Phosphodiesterases (PDEs) [2]
Adenosine receptors (antagonistic effect) [2]

The bronchodilator theophylline is combined in a 2:1 ratio with ethylenediamine to form aminophylline. Aminophylline is typically found as a dihydrate, and the ethylenediamine increases solubility. Theophylline is more potent and has a longer half-life than aminophylline. It is most frequently used to treat airway obstruction brought on by COPD or asthma. Off-label, it is employed in nuclear stress testing as a reversal agent. Aminophylline is a phosphodiesterase inhibitor and nonselective antagonist of adenosine receptors. An extracellular messenger found in the body that can control myocardial oxygen requirements is adenosine. It works by blocking atrioventricular node conduction, increasing coronary artery blood flow, slowing heart rate, suppressing cardiac automaticity, and reducing the effects of β-adrenergic on contractility through cellular surface receptors that alter intracellular signaling pathways. Additionally, circulating catecholamines' chronotropic and ionotropic effects are countered by adenosine. All things considered, adenosine lowers the heart's contraction force and rate, which increases blood flow to the heart muscle. This mechanism (meant to protect the heart) may result in atropine-resistant refractory bradyasystole under certain conditions. The effects of adenosine vary with concentration. Aminophylline and other methylxanthines are competitive antagonists of adenosine receptors. Adenosine's cardiac effects are competitively inhibited by aminophylline at the cell surface receptors. It consequently raises contractility and heart rate.

The study recruited patients with acute asthma exacerbation to assess the impact of Aminophylline on arterial blood gas tensions. [2]
Patients received intravenous Aminophylline at a dosage of 5 mg/kg as an initial bolus, followed by a continuous infusion of 0.9 mg/kg per hour. [2]
Arterial blood samples were collected before drug administration and at 30-minute intervals for 2 hours after the start of treatment to measure PaO2 (arterial oxygen tension) and PaCO2 (arterial carbon dioxide tension). [2]
After administration of Aminophylline, PaO2 increased significantly from baseline (mean increase of 12.3 mmHg at 2 hours) and PaCO2 decreased significantly (mean decrease of 8.7 mmHg at 2 hours) in the enrolled patients. [2]
In comparison with isoprenaline (administered as 0.5 mg subcutaneous injection), Aminophylline produced a more prolonged improvement in blood gas homeostasis, with PaO2 and PaCO2 remaining within normal ranges for up to 2 hours post-infusion, whereas the effect of isoprenaline peaked at 30 minutes and declined thereafter. [2]

Absorption, Distribution and Excretion
0.3 to 0.7 L/kg 0.29 mL/kg/min [3-15 days after birth] 0.64 mL/kg/min [25-57 days after birth] 1.7 mL/kg/min [1-4 years] 1.6 mL/kg/min [4-12 years] 0.9 mL/kg/min [13-15 years] 1.4 mL/kg/min [16-17 years] 0.65 mL/kg/min [Adults (16-60 years), non-smoking asthmatic patients] 0.41 mL/kg/min [Elderly (>60 years)] [Liver and Kidney Function] 0.33 mL/kg/min [Acute pulmonary edema] 0.54 mL/kg/min [Chronic obstructive pulmonary disease (COPD) > 60 years old, stable condition, non-smoker > 1 year]
0.48 mL/kg/min [COPD with pulmonary heart disease]
1.25 mL/kg/min [Cystic fibrosis (14-28 years old)]
0.31 mL/kg/min [Liver disease - cholestasis]
0.35 mL/kg/min [Liver cirrhosis]
0.65 mL/kg/min [Acute hepatitis]
0.47 mL/kg/min [Sepsis with multiple organ failure]
0.38 mL/kg/min [Hypothyroidism]
0.8 mL/kg/min [Hyperthyroidism]
Intravenous theophylline can produce the highest and fastest serum theophylline concentration. In healthy adults, a single intravenous injection of approximately 5 mg/kg of theophylline (in aminophylline form) administered over 30 minutes results in a mean peak serum theophylline concentration of approximately 10 μg/mL. Approximately 50% of the theophylline dose in newborns is excreted unchanged in the urine. After three months of age, approximately 10% of the theophylline dose is excreted unchanged in the urine. /Theophylline/
Intramuscular absorption of theophylline is generally slow and incomplete. Rectal suppositories (no longer sold in the US) are absorbed slowly and inconsistently, regardless of whether the suppository matrix is hydrophilic or lipophilic. /Theophylline/
The rate-limiting step in the absorption of oral theophylline appears to be dissolution. In the acidic environment of the stomach, theophylline salts and compounds release free theophylline. Microcrystalline theophylline and oral solutions are absorbed faster than uncoated tablets, but not to a greater extent. While extended-release theophylline formulations (capsules and tablets) are absorbed more slowly, their extent of absorption is generally the same as uncoated tablets; however, the actual absorption rate of extended-release formulations may vary. Theophylline extended-release formulations are formulated to produce varying rates of drug release, suitable for administration every 8–12 hours, 12 hours, or 24 hours; however, the actual dosing frequency for a particular patient depends on their individual pharmacokinetic parameters. Because absorption rates and extent can vary between different extended-release formulations, and even between different doses of the same formulation, patients should generally be kept on a stable regimen; one extended-release formulation should only be substituted for another if the two formulations have demonstrated comparable efficacy and/or if a pharmacokinetic assessment is performed on the patient during the transition period. The presence of food may delay theophylline absorption, but generally does not reduce it; however, the effect of food on the absorption of extended-release formulations appears to vary from person to person, and therefore the manufacturer's dosing recommendations for the specific formulation should be followed. /Theophylline/
For more complete data on the absorption, distribution, and excretion of aminophylline (12 in total), please visit the HSDB record page.

