Macrolide
Ribosome (large ribosomal subunit, near the peptidyltransferase center and peptide exit tunnel) (Ki = 5.5 nM) [1]
The primary target of josamycin is the 50S large subunit of the bacterial ribosome. By reversibly binding to the 50S subunit, josamycin blocks the elongation of nascent peptides through the ribosomal exit tunnel, thereby inhibiting bacterial protein synthesis. Compared to 14-membered ring macrolides such as erythromycin, josamycin exhibits a significantly longer average residence time on the ribosome (approximately 3 hours versus less than 2 minutes for erythromycin), with a dissociation constant Kd of 5.5 nM for ribosome binding. This compound completely shuts down full-length protein synthesis at saturating drug concentrations, a mechanistic difference that may explain its more potent bactericidal effect.

According to studies, josamycin has an average lifetime of 3 hours on the ribosome and a dissociation constant of 5.5 nM when it binds to the ribosome. Josamycin completely prevents the synthesis of full-length proteins at saturating drug concentrations by completely inhibiting the formation of the second or third peptide bond, depending on the peptide sequence. Josamycin also slows down the formation of the first peptide bond of a nascent peptide in an amino acid-dependent manner. Josamycin completely inhibits the synthesis of full-length proteins at a saturating drug concentration[1].

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As a 16-membered lactone ring macrolide antibiotic, josamycin inhibits peptide elongation on the ribosome by binding close to the peptidyltransferase center and blocking the peptide exit tunnel in the large ribosomal subunit [1]
- In a cell-free mRNA translation system with pure components from Escherichia coli, the dissociation constant (Ki) for josamycin binding to the ribosome is 5.5 nM, and its average lifetime on the ribosome is 3 hours [1]
- Josamycin slows down the formation of the first peptide bond of a nascent peptide in an amino acid-dependent manner and completely inhibits the formation of the second or third peptide bond, depending on the peptide sequence [1]
- Josamycin stimulates the rate constant for drop-off of peptidyl-tRNA from the ribosome, and this drop-off rate is much faster than the drug dissociation rate [1]
- At a saturating concentration, josamycin completely shuts down the synthesis of full-length proteins [1]

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In vitro studies demonstrate that josamycin exhibits good antibacterial activity against various Gram-positive cocci. The minimum inhibitory concentrations against pneumococci, Streptococcus pyogenes, Staphylococcus aureus, and Staphylococcus epidermidis are comparable to those of erythromycin and clindamycin. Josamycin shows superior activity against enterococci compared to clindamycin. Against anaerobes, josamycin demonstrates potent in vitro activity against various anaerobes including Bacteroides fragilis. This antibiotic is concentrated up to 20-fold in phagocytic cells. Josamycin-pretreated human neutrophils exhibit approximately 425-460% enhanced killing capacity against Staphylococcus aureus. Additionally, josamycin enhances the bactericidal activity of oxidative stress against S. aureus, with greater efficacy than erythromycin.

After giving rabbits 200 mg/kg of josamycin orally, the drug is present in their blood and tissues. In general, tissue levels are significantly higher than blood levels, and they are also somewhat higher three hours after the dose than they are one hour later, when the blood level is at its lowest. The level in the lungs is the highest of all tissue levels one hour after the medication[2].

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In vivo studies confirm that josamycin exhibits significant efficacy in infected animal models. In a rabbit syphiloma model, although josamycin showed lower efficacy than penicillin, it still demonstrated sufficient anti-Treponema pallidum activity. In clinical application involving 19 patients with pyoderma, josamycin achieved an effective rate of 63.2%, with particularly satisfactory results in furuncles and follicular infections; only one patient reported epigastric burning sensation. Josamycin also enhances whole blood bactericidal function, with significantly increased killing capacity against Pseudomonas aeruginosa at a concentration of 10 mg/L.

0.5 mM calcium chloride, 5 mM magnesium acetate, 5 mM ammonium chloride, 95 mM potassium chloride, 8 mM putrescine, 1 mM spermidine, 5 mM potassium phosphate, and 1 mM dithioerythritol are the ingredients used to prepare josamycin in polymix buffer. Preinitiated ribosomes are treated with varying concentrations of josamycin (2, 3, 4, and 6 μM) to initiate the incubation process. At each incubation period, one volume of the reaction mix is mixed with one volume of the elongation mix. The reaction is then quenched with formic acid after ten seconds. The fraction of ribosomes that form tripeptides is used to estimate the association rates[1].

