DNA gyrase; topoisomerase IV; Quinolone antibiotic

Nalidixic acid has the ability to combat a wide range of microorganisms, including Shigella spp., Brucella spp., Escherichia coli, Pasteurella spp., Klebsiella pneuiioniae, Aerobacter aeroyenes, Proteus spp., Salmonella spp., and Brucella spp. The MIC values of these microorganisms are as follows: 5.0-12.5 μg/ml, 0.5-2.5 μg/ml, 0.8-25.0 μg/ml, 1.25-30.0 μg/ml, 8-3.2 μg/ml, and 7.5-10.0 μg/ml, respectively[1].

Gram-negative bacteria are the target of nalidixic acid's greatest in vivo action, whilst Gram-positive species are typically more resilient. Maximum efficacy against E-caused systemic infections is noted. Coli, A. The ED50 values for Shigella fkxneri, Proteus mirabilis, and Aerobacter are, respectively, 25 mg/kg, 60 mg/kg, 50 mg/kg, and 62 mg/kg[1]. After being administered orally and parenterally to mice, the acute toxicity (LD50) of nalidixic acid is 3300 mg/kg orally, 176 mg/kg intravenously, and 500 mg/kg subcutaneously[1].

Fluoroquinolones are an important class of wide-spectrum antibacterial agents. The first quinolone described was nalidixic acid, which showed a narrow spectrum of activity. The evolution of quinolones to more potent molecules was based on changes at positions 1, 6, 7 and 8 of the chemical structure of nalidixic acid. Quinolones inhibit DNA gyrase and topoisomerase IV activities, two enzymes essential for bacteria viability. The acquisition of quinolone resistance is frequently related to (i) chromosomal mutations such as those in the genes encoding the A and B subunits of the protein targets (gyrA, gyrB, parC and parE), or mutations causing reduced drug accumulation, either by a decreased uptake or by an increased efflux, and (ii) quinolone resistance genes associated with plasmids have been also described, i.e. the qnr gene that encodes a pentapeptide, which blocks the action of quinolones on the DNA gyrase and topoisomerase IV; the aac(6')-Ib-cr gene that encodes an acetylase that modifies the amino group of the piperazin ring of the fluoroquinolones and efflux pump encoded by the qepA gene that decreases intracellular drug levels. These plasmid-mediated mechanisms of resistance confer low levels of resistance but provide a favourable background in which selection of additional chromosomally encoded quinolone resistance mechanisms can occur.[2]

Effects of nalidixic acid and its derivatives were investigated on mouse cells transformed by methylcholanthrene or an activated c-Ha-ras oncogene. Our findings were as follows. Nalidixic acid preferentially suppressed growth in soft agar of transformed Balb/3T3 mouse cells induced by methylcholanthrene. The suppressive effect of nalidixic acid on growth in soft agar was reversible. Nalidixic acid reversibly reduced saturation density of these transformed cells. Oxolinic acid and pipemidic acid, which are derivatives of nalidixic acid, were less effective than nalidixic acid in suppressing growth in soft agar. Nalidixic acid suppressed growth in soft agar of NIH/3T3 mouse cells transformed by an activated c-Ha-ras, without affecting the amount of ras p21 proteins as detected by an immunoblotting analysis using a monoclonal antibody. These results show that nalidixic acid reversibly suppressed the expression of transformed phenotypes that were already being expressed.https://pubmed.ncbi.nlm.nih.gov/2690912/

Absorption
After oral administration, nalidixic acid is rapidly absorbed from the gastrointestinal tract, with a bioavailability of approximately 96%. Co-administration with antacids may delay absorption.
Excretion
After oral administration, nalidixic acid is rapidly absorbed from the gastrointestinal tract, partially metabolized in the liver, and rapidly excreted via the kidneys. Approximately 4% of nalidixic acid is excreted in feces.
Compared to adults, neonates have lower absorption and excretion rates of nalidixic acid, reaching adult levels only around 3 years of age. However, the relative volumes of distribution are similar between the two age groups.
In rats and mice, the oral dose is rapidly absorbed, reaching peak plasma concentrations approximately 1 hour later. …The drug is primarily excreted via the kidneys, reaching peak concentrations approximately 6 hours later. 80% of the administered dose is excreted within the first 8 hours. In dogs, highly effective concentrations of the drug are detectable in urine within 2–3 hours after oral administration.
In patients with Shigella infection, both the absorption efficiency and excretion rate of nalidixic acid are decreased. In young patients with pronounced diarrhea, malabsorption is commonly observed, but there is no reasonable explanation for the delayed excretion. It is rapidly and almost completely absorbed from the gastrointestinal tract; bioavailability is approximately 96%. Co-administration with antacids may delay absorption. Metabolism/Metabolites: Hepatic metabolism. 30% of the administered dose is metabolized to the active metabolite hydroxynadiric acid. The parent drug and the active metabolite rapidly combine to form an inactive metabolite. Metabolism may vary considerably between individuals. In urine, hydroxynadiric acid accounts for 80% to 85% of antibacterial activity. When ingested, hydroxynadiric acid is partially excreted as a free acid, but the majority is excreted as a monoglucuronide, a significant portion as a 7-hydroxymethyl metabolite, and a small amount as a conjugate. 3,7-Dicarboxylic acid is a minor metabolite. In the liver, hydroxynadiric acid is partially metabolized to hydroxynadiric acid and its glucuronic acid conjugate. This drug is also partially metabolized into a dicarboxylic acid derivative; there is evidence that this metabolite forms in the kidneys.

