Chlorzoxazone

Limits on what this shows

• Key null findings: Repair-pass PubMed listing for chlorzoxazone with respiration, oxidative phosphorylation or mitochondrial toxicity returned 7 records; none measured mitochondrial function with chlorzoxazone as the exposure (chlorzoxazone appeared as a CYP2E1 substrate in HepG2 hypoxia-reoxygenation work, as a P450 inhibitor in a cumene hydroperoxide hepatocyte study, in a spasticity trial and in an overdose report), and Xu 2008 was the one record that named it as a hepatotoxicant [24][27]. The earlier listing for chlorzoxazone and mitochondria (8 records) also had no chlorzoxazone-exposed mitochondrial result [24]. No circulating consumer claim about mitochondrial damage by chlorzoxazone was retrieved.
• At-risk subgroups: None identified.

Direction

Drug toxicity burden signal in cultured cells (mechanistic, Tier B). Not a depletion, not a retention effect and not a nutrient deficiency (Hard Rule 2). Whether the flag arose from the mitochondrial readout, the oxidative stress readout or the glutathione readout is unknown, so this entry does not establish a mitochondrial effect on its own.

Regulatory status

No mitochondrial content in the label texts read [1][2].

Population evidence

None in humans. Xu et al. 2008 (Toxicol Sci 105:97): imaging in primary human hepatocyte cultures of mitochondrial damage, oxidative stress and intracellular glutathione across more than 300 drugs and chemicals; the abstract, opened on its PubMed record, reports a true-positive rate of 50 to 60 percent with a false-positive rate of 0 to 5 percent and states that "these in vitro predictions can augment the performance of the combined traditional preclinical animal tests by identifying idiosyncratic human hepatotoxicants such as nimesulide, telithromycin, nefazodone, troglitazone, tetracycline, sulindac, zileuton, labetalol, diclofenac, chlorzoxazone, dantrolene, and many others." Reading chlorzoxazone as a drug the panel flagged is an inference from that sentence. The full text, tables, supplementary data and per-drug results were not accessible: the publisher page and the PDF link opened only abstract-level content (the DOI redirected to an abstract-only page), and OpenAlex returned an inverted index of the abstract [8][24].

Mechanism

Named only at panel level: mitochondrial damage, oxidative stress or glutathione change in hepatocytes as DILI-linked cellular signals [8]. For chlorzoxazone specifically, a CYP3A-generated oxirane trapped by glutathione is the only named pathway, in vitro and in rat (see Hepatic function) [7]. No electron transport, membrane potential or biogenesis result for chlorzoxazone was read.

Counter-evidence held on record

Counter-evidence and limits This is the weakest yellow in the dossier. It is a single abstract sentence in a screening paper, the per-drug readout is unseen, the concentration is unknown, and an in vitro flag does not show a mitochondrial effect at therapeutic exposure in people. The panel's reported true-positive rate was only 50 to 60 percent, and a flag could come from any of three different readouts, so the mitochondrial component cannot be separated. Graded yellow only because a Tier B study using this drug exists whose design includes a mitochondrial endpoint; a reviewer who requires a per-drug mitochondrial result would return it to ungraded. Framing caution Do not say chlorzoxazone damages mitochondria in patients or lowers cellular energy. This is a cell-culture screening statement about liver injury potential. No supplement (for example a mitochondrial or antioxidant product) is implied.

Limits on what this shows

• Key null findings: Two human studies measured oxidative variables around chlorzoxazone but gave it only as a probe to all participants: Lainé 2006 (phlebotomy trial, 48 participants) and Somparn 2007 (35 thalassemia patients, 42 controls), where blood glutathione fell and protein carbonyl and lipid peroxidation rose in thalassemia and "no correlations between CYP2E1 activity and oxidant stress or antioxidant parameters" were found [15][16]. These measure disease or iron-removal effects, not an effect of chlorzoxazone, so they do not test the claim. PubMed listings for chlorzoxazone AND oxidative stress, reactive oxygen species, antioxidant, lipid peroxidation and glutathione were all refused (HTTP 429) and not retrieved.
• At-risk subgroups: None identified.