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Ribosome binding and peptide elongation inhibition assay: Prepare a cell-free mRNA translation system using pure components isolated from Escherichia coli. Dilute josamycin to appropriate concentrations and add it to the reaction system containing ribosomes, mRNA, aminoacyl-tRNAs, and other necessary translation factors. Incubate the system under optimal conditions for protein synthesis. Monitor the formation of peptide bonds (e.g., by detecting radioactive-labeled peptides or using other peptide bond detection methods) to analyze the effect of josamycin on the rate of peptide bond formation. Determine the dissociation constant (Ki) of josamycin binding to the ribosome through quantitative analysis of binding affinity. Measure the lifetime of josamycin on the ribosome and the rate constant of peptidyl-tRNA drop-off using kinetic detection techniques [1]

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Ribosome binding studies for josamycin can be conducted using cell-free translation systems: Reaction System Construction: Construct a cell-free mRNA translation system using purified components from E. coli, including 70S ribosomes, aminoacyl-tRNA synthetases, initiation factors, and elongation factors. Drug Treatment: Pre-incubate varying concentrations of josamycin (0.5-100 nM) with ribosomes for 10-30 minutes. Peptide Synthesis Detection: Add radiolabeled amino acids (e.g., [³⁵S]-methionine) or polypeptide templates, incubate at 37°C, and detect radioactivity of newly synthesized peptide chains by scintillation counting. Dissociation Kinetics Determination: Monitor the dissociation rate of the josamycin-ribosome complex using excess unlabeled drug or rapid dilution methods. Results show that josamycin has an average residence time of 3 hours on the ribosome. Data Analysis: Calculate the dissociation constant (Kd = 5.5 nM) and binding occupancy of the drug with the ribosome.

Cell Culture: Isolate and purify human peripheral blood neutrophils from healthy volunteer blood by Ficoll-Hypaque density gradient centrifugation. Drug Treatment: Pre-incubate adherent neutrophils with josamycin (0.1-25 mg/L) for 30-60 minutes, with a vehicle control group. Antibacterial Activity Assay: Add S. aureus (approximately 1-5×10⁶ CFU/mL) at a bacteria-to-cell ratio of approximately 10:1, and incubate at 37°C for 60-120 minutes. Bacterial Killing Detection: Lyse neutrophils with saponin to release engulfed bacteria, plate on agar plates for colony counting, and calculate killing rates. Phagocytosis Activity Detection: Assess neutrophil phagocytic capacity using microscopy or flow cytometry. Results show that josamycin-pretreated PMNs exhibit 425-460% increased killing capacity against S. aureus. Chemotaxis Detection: Evaluate the effect of josamycin on neutrophil chemotactic function using the Boyden chamber method with fMLP as the chemoattractant.

Rat: Rats are given 200 mg/kg of oral Josamycin labeled with tritium. Bioassay is used to determine the blood and tissue levels at one hour and three hours[2].
Mouse: Mice are given 200 mg/kg of josamycin labeled with tritium orally. Bioassay is used to determine the blood and tissue levels at one hour and three hours[2].

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Animal Models: Skin infection model: Use Japanese white rabbits to establish syphiloma models by intradermal injection of Treponema pallidum. Systemic infection model: Use mice to establish systemic infection models by intraperitoneal injection of S. aureus suspension. Dosing Regimen: Administer josamycin by oral gavage. Rabbit models receive 50-100 mg/kg daily for 7-14 days; mouse model doses are calculated based on body weight. Efficacy Assessment: Syphiloma model: Observe changes in lesion size and collect lesion tissue for dark-field microscopy to detect spirochetes. Systemic infection model: Record animal survival rates and collect spleen, kidney, and other tissues for bacterial enumeration. Clinical Observation: Record daily animal general status, behavior, food intake, and body weight changes, and observe injection site reactions. Data Analysis: Compare differences in lesion regression time, bacterial clearance rates, and survival rates between treatment and control groups.

Josamycin is rapidly absorbed after oral administration, reaching peak plasma concentrations approximately 1 hour after a 1 g oral dose in healthy volunteers. This drug has at least 15-fold higher lipophilicity than erythromycin, with a plasma protein binding rate of approximately 15%, significantly lower than other macrolide antibiotics. Josamycin exhibits high accumulation in lung tissue, with lung tissue concentrations 2-3 times higher than plasma concentrations, reaching a peak concentration of 3.68 μg/g at 2-3 hours. In a study of 21 patients, josamycin concentrations in lung tissue consistently reached the minimum inhibitory concentrations required for treating infections. This drug can be concentrated up to 20-fold in phagocytic cells, suggesting advantages for targeted delivery to infection sites.

Josamycin, as a protein synthesis inhibitor, is itself toxic to humans[2] - It is hepatotoxic, which hinders efforts to increase its blood concentrations[2] - Intravenous josamycin is associated with a higher incidence of serious side effects[2] - Oral administration allows for a delicate balance between tissue affinity and metabolism, thereby minimizing side effects while ensuring efficacy[2]

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Toxicological studies of josamycin demonstrate a favorable safety profile at therapeutic doses. In long-term carcinogenicity studies in rodents, josamycin showed no positive tumorigenic results, with no tumorigenicity observed in either male or female rats. Long-term clinical data in humans indicate that the common adverse reactions to josamycin are primarily gastrointestinal, including nausea, abdominal discomfort, and diarrhea, with relatively low incidence rates. In a clinical observation of 19 patients with skin infections, only one patient reported epigastric burning sensation, with no other notable side effects. Compared to other macrolide antibiotics, josamycin has a lower risk of drug interactions, with weaker inhibition of CYP3A4. The drug is contraindicated in patients with known hypersensitivity and should be used with caution in those with severe hepatic impairment. Store protected from light in sealed containers.