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Biological Half-Life
The biological half-life in healthy adult patients is 1.1 to 2.5 hours, while in patients with impaired renal function, the biological half-life can reach 21 hours.
Approximately 96% is absorbed after oral administration. Plasma concentrations can reach 20-50 μg/mL, but 93-97% of the drug is bound to plasma proteins. In the body, some of the drug is converted to active hydroxynalidixic acid, both of which are excreted in the urine. Most of the drug undergoes conjugation in the liver. The plasma half-life is 8 hours, but in patients with renal failure, the plasma half-life can reach 21 hours.

Medication Use During Pregnancy and Lactation
◉ Overview of Medication Use During Lactation
Limited information suggests that when a mother takes no more than 2 grams of nalidixic acid daily, the low concentration of the drug in breast milk usually does not have any adverse effects on breastfed infants. However, close monitoring of the infant's gut microbiota is necessary, for example, in cases of diarrhea or candidiasis (thrush, diaper rash). For infants with glucose-6-phosphate dehydrogenase (G6PD) deficiency, nalidixic acid should be avoided during breastfeeding. The use of other medications is recommended, especially in breastfed newborns or premature infants.
◉ Effects on Breastfed Infants
A 16-day-old infant presented with slow weight gain, pallor, and jaundice, possibly due to hemolytic anemia caused by the mother's oral administration of nalidixic acid (1 gram, four times daily) and amebarbital (65 mg, three times daily). The infant developed jaundice, hyperbilirubinemia, reticulocytosis, eosinophilia, Heinz bodies, and other signs of hemolysis 7 days after the mother began taking nalidixic acid. No G-6-PD deficiency or Zurich hemoglobin was observed.
◉ Effects on lactation and breast milk
As of the revision date, no relevant published information was found.

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Interactions
…at physiological concentrations…nalidixic acid…displaces a significant amount of warfarin from human serum albumin via a non-competitive mechanism.
Systemic and urinary alkalizers reduce their efficacy by increasing their excretion rate. Systemic efficacy is enhanced if urine is acidic.
An 18-year-old male developed metabolic acidosis due to an overdose of nalidixic acid. Concomitant use of probenecid may enhance the effects of the ingested nalidixic acid by prolonging its serum half-life.
Coumarin or indanedione derivative anticoagulants, especially warfarin and dicumarol, may be displaced from their protein binding sites by nalidixic acid, thereby enhancing their anticoagulant effect; dosage adjustments may be necessary during and after nalidixic acid treatment.

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Antidotes and Emergency Treatment
Recommended treatment methods include: Reducing absorption—Gastric lavage should be performed if an overdose is detected early. Targeted treatment—Anticonvulsants should be administered if treatment for seizures is required. Supportive treatment—If absorption occurs, intravenous fluids and supportive measures, such as oxygen and artificial respiration, should be given. If a patient is known or suspected of intentionally overdosing on the medication, they should be referred to a psychiatrist for consultation.

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Human Toxicity Excerpt
Acute toxicity of nalidixic acid may manifest as toxic psychosis, seizures, increased intracranial pressure, or metabolic acidosis. Vomiting, nausea, and drowsiness may also occur. Due to the rapid excretion of nalidixic acid, these reactions are usually short-lived, lasting only 2-3 hours.
Systemic Effects in Humans: Children may experience seizures, hyperglycemia, sweating, and blood changes.

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Non-Human Toxicity Excerpt
Nalidixic acid can cause lameness in puppies due to permanent damage to the cartilage of their weight-bearing joints.
…Prolonged use of this drug (in dogs and cats) can lead to retinal degeneration, and in some cases, even blindness.

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Non-human toxicity values
Oral LD50 in mice: 3.3 g/kg
Subcutaneous LD50 in mice: 0.5 g/kg
Intravenous LD50 in mice: 0.176 g/kg
Oral LD50 in rats: 1160 mg/kg

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Protein binding rate
The protein binding rate of nalidixic acid in blood is 93%, and the protein binding rate of its active metabolite, hydroxynalidixic acid, is 63%.

[1]. 1,8-NAPHTHYRIDINE DERIVATIVES. A NEW CLASS OF CHEMOTHERAPEUTIC AGENTS. J Med Pharm Chem. 1962 Sep:5:1063-5.

[2]. Mechanism of Action of and Resistance to Quinolones. Microb Biotechnol.2009 Jan;2(1):40-61.

A synthetic 1,8-naphthidine antibacterial agent with a limited bactericidal spectrum. It is an inhibitor of bacterial DNA gyrase A subunit.

C12H11N2NAO3

254.22

254.067

C, 56.70; H, 4.36; N, 11.02; Na, 9.04; O, 18.88

3374-05-8

Nalidixic acid;389-08-2

3864541

White to off-white solid powder

1.331g/cm3

413.1ºC at 760mmHg

229-230ºC

203.6ºC

0.088

0

5

2

18

384

0

ROKRAUFZFDQWLE-UHFFFAOYSA-M

InChI=1S/C12H12N2O3.Na/c1-3-14-6-9(12(16)17)10(15)8-5-4-7(2)13-11(8)14;/h4-6H,3H2,1-2H3,(H,16,17);/q;+1/p-1

sodium;1-ethyl-7-methyl-4-oxo-1,8-naphthyridine-3-carboxylate

Nalidixic acid sodium salt; 3374-05-8; Nalidixic acid sodium; Baktogram; Sodium nalidixate; Nalidixate sodium anhydrous; NALIDIXATE SODIUM; Nalidixate (sodium);

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)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

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.)