Direction

Burden-direction mechanism (drug toxicity pathway). Not a depletion finding: whether cellular glutathione fell was not stated in the abstract read. Two further studies name chlorzoxazone among tested agents without a per-drug direction available to this review (see Population evidence).

Regulatory status

No oxidative or antioxidant content in the label texts read [1][2].

Population evidence

None in humans for chlorzoxazone as the exposure. Sun et al. 2024: glutathione and N-acetylcysteine conjugates of an oxirane intermediate, as above [7]. Xu et al. 2008 (Toxicol Sci): in primary human hepatocyte cultures, high-content imaging of mitochondrial damage, oxidative stress and intracellular glutathione across over 300 chemicals; the abstract names chlorzoxazone among the drugs examined, but the per-drug result, concentration and DILI category were not visible (abstract only on OpenAlex and the publisher page) [8]. Patel et al. 2009 (Free Radic Res): rat neutrophils, the abstract lists chlorzoxazone among intermediate and small conductance calcium-activated K+ channel modulators that altered NO-induced free radical generation and membrane polarity; its direction for chlorzoxazone was not stated [19].

Mechanism

Named, in vitro and rat: CYP3A bioactivation and glutathione trapping [7]. K+ channel modulation altering neutrophil radical generation is a pharmacology signal at an unstated concentration [19].

Counter-evidence held on record

Counter-evidence and limits Tier B only (mechanistic studies using this drug). The 200 µM microsomal exposure and the 10 to 400 µM hepatocyte range are not related here to therapeutic plasma concentrations (about 7 µg/mL, roughly 40 µM by this dossier's own conversion of the 400 mg Cmax in [14] using a molar mass of 169.6 g/mol). The conjugates show a reactive metabolite forms; they do not show an antioxidant shortfall in people. Framing caution Do not describe chlorzoxazone as causing glutathione depletion or oxidative stress in patients, and do not describe it as an antioxidant. These are test-tube and animal findings. No antioxidant supplement is implied.

Limits on what this shows

• Key null findings: PubMed listing for chlorzoxazone AND iron returned 8 records; apart from [15] and [16] they were CYP2E1 variant, voltammetry, ligand-binding and hepatoma-cell papers with no iron outcome in chlorzoxazone users [24]. A listing for chlorzoxazone with copper, cupric or ceruloplasmin returned no record with chlorzoxazone as an agent. No hemoglobin, ferritin or transferrin saturation result in chlorzoxazone users was found.
• At-risk subgroups: None supported. People with iron overload or thalassemia may metabolize chlorzoxazone differently, per [16], with unclear direction.

Direction

Two separate things. (1) Labeled bleeding is an adverse-reaction outcome; any resulting iron loss is hypothetical and unmeasured (Hard Rule 2: bleeding or anemia is not iron deficiency). (2) The probe studies run from iron status to enzyme activity, which is nutrient-to-drug, not depletion (Hard Rule 4).

Regulatory status

Parafon DSC label as summarized: "Rare: gastrointestinal bleeding" among adverse reactions [1]. The other label summary read did not show the adverse reaction section beyond urine discoloration [2], so this is one label version and the exact wording and causality statement were not seen.

Population evidence

No study measures iron as an outcome of chlorzoxazone. Lainé et al. 2006 (Aliment Pharmacol Ther), randomized trial, 48 participants with dysmetabolic iron overload: phlebotomy (about 3.9 L removed, ferritin below 100 µg/L) versus none; CYP2E1 activity measured by chlorzoxazone metabolism showed no significant group difference [15]. Somparn et al. 2007 (Eur J Clin Pharmacol): 35 hemoglobin E-beta thalassemia patients and 42 controls; plasma 6-hydroxychlorzoxazone to chlorzoxazone ratio used as the CYP2E1 index; serum iron, protein carbonyl and lipid peroxidation were higher and blood glutathione lower in patients, with altered CYP2E1 and CYP3A4 activity (direction not stated in the abstract read) [16]. Abstracts only.

Mechanism

For the drug-to-iron direction, none named. For iron to enzyme activity, the abstracts give no mechanism beyond association.

Counter-evidence held on record

Counter-evidence and limits The bleeding statement appears in a 2010 label summary and a causal relationship was not confirmed in anything read. The Lainé trial is null for iron removal affecting CYP2E1, and in both probe studies all participants received chlorzoxazone, so neither can say what chlorzoxazone does to iron. Framing caution Do not say chlorzoxazone lowers iron. Do not suggest iron supplements; in people with iron overload they would be the hazard. This entry is a context flag only.

