Bacterial DNA, through intercalation and induction of mutations in the bacterial genome. The primary mechanism of carbadox's antibacterial activity is the induction of DNA damage, which consequently triggers the bacterial SOS response and, notably, the lytic cycle of prophages within the bacterial genome. Carbadox inhibits DNA synthesis and can also break existing DNA, exerting a bactericidal effect.

The MTT assay results demonstrated that in Vero cells, mitochondrial activity decreased in a dose-dependent manner across all Carbadox doses. When Carbadox was used at its maximum dose of 160 μg/mL, the percentage of viable cells decreased to just 12%. DNA migration increased in cells treated with carbadox in a dose-dependent manner (p<0.01). The mitotic index (NDI) dramatically drops as carbadox dosage rises [1].
Carbadox exerts its antibacterial activity by inhibiting bacterial DNA synthesis and protein synthesis. The primary mechanism is the induction of DNA damage, which triggers the bacterial SOS response and the lytic cycle of prophages within the bacterial genome. Carbadox inhibits DNA synthesis and can also break existing DNA, exerting a bactericidal effect. The compound is most effective against Gram-positive bacteria and under anaerobic conditions, with high activity against Clostridium spp. and Brachyspira spp.

The medicated piglet samples' alpha diversity (Shannon diversity, Heips evenness, and inverse Simpson index) at 2, 3, and 4 days following continuous Carbadox differed considerably from those of the unmedicated piglets, but not at the late Carbadox or at any other time. same thing. period of drug withdrawal. There was no significant difference in the animals' prior antibiotic therapy (p=0.82). However, analysis of the animals' bacterial community structure revealed significant alterations on days 3 and 4 of early Carbadox administration ([R=0.32, p=0.015] and [R=0.54, p=0.003], respectively). E did not differ significantly. Colony-forming units (CFU) of E. Coli were detected either late in the study's withdrawal period or during the Carbadox treatment period. After drug discontinuation, E changed significantly on the second day. coli CFU in comparison to the non-drug group in the drug group [2].
In vivo, carbadox is used as a growth-promoting and antibacterial drug in swine feed to prevent dysentery and improve feed efficiency. It is effective against bacterial pathogens that cause swine dysentery and proliferative enteropathy. The compound is used in animals up to 4 months of age with a 4-week withdrawal period to ensure that drug residues are cleared before slaughter. Its use is regulated in many countries due to concerns about potential carcinogenicity.

In vitro susceptibility testing for carbadox is performed using standard broth microdilution or agar dilution methods against bacterial isolates. Minimum inhibitory concentrations (MICs) are determined for various bacterial species. Carbadox shows activity against S. typhimurium strains with MICs of 3.6-14.3 µM under aerobic conditions. The compound's mechanism of action can be studied by measuring DNA damage or SOS response induction in bacterial cultures.

In vitro cellular assays for carbadox involve treating bacterial cells with the compound and measuring DNA damage, SOS response induction, or cell viability. The compound's ability to inhibit DNA synthesis and cause bactericidal effects is demonstrated in these assays. Carbadox is most effective against Gram-positive bacteria and under anaerobic conditions.

In vivo animal studies for carbadox typically involve administration to swine through feed at a rate of 50 ppm. Efficacy is assessed by measuring the reduction in bacterial infections (such as swine dysentery) and improvement in feed efficiency and weight gain. Withdrawal period studies are conducted to determine the time required for drug residues to decline to safe levels (4 weeks).

Absorption, Distribution and Excretion
High-performance liquid chromatography (HPLC) was used to determine the concentration of quinoxaloline-2-carboxylic acid (QCA) in porcine liver and muscle after alkaline hydrolysis. The QCA concentration range in liver was <3 ng/g to 45.3 ng/g, and in muscle it was <3 ng/g to 10.8 ng/g. QCA was detected in liver samples (9.7 ng/g) after 77 days of treatment. The recoveries of QCA in liver and muscle were 70% at 5 ng/g, 77% and 75% at 10 ng/g, respectively, and 90% at 30 ng/g. This experiment was conducted within the framework of the National Surveillance Program for Animal Tissue Residues in the Republic of Croatia. Concentrations of Cabardos and its first metabolite, deoxycabados, in the gastrointestinal contents of pigs were determined after the addition of therapeutic doses (100–150 ppm) of cabardos to the feed. The results showed that the concentration of cabardosin in the gastrointestinal tract was lower than the reported minimum inhibitory concentration (MIC) values for sites of action of enteropathogenic microorganisms. Therefore, these observations do not support the pharmacological basis for using cabardosin to treat swine dysentery and diarrhea. However, the cabardosin levels detected in the proximal intestine (stomach and duodenum) appear to suggest that feed supplementation with 50 ppm cabardosin can effectively prevent Treponema pallidum (the pathogen of swine dysentery). This article discusses the changes in blood and deoxycabachosin concentrations and the concentration distribution in the gastrointestinal tract after feed supplementation with cabardosin (50 ppm), and explores the distribution of this drug in pigs. The elimination of cabardosin was investigated in mice, pigs, and monkeys. Pigs were fed a diet containing 50 g/t of unlabeled cabardos for several weeks before receiving 3.5 mg/kg of (14)C-cabbardos, while mice and monkeys received a single dose of 5 mg/kg of (14)C-cabbardos. Urine and fecal samples were collected and their radioactivity was determined. Urinary metabolites were qualitatively analyzed using thin-layer chromatography (TLC). Almost all metabolites present in pig urine (13 out of 15) were also present in rat and monkey urine. Of the other two metabolites, one was a glycine conjugate of quinoxaline-2-carboxylic acid. During the 72-hour collection period, all animals excreted more than 50% of the dose via urine (pigs: 74%, monkeys: 61%, rats: 54%). The radioactivity in feces over 72 hours was: pigs 17%, monkeys 8-10%, and rats 29%. The total excretion rate over 72 hours appeared to be between 70-90%. The authors concluded that the radioactivity distribution was similar across the three species.