[1]. Kinetics of macrolide action: the Josamycin and erythromycin cases. J Biol Chem. 2004 Dec 17;279(51):53506-15.

[2]. Pharmacokinetics of macrolides, lincosamides and streptogramins. J Antimicrob Chemother. 1985 Jul;16 Suppl A:151-66.

Josamycin is a macrolide antibiotic produced by certain strains of Streptomyces narbonensis var. josamyceticus. It is both an antibacterial drug and a metabolite. It is a macrolide antibiotic, and also an aldehyde, tertiary amine, tertiary alcohol, acetate, disaccharide derivative, and glycoside compound.
A macrolide antibiotic produced by Streptomyces narbonensis. This drug has antibacterial activity against a variety of pathogens.
A macrolide antibiotic produced by Streptomyces narbonensis. This drug has antibacterial activity against a variety of pathogens.

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Drug Indications
Used to treat bacterial infections.

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Mechanism of Action
The mechanism of action of macrolide antibiotics (such as josamycin) is through reversible binding to the 50S subunit of bacterial ribosomes, inhibiting bacterial protein biosynthesis, thereby inhibiting the translocation of peptidyl-tRNA. This action is mainly bacteriostatic, but at high concentrations it may also have bactericidal activity. Macrolide antibiotics tend to accumulate in leukocytes, thus enabling them to be transported to the site of infection.

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Pharmacodynamics
Josamycin is a macrolide antibiotic derived from Streptomyces. This drug has antibacterial activity against a variety of pathogens.

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Josamycin is a macrolide antibiotic containing a 16-membered lactone ring[1]
- The bactericidal effect of josamycin is thought to be caused by two factors: inhibition of protein elongation, and depletion of the intracellular pool of aminoacyl-tRNA available for protein synthesis due to peptidyl-tRNA shedding and incomplete peptidyl-tRNA hydrolase activity[1]
- For macrolide antibiotics like josamycin, their in vivo therapeutic effect cannot be assessed solely by blood concentration and minimum inhibitory concentration (MIC), which differs from the conventional rules of antibiotic evaluation[2]

C42H69NO15

827.99496

827.466

C, 60.92; H, 8.40; N, 1.69; O, 28.98

16846-24-5

31674-19-8 (Josamycin propionate); 11033-19-5 (Josamycin HCl); 4564-87-8 (Carbomycin); 35775-82-7 (Maridomycin); 20283-69-6 (niddamycin); 1404-48-4 (relomycin);35834-26-5 (rosaramicin); 497-72-3 (methymycin); 30042-37-6 (lankamycin )

5282165

Solid powder

1.2±0.1 g/cm3

877.8±65.0 °C at 760 mmHg

131.5℃

484.7±34.3 °C

0.0±0.6 mmHg at 25°C

1.535

3.88

3

16

14

58

1390

16

O[C@H]1[C@](O[C@H](C)[C@@H](O[C@@](O[C@@H](C)[C@@H]2OC(CC(C)C)=O)([H])C[C@]2(O)C)[C@@H]1N(C)C)([H])O[C@@H]([C@H](C[C@H]([C@H](/C=C/C=C/C3)O)C)CC=O)[C@H]([C@](CC(O[C@@H]3C)=O)([H])OC(C)=O)OC

XJSFLOJWULLJQS-DJBGMQKKSA-N

InChI=1S/C42H69NO15/c1-23(2)19-32(47)56-40-27(6)53-34(22-42(40,8)50)57-37-26(5)54-41(36(49)35(37)43(9)10)58-38-29(17-18-44)20-24(3)30(46)16-14-12-13-15-25(4)52-33(48)21-31(39(38)51-11)55-28(7)45/h12-14,16,18,23-27,29-31,34-41,46,49-50H,15,17,19-22H2,1-11H3/b13-12-,16-14+/t24-,25-,26-,27+,29+,30+,31-,34+,35-,36-,37-,38+,39+,40+,41+,42-/m1/s1

Leucomycin V, 3-acetate 4(sup B)-(3-methylbutanoate)

Josamycin; BRN-1677122; CCRIS 8511; josamycin; Leucomycin A3; Turimycin A5; Kitasamycin A3; 16846-24-5; BRN 1677122; BRN1677122; Josamycin hydrochloride; Leucomycin A3 hydrochloride;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

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

DMSO : 41.67~100 mg/mL (50.33~120.77 mM)
H2O : 8.33 mg/mL (10.06 mM)
Ethanol : ~100 mg/mL

Solubility in Formulation 1: 2.5 mg/mL (3.02 mM) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with sonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: 2.5 mg/mL (3.02 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
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.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (3.02 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 4: 10% DMSO+40% PEG300+5% Tween-80+45% Saline: 2.5 mg/mL (3.02 mM)

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