Limits on what this shows

• Why no grade: Dual condition met: after the clean drug-plus-copper listing was finally delivered in the repair pass, it held no experimental or observational record in which chlorzoxazone affected copper, and no circulating claim was retrieved. In silico predictions are not graded. Mostly a genuine gap for PubMed-indexed records; the consumer-claim side stays tooling-limited because web search was unavailable.

Channels tried

PubMed listing "chlorzoxazone copper" refused (HTTP 429) twice with different parameters; "chlorzoxazone copper OR cupric OR ceruloplasmin" returned 10 records on copper and ceruloplasmin with no chlorzoxazone record (an OR query that does not isolate the drug, so a weak null) [24]. OpenAlex and Crossref search endpoints refused (429). Label texts [1][2] and drugs.com [21]: no copper content. DailyMed and web search for depletion claims: search budget exhausted. Repair pass: PubMed "chlorzoxazone AND copper" returned 4 records [27]: an electrochemical detection method using a copper sulfide electrode (PMID 35783211), a chlorophyllin (a copper-containing compound) inhibition of P450 activities including chlorzoxazone 6-hydroxylation in microsomes (PMID 7788866), and two bioinformatic cuproptosis papers (PMIDs 37435496 and 36389705) in which chlorzoxazone is listed as a database-predicted agent. The first of these, Frontiers in Pharmacology 2023, was opened (OpenAlex abstract): the chlorzoxazone prediction came from the Enrichr database screen, not from an experiment, so it is a computational prediction and not Tier B or C evidence [26]. PMID 36389705 was seen as a listing sentence only. None measures copper, ceruloplasmin or copper enzymes in chlorzoxazone-exposed cells, animals or people.

Limits on what this shows

• Why no grade: Dual condition met: no Tier B or C evidence for a drug effect on the microbiome (the retrieved rodent studies ask the reverse question), and no circulating claim retrieved. The repair-pass query with different terms confirmed the gap in PubMed-indexed records; drug-by-bacteria screening datasets and consumer claims remain unsearched, so partly tooling-limited.

Channels tried

PubMed "chlorzoxazone microbiota": 4 records, all CYP probe-cocktail studies in rodents where microbiota or probiotics were the variable and chlorzoxazone was a probe (NASH mice, antibiotic-depleted rats, probiotic-fed rats, aging mice) [24]. "chlorzoxazone intestinal bacteria" returned a 404 no-match page. "chlorzoxazone gut microbiome" and "chlorzoxazone microbiome" were refused (429). Drug-by-bacteria screens were not searched for this drug. Repair pass: PubMed "chlorzoxazone AND (dysbiosis OR 16S OR fecal OR gut bacteria OR gut flora)" returned 3 records, all with chlorzoxazone as a P450 probe or interaction example (compound probiotics in rats, PMID 39214665; rilpivirine drug interactions, PMID 23681436; gut microbiota in aging mice, PMID 41687425), none with chlorzoxazone as an exposure acting on the microbiome [27]. Read as listing text only; abstracts were not separately opened.

Closest source

Guo et al. 2021 (J Adv Res), NASH mice given six probe drugs: "no significant increase in exposure of chlorzoxazone was observed in NASH mice", with reduced clearance of other drugs tied to lower microbiota-mediated metabolism [18]. This is microbiota acting on a drug, in animals, and does not test a drug effect on the microbiome. The other three records had no abstract available to this review.

Limits on what this shows

• Key null findings: PubMed listing for chlorzoxazone AND hemolytic anemia returned 2 records: the HUS case [10] and the thalassemia probe study [16], which concerns an existing hemoglobinopathy and not a chlorzoxazone effect. Listing for chlorzoxazone AND methemoglobin returned 2 records: [9] and a cytochrome b5 regulation paper where chlorzoxazone appears only in a probe context (title only, not opened). The listing for chlorzoxazone AND hemolysis was refused (HTTP 429). No cohort, pharmacovigilance study, hemoglobin outcome or incidence figure was retrieved.
• At-risk subgroups: Not supported by retrieved data. G6PD deficiency is covered in its own ungraded entry.

