PK variability is not explained by fecal microbial composition or direct microbial metabolism according to these experiments. - Methods and analytic details: 16S rRNA sequencing performed with Illumina NovaSeq and EasyAmplicon pipeline; plasma quantification by AB Sciex Qtrap 6500 UPLC-MS/MS; statistical tests included unpaired t-test or Mann–Whitney with p < 0.05 significance threshold.
## Clinical Analysis & Structured Key Points
Ticagrelor and gut microbiota: Absence of pharmacokinetic interaction in vitro and in vivo studies | PLOS One Browse Subject Areas ? Click through the PLOS taxonomy to find articles in your field. For more information about PLOS Subject Areas, click here . Article Authors Metrics Comments Media Coverage Reader Comments Figures Figures Abstract Ticagrelor, a commonly used antiplatelet medication, has been found to exhibit considerable inter-individual pharmacodynamic variability. Recent studies have revealed a close relationship between pharmacokinetics and pharmacodynamics, both of which can be influenced by gut microbiota. In this study, we explored whether the pharmacokinetics of ticagrelor is mediated by intestinal flora. We enrolled 22 healthy participants to assess the clinical pharmacokinetic profiles of ticagrelor and its metabolite AR-C124910XX after a single oral dose of ticagrelor. The fecal microbial compositions of individuals who exhibited distinct pharmacokinetic parameters were analyzed using 16S rRNA gene sequencing. Additionally, we conducted experiments involving the co-incubation of ticagrelor with fecal bacteria and a systematic exploration of the pharmacokinetics of ticagrelor in pseudo-germ-free (PGF) rats to illustrate the role of gut microbiota in ticagrelor metabolism, both in vitro and in vivo . Our findings revealed a three-fold difference in the bioavailability of ticagrelor among individuals. However, no difference was found in the composition of gut microbiota among individuals with different pharmacokinetic parameters. Quantitative analysis showed that gut microbiota cannot directly metabolize ticagrelor. Furthermore, there were no significant differences in AUC (0–48) , AUC (0-∞), and C max between ticagrelor and AR-C124910XX in PGF rats compared to normal control, either by single intragastric or intravenous administration. In conclusion, these findings suggest that gut microbiota has no impact on the pharmacokinetics of ticagrelor. Citation: Qu P, Gao Y, Li X, Yang L, Tang J, Chen M, et al. (2026) Ticagrelor and gut microbiota: Absence of pharmacokinetic interaction in vitro and in vivo studies. PLoS One 21(10): e0359660. https://doi.org/10.1371/journal.pone.0359660 Editor: Hideo Kato, Mie University Hospital: Mie Daigaku Igakubu Fuzoku Byoin, JAPAN Received: January 19, 2026; Accepted: September 16, 2026; Published: October 1, 2026 Copyright: © 2026 Qu et al. This is an open access article distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Data Availability: The 16S sequencing microbiota data reported in this paper have been deposited in the Genome Sequence Archive for Human (GSA-Human) at the National Genomics Data Center, China National Center for Bioinformation (GSA Accession Number: HRA019780). These data are publicly accessible via the GSA-Human browse page: https://ngdc.cncb.ac.cn/gsa-human/browse/ . Any additional information required to reanalyze the data reported in this work is available from the institutional contact ( fanlan_038038@163.com ) upon reasonable request. Funding: This study was partly supported by the National Key Research and Development Program of China (2021YFA1301200), the National Natural Science Foundation of China (82373961, 82073945), the scientific research project of Furong Laboratory of Central South University (2023SK2083), and the Natural Science Foundation of Hunan Province (2022JJ30949). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. There was no additional external funding received for this study. Competing interests: The authors have declared that no competing interests exist. 