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Metabolism/Metabolites
Seven-week-old pigs were fed unlabeled carbadox at a rate of 50 g/t for several weeks, followed by a single oral administration of carbonyl-labeled 14C-carbadox via gastric tube. The following assessments were performed: detection of 14CO2 in exhaled breath, detection of the labeled substance in liver and urine to determine its association with methylcarbazine, and detection of free hydrazine in plasma and urine. Peak plasma concentrations of the radiolabeled substance occurred at approximately 3 hours. While early plasma concentrations were similar to those of ring-labeled carbadox, concentrations remained slightly higher after 24 hours. At 3 hours, approximately 50% of the radiolabeled substance in plasma was identified as carbadox, compared to an estimated 30% for methylcarbazine. The primary route of excretion of the radiolabeled substance was urine. However, of the total amount recovered from ring-labeled carbadox, less than half of that recovered from carbonyl-labeled carbadox was recovered (37% vs 88%). This is likely due to a higher conversion rate of the radiolabeled carbadox to carbon dioxide (up to 36% in rats). Five days later, radiolabeled carbadox equivalents of 0.1–0.34 ppm were detected in the liver. The authors inferred that a portion of this was present as carbon dioxide (possibly 25%). Pigs receiving a 7 mg/kg dose excreted 7% of the dose in the urine as free hydrazine after 24 hours, while no identifiable hydrazine was detected in the urine of pigs receiving lower doses. Seven-week-old pigs were fed a diet containing 50 g/t of unlabeled carbadox for several weeks, followed by a single oral dose of 3.5 mg/kg or 0.8 mg/kg of benzene ring-labeled (14)C-carbadox. A peak in plasma was observed approximately 3 hours after administration. Five to eight hours after administration, the following substances were detected in plasma: carbadox (13%), deoxycarbadox (9-19%), carbadox aldehyde (13%), and quinoxaloline-2-carboxylic acid (19%) (all values are expressed as total plasma radioactivity). Carbadox aldehyde was also confirmed in gastric contents. Carbadox is rapidly excreted from the body. Approximately two-thirds of the dose is excreted in the urine within 48-72 hours, with the remainder excreted in the feces (approximately 90% in total). Fourteen days after administration, the residual radioactive material in the liver was equivalent to approximately 0.1 ppm of carbadox. Attempts to identify these residual radioactive materials were unsuccessful. Twenty-four hours after administration, the only metabolite identified in the liver was quinoxaloline-2-carboxylic acid, a metabolite primarily excreted in the urine.
Pharmacokinetic studies of carbadox in swine have been performed using both ring-labeled and carbonyl-labeled 14C-carbadox. Studies with carbonyl-labeled carbadox demonstrate that methylcarbazate is generated. Probably as much as 25% is excreted in urine in the form of methylcarbazate derivatives. Carbadox stimulates the renin-angiotensin system and suppresses aldosterone production by inducing morphologic changes in adrenal cortex (plasma Na↓ and K↑).

Preclinical toxicity studies have raised concerns about the carcinogenicity of carbadox. The compound's mechanism of action involves DNA damage, which is consistent with its potential genotoxicity and carcinogenicity. Due to these concerns, the use of carbadox is regulated in many countries, and it is only approved for use in swine up to 4 months of age with a 4-week withdrawal period.

[1]. Investigation of the genotoxicity of quinocetone, carbadox and olaquindox in vitro using Vero cells. Food Chem Toxicol. 2009 Feb;47(2):328-34.

[2]. Carbadox has both temporary and lasting effects on the swine gut microbiota. Front Microbiol. 2014 Jun 10;5:276.

Carbadox is an antimicrobial agent used in pigs. It is a mutagenic/carcinogenic agent and has been banned in Canada, Australia, and the European Union. Carbadox was previously used in veterinary clinical practice to treat swine dysentery and enteritis and to promote growth. However, due to reports of carcinogenicity and mutagenicity, it has been banned in the UK. (Excerpt from Martindale Pharmacopoeia, 30th edition, p. 125) See also: Carbadox; Oxytetracycline (ingredient); Carbadox; Praziquantel tartrate (ingredient).
Carbadox is a quinoxaline-di-N-oxide antimicrobial agent used in veterinary medicine as a growth promoter and antibacterial agent in swine feed. It is used to prevent swine dysentery and improve feed efficiency. The compound's mechanism of action involves DNA damage and induction of the bacterial SOS response. Its use is regulated in many countries due to concerns about potential carcinogenicity. Carbadox remains an important tool in swine production in some regions, but its use is increasingly restricted.

C11H9N4O4

261.21

262.07

6804-07-5

Carbadox-d3;1185240-06-5

135403805

Light yellow to yellow solid powder

1.447g/cm3

405.47°C (rough estimate)

239-240ºC

18°(64°F)

1.648

1.777

1

5

3

19

352

0

COC(=O)N/N=C/C1=[N+](C2=CC=CC=C2[N+](=C1)[O-])[O-]

OVGGLBAWFMIPPY-WUXMJOGZSA-N

InChI=1S/C11H10N4O4/c1-19-11(16)13-12-6-8-7-14(17)9-4-2-3-5-10(9)15(8)18/h2-7H,1H3,(H,13,16)/b12-6+

methyl N-[(E)-(1,4-dioxidoquinoxaline-1,4-diium-2-yl)methylideneamino]carbamate

Getroxel; Fortigro; Carbadoxum

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~3.57 mg/mL (~13.61 mM)
H2O : < 0.1 mg/mL

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)