Direction

Drug toxicity at most (idiosyncratic hemolysis; oxidant overdose). Not a depletion, not a retention effect and not an oxygen-delivery deficiency in the nutrient sense. Anemia and hemolysis are outcomes, not nutrients (Hard Rule 2).

Regulatory status

The label texts read, as summarized, do not mention hemolysis, anemia or methemoglobinemia; the Parafon DSC adverse reactions summary lists drowsiness, dizziness, lightheadedness, malaise, overstimulation, and rare gastrointestinal bleeding, rash, angioneurotic edema and anaphylaxis [1]. The full adverse reaction section was not seen as raw text.

Population evidence

Wang et al. 2025 (Cureus), case report, n=1: a 57-year-old woman with cognitive decline, psychomotor retardation and tremor after recent exposure to "a chlorzoxazone-containing compound"; hemolysis, thrombocytopenia and renal dysfunction, low complement C3, infectious causes and TTP excluded, "probable drug-induced HUS" by Naranjo score, recovery with supportive care; the other ingredients of the compound were not given in the abstract [10]. Ramírez Rivera and Garayúa 2009 (Bol Asoc Med P R), case report, n=1: a 46-year-old man with depression who took 30 diphenhydramine, 30 haloperidol, 20 dolagesic, 20 cyclobenzaprine, naproxen, 14 cephalexin and 48 chlorzoxazone tablets, developed cyanosis with SpO2 85% and PaO2 290.8 mm Hg at 99% SaO2, and improved after methylene blue 1 mg/kg; no single agent was blamed [9]. Both abstracts only (OpenAlex reconstructions).

Mechanism

Complement-mediated thrombotic microangiopathy suggested in [10]; the oxidant agent was not identified in [9]. Neither study tested whether chlorzoxazone itself is the oxidant or hemolytic agent.

Counter-evidence held on record

Counter-evidence and limits Both reports are confounded by co-ingestants. In [10] the exposure is a compound product and chlorzoxazone's share of the effect cannot be separated. In [9] six or more drugs were taken in overdose, including diphenhydramine, haloperidol and cyclobenzaprine, and the cited case does not attribute methemoglobinemia to chlorzoxazone. The cyclobenzaprine sibling dossier cites the same overdose case; it was opened here separately and no sibling text was relied on. Framing caution Do not present chlorzoxazone as lowering hemoglobin, iron or oxygen delivery through a nutrient mechanism. These are single-patient reports with co-exposures. No supplement is implied.

Limits on what this shows

• Why no grade: Dual condition met: no Tier B or C source on ATP synthesis for this drug and no circulating claim retrieved. The respiration and oxidative phosphorylation phrasing was delivered in the repair pass and returned nothing relevant, so this is now mostly a genuine gap in PubMed-indexed records; consumer claims remain unsearched (web search unavailable).

Channels tried

PubMed "chlorzoxazone ATP": 9 records, all either CYP2E1 probe-substrate uses or potassium-channel pharmacology (chlorzoxazone as a channel opener) [24]. "chlorzoxazone oxidative phosphorylation" and "chlorzoxazone mitochondrial respiration" refused (429). Overlaps with the mitochondrial search. Repair pass: "chlorzoxazone AND (respiration OR oxidative phosphorylation OR mitochondrial toxicity)" returned 7 records with no ATP or respiration result for chlorzoxazone as exposure [27]; "chlorzoxazone AND (bioenergetics OR ATP synthesis OR energy metabolism OR ATP content OR ATP depletion)" was read by the fetch tool as a no-match page (0 records) [27]. The Xu 2008 panel [8] scores mitochondrial damage and glutathione, not ATP, and gives no per-drug ATP result, so it is not used here.

Closest source

Ito et al. 2003 (Am J Respir Cell Mol Biol): in human airway Calu-3 cells, chlorzoxazone 1 mM and 1-EBIO 1 mM increased release of ATP into the apical compartment through intermediate-conductance Ca2+-activated K+ channel activation and cell-volume signaling [20]. This is extracellular ATP release in one cell line at a high concentration, not ATP production, so it is not used to grade the category (reasoning stated, not a hidden grade).