1. Introduction Ticagrelor, a cyclopentyltriazole compound, is a selective adenosine diphosphate receptor antagonist with a dual pathway inhibitory effect on platelets [ 1 , 2 ]. The latest American College of Cardiology and American Heart Association guidelines recommend ticagrelor as a first-line agent for the treatment of acute ST-segment elevation myocardial infarction (STEMI) [ 3 ]. However, there are significant individual differences in the clinical efficacy of ticagrelor. The PLATO subgroup study found that approximately 3%−27% of patients had a low response to ticagrelor (platelet reactivity index > 50%) [ 4 ]. Gene polymorphisms have limited effects on ticagrelor: P2RY12, P2RY1, and ITGB3 polymorphisms do not affect the inhibition of platelet aggregation by ticagrelor [ 5 ], and CYP2C19 and ABCB1 polymorphisms do not affect the clinical efficacy of ticagrelor [ 6 ]. Therefore, the factors that contribute to the individual differences in ticagrelor efficacy need to be further explored. Studies have shown that changes in pharmacokinetics can alter drug efficacy and side effects [ 7 ]. Gut microbiota has been identified as an important factor in the regulation of drug pharmacokinetics and pharmacodynamics, as well as individual variability in drug response [ 8 ]. Recent studies have demonstrated that gut microbiota may affect drug biotransformation including absorption, distribution, metabolism, and excretion, which in turn affects pharmacodynamics, such as the ability of intestinal bacteria to convert drugs into biologically active substances (Sulfasalazine, lovastatin, and ginsenoside Rb1), inactive substances (Ranitidine and metronidazole), and toxic metabolites, etc [ 8 – 12 ]. Currently, the mechanisms of drug pharmacokinetics influenced by gut microbiota can be divided into three main categories: (1) Direct metabolism [ 13 ]; (2) Bioaccumulation [ 14 ]; (3) Gut microbiota metabolites alter drug pharmacokinetics by affecting metabolic enzymes, transporters [ 15 ]. Previous research has tentatively established a potential relationship between the plasma concentration of ticagrelor and its metabolite AR-C124910XX and gut microbiota [ 16 ]. However, the effects of gut microbiota on the pharmacokinetics of ticagrelor are not well explored. Herein, with genetic polymorphism almost ruled out, we intended to explore whether the gut microbiota affects the pharmacokinetics of ticagrelor. By enrolling healthy subjects, we found significant individual differences in the pharmacokinetic profiles of ticagrelor and its metabolite AR-C124910XX, but no differences in the composition of gut microbiota among individuals with distinct pharmacokinetic parameters. Ticagrelor couldn’t be metabolized by gut microbiota in vitro . By comparing the plasma concentration of ticagrelor and AR-C124910XX in pseudo-germ-free (PGF) rats with control rats, we confirmed that gut microbiota had no influence on the pharmacokinetics of ticagrelor in vivo . Our results implied that gut microbiota may not be responsible for the pharmacokinetic differences in ticagrelor. 2. Materials and methods 2.1. Materials and reagents Ticagrelor tablets (AstraZeneca Co., Ltd.), ticagrelor (98% purity, Aladdin Co., Ltd.), and AR-C124910XX (Push Bio-technology Co., Ltd.). Carbamazepine (98% purity) was provided by the National Institutes for Food and Drug Control (Beijing, China) and was selected as the internal standard (IS) for the pharmacokinetics experiment. Ampicillin, metronidazole, vancomycin hydrochloride, and neomycin sulphate were purchased from Sangon Biotech Co., Ltd (Shanghai, China). Gifu Anaerobic Medium, modified (mGAM) was purchased from Nissui Co., Ltd (Tokyo, Japan). QIAamp ® PowerFecal ® Pro DNA Kit was purchased from QIAGEN (Hilden, Germany). Methanol was purchased from Merck & Co., Inc. (Kenilworth, NJ, USA). Formic acid was purchased from ANPEL Laboratory Technologies Inc.(Shanghai, China). Unless otherwise stated, all reagents were of analytical grade. 2.2. Human studies The recruitment period for this study was conducted from June 1, 2020, to August 31, 2020. All participants in this study have signed the informed consent form, and the study protocol has been approved by the Institutional Review Board of the First Affiliated Hospital of Shantou University Medical College (ethical number: 2019005). All subjects were 19–36 years old and in good health. The health status of subjects was judged based on medical history, physical examination, and routine laboratory tests. These subjects had not taken any medications (prescription or nonprescription) for 2 weeks before the study and did not anticipate any need for medication during the study. The clinical details of the included cases are listed in S1 Table . Twenty-two subjects received oral 90 mg ticagrelor with 240 mL water, which was to be swallowed whole, not swallowed or chewed. Plasma samples were collected at designated times for pharmacokinetic analysis. For the subjects, blood samples were collected before drug administration (0 h), and at 20, 40 min, 1, 1.5, 2, 2.5, 3, 4, 6, 8, 12, 24, 36, and 48 h after dosing. 4 mL of blood was collected into a labeled K2-EDTA anticoagulation tube, which was centrifuged at 2,000 × g at 4 °C for 10 min. The plasma supernatants were stored below −80 °C until assay. Hierarchical clustering analysis was applied to identify subject clusters with different pharmacokinetic parameters. 2.3. 16S rRNA sequencing for human gut microbiota The compositions of the intestinal flora of fecal samples from 22 volunteers were examined by 16S rRNA sequencing analysis. The total DNA was isolated from stool samples. After extracting the total DNA of the samples, PCR amplification was performed and the products were purified, quantified, and homogenized to form sequencing libraries. The constructed libraries were first subjected to quality control, and the libraries that passed the quality control were sequenced by Illumina NovaSeq. The results are stored in FASTQ (fq) file format, which contains the sequence information and sequencing quality information. High-quality clean tags were obtained by splicing and filtering the original Reads, followed by subsequent analysis using the EasyAmplicon [ 17 ] pipeline. Then the data were subjected to basic analysis and the processed tags were denoised with “unoise3” to obtain single-base precision ASVs, which were dechimerized, deplasmidized, and non-bacterialized. Species annotation, diversity analysis, and species composition analysis of ASV representative sequences were performed using the Silva database. The α -diversity was computed by the R package ‘vegan’, and the β -diversity was calculated by principal coordinate analysis (PCoA) at the ASV level. The linear discriminant analysis (LDA) effect size (LEfSe) method was applied to explore biomarker features. 2.4. Gut microbiota preparation An appropriate amount of human or rat fresh fecal sample was suspended well in cold physiological saline (w/v, 1:20) before vortexing for adequate mixing and then centrifuged at 500 rpm for 5 min at 4 °C. The resultant fecal supernatant was decanted and centrifuged at 4,000 rpm for 30 min at 4 °C, then the precipitate was re-suspended with 2.5 mL of mGAM to produce gut microbiota suspension. Stocks were generated by adding glycerol to the gut microbiota suspension (final concentration of glycerol, 20%) and were anaerobically stored in −80 °C freezer [ 18 ]. 2.5. Ticagrelor preparation and co-incubation with fecal samples Gut microbiota stock was activated with mGAM broth (1:20). After incubation for 24 h, the medium was centrifuged at 4,000 rpm for 30 min at 4 °C. The supernatant was discarded and the precipitate was re-suspended in mGAM broth as the gut microbiota work solution. The biotransformation of the ticagrelor by the gut microbiota was performed in an incubation system containing 500 μL of the gut microbiota work solution and 5 μL of ticagrelor solution (1.07 μg/mL, solution in dimethyl sulphoxide). Negative control was performed in an incubation system containing 500 μL of the mGAM broth and 5 μL of ticagrelor solution. The incubation system was anaerobically incubated at 37 °C for 24 h and stopped with the addition of 600 μL cold methanol including IS (carbamazepine) in a 200 μL incubation system. The samples were centrifuged at 14,000 rpm for 10 min and detected by UPLC-MS/MS. 2.6. Animals SD rats (specific pathogen-free (SPF), Certificate No. SCXK2019–0004), 250 ± 20 g , were purchased from Hunan SJA Laboratory Animal Co., Ltd (Changsha, China). The animals were maintained in the SPF animal facilities of the Animal Experimental Center of Central South University and natural light-dark cycle conditions with controlled temperature (22 ± 2°C), food and drinking water supplied unrestricted. All animal work was done in accordance with the National Institutes of Health’s Guide for the Use and Care of Laboratory Animals and was approved by the Institutional Review Board of Central South University (Hunan, China. permission numbers: CSU-2022-0001-0017). Rats were acclimated to the laboratory for 1 week before the experiment and randomized into two treatment groups: control group and PGF group (pseudo-germ-free) (n = 8). Rats in the PGF group were gavage with an antibiotic cocktail (100 mg/kg ampicillin, 100 mg/kg neomycin sulfate, 100 mg/kg metronidazole, and 50 mg/kg vancomycin), and provided with an antibiotic cocktail (0.25 g/L ampicillin, 0.25 g/L neomycin sulfate, 0.25 g/L metronidazole and 0.125 g/L vancomycin) in drinking water for 2 weeks to deplete the intestinal flora. To validate the pseudo-sterile rat model, rat feces were collected and fecal DNA was extracted using a QIAamp ® PowerFecal ® Pro DNA kit. The amount of intestinal microbiota DNA was measured using a BioSpec-nano spectrophotometer to assess the stability and validity of the pseudo-sterility model [ 19 ]. All surgeries were performed under anesthesia (isoflurane). Animals were euthanized when humane endpoints were reached, which included abnormal posture, failure to groom, lethargy, impaired ambulation or paralysis, weight loss exceeding 20% normal body weight, and the inability to voluntarily access food and water. Rats reaching these endpoints were euthanized immediately via CO2 inhalation followed by cervical dislocation to minimize suffering and distress. Animals were monitored daily for health and behaviour, and no animals died before meeting euthanasia criteria. All research staff received training in animal anaesthesia, animal surgery, experimental procedures, and monitoring/nursing. All animal welfare considerations were taken to minimize suffering and distress, including frequent monitoring and immediate euthanasia at endpoint. 2.7. Pharmacokinetic experiments Rats were fasted 12 hours before the pharmacokinetic experiments. 18 mg/kg of ticagrelor (0.5% CMC-Na solution as solvent) was administered to the rats by intragastric gavage. Blood samples were collected from the tail vein at 0, 0.25, 0.5, 1, 2, 3, 4, 6, 7.5 and 24 h. Additionally, 3 mg/kg of ticagrelor (2% Tween 80 solution as solvent) was intravenously administered to the rats. The blood samples were collected from the tail vein at 0, 0.25, 0.5, 1, 2, 3, 4, 5, 6, and 24 h. The blood sample was centrifuged at 4,000 rpm at 4 °C for 15 min and supernatant plasma was collected and stored at −80 °C for further analysis. 2.8. Quantification of plasma ticagrelor and AR-C124910XX Compounds were extracted from plasma by albumen precipitation method. The concentration of the ticagrelor and AR-C124910XX was determined on the AB Sciex Qtrap 6500 UPLC-MS/MS system (AB Sciex, Massachusetts, USA). Targeted analytes were separated using an ACE Excel 5 Super C18 column (150 mm x 2.1 mm, Radnor, PA, USA); the flow rate was maintained at 0.4 mL/min. The mobile phase consisted of solvent A (deionized water with 0.1% formic acid) and solvent B (methanol). A gradient program was used for the HPLC separation: 0–0.5 min (30% A); 0.5–1.5 min (30%−5% A); 1.5–2.0 min (5% A); 2.0–2.5 min (5%−30% A); 2.5–4.0 min (30% A). The oven temperature was maintained at 40℃ and the injection volume was set at 2 μL. The mass spectrometer was operated in positive ion mode through multiple-reaction monitoring. The precursor-product ion pairs monitored were m/z 523.4 → 152.9 for ticagrelor (DP 185 V, CE 46 eV), m/z 479.3 → 153.2 for AR-C124910XX (DP 155 V, CE 44 eV), and m/z 237.1 → 194.1 for carbamazepine (DP 130 V, CE 15 eV). Data were collected and processed using AB Sciex Analyst 1.4.2 data collection and integration software. 2.9. Statistical analysis Pharmacokinetic parameters were analyzed by DAS 2.0 software (Chinese Pharmacological Association, China) using non-compartmental analysis. All data were represented as mean ± standard deviation (SD). The significance between the two groups was performed by using the unpaired two-tailed t-test or Mann-Whitney test, and p < 0.05 indicated that the difference was statistically significant. 3. Results 3.1. Individual variability in clinical pharmacokinetics of ticagrelor To evaluate the population pharmacokinetic profile of ticagrelor in Chinese healthy individuals, 22 healthy individuals were recruited and received a single oral dose of ticagrelor (90 mg), then measured by UPLC-MS/MS. The plasma concentration-time curves of ticagrelor and AR-C124910XX within 48 h in healthy subjects are shown in Fig 1 , and the mean pharmacokinetic parameters are summarized in Table 1 . For ticagrelor, there was a 3-fold difference between individuals in AUC (0–48) (1,429 ng·h/mL −5,582 ng·h/mL) and AUC (0-∞) (1,568 ng·h/mL −5,746 ng·h/mL). At the same time, we found a 7-fold difference in the C max of ticagrelor (137 ng/mL −953 ng/mL). The AUC (0–48) , AUC (0-∞) , and C max of AR-C124910XX also showed significant individual differences. Altogether, these results indicated that large individual differences in the population pharmacokinetics of ticagrelor. Download: PNG larger image TIFF original image Table 1. Pharmacokinetic parameters of ticagrelor and AR-C124910XX in healthy subjects. https://doi.org/10.1371/journal.pone.0359660.t001 Download: PNG larger image TIFF original image Fig 1. The plasma concentration-time curve of ticagrelor and AR-C124910XX within 48 h in healthy subjects. Data are represented as mean ± SD. n = 22. https://doi.org/10.1371/journal.pone.0359660.g001 3.2. Characteristics of gut microbiome of individuals with different pharmacokinetic parameters of ticagrelor To explore whether individual differences in pharmacokinetic parameters of ticagrelor were related to gut microbiota. Subjects were divided into two groups based on ticagrelor pharmacokinetic parameters by using hierarchical clustering, Group H (with higher AUC, n = 12) versus Group L (with lower AUC, n = 10) ( S1 Fig ). Compared to Group L, the AUC (0–48) , AUC (0-∞) , C max , and t 1/2 of ticagrelor were significantly higher than in Group H, with AUC (0–48) increased by 79.51%, AUC (0-∞) increased by 80.77%, C max increased by 62.43% and t 1/2 increased by 30.34%. However, CL was significantly reduced by 45.51%. In addition, the AUC (0–48) , AUC (0-∞), and C max of AR-C124910XX in Group H showed an increasing trend, but there was no statistical significance compared to Group L ( S2 Fig , Table 2 ). These results suggested a significant increase in the bioavailability of ticagrelor for individuals in Group H. Download: PNG larger image TIFF original image Table 2. Pharmacokinetic parameters of ticagrelor and AR-C124910XX between the Group H and Group L. https://doi.org/10.1371/journal.pone.0359660.t002 Microbial DNA was isolated from fecal samples collected before ticagrelor administration and ed by 16S rRNA amplicon sequencing was performed to investigate the differences in gut microbiota between Group H and Group L. The results revealed no notable differences in α - (ACE, Chao, Simpson, and Shannon index) and β -diversities between Group H and Group L ( Figs 2A and 2B). Additionally, we compared the abundance of bacteria taxa
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