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Metabolism/Metabolites
Both the N-demethylation and hydroxylation pathways of theophylline biotransformation are capacity-limited. Due to significant individual variability in the rate of theophylline metabolism, some patients may experience nonlinear elimination at serum theophylline concentrations <10 mcg/mL. Because this nonlinearity leads to disproportionate changes in serum theophylline concentration with dose, it is recommended to increase or decrease the dose in small increments to achieve the desired serum theophylline concentration change. The dose-dependent nature of theophylline metabolism cannot be accurately predicted in advance, but patients with extremely high initial clearance (i.e., lower steady-state serum theophylline concentrations at above-average doses) are most likely to experience large fluctuations in serum theophylline concentrations with dose changes. /Theophylline/
Caffeine and 3-methylxanthine are the only two pharmacologically active theophylline metabolites. 3-methylxanthine has approximately one-tenth the pharmacological activity of theophylline, with serum concentrations below 1 μg/mL in adults with normal renal function. In patients with end-stage renal disease, the accumulated concentration of 3-methylxanthine may approach that of unmetabolized theophylline. Caffeine concentrations in adults are typically undetectable regardless of renal function. In newborns, the cumulative concentration of caffeine may approach that of unmetabolized theophylline, thus producing pharmacological effects. Theophylline is metabolized in the liver to 1,3-dimethyluric acid, 1-methyluric acid, and 3-methylxanthine. …Individual rates of theophylline metabolism vary; however, individual metabolism of this drug is generally reproducible. Theophylline and its metabolites are primarily excreted by the kidneys. However, renal clearance accounts for only 8–12% of total plasma clearance of theophylline. A small amount of theophylline is excreted unchanged in feces. Theophylline is primarily metabolized via the microsomal cytochrome P450 system, particularly by the isoenzyme CYP1A2. The main metabolic pathway of theophylline is demethylation to 3-methylxanthine, and it also undergoes demethylation or oxidation to produce other metabolites. Less than 10% of theophylline is excreted unchanged in the urine. /Theophylline/
No measurable first-pass elimination occurs after oral administration of theophylline. In adults and children over one year of age, approximately 90% of the dose is metabolized in the liver. Biotransformation processes include demethylation to 1-methylxanthine and 3-methylxanthine, and hydroxylation to 1,3-dimethyluric acid. 1-Methylxanthine is further hydroxylated by xanthine oxidase to 1-methyluric acid. Approximately 6% of theophylline doses undergo N-methylation to generate caffeine. Theophylline demethylation to 3-methylxanthine is catalyzed by cytochrome P-450 1A2, while hydroxylation to 1,3-dimethyluric acid is catalyzed by cytochrome P-450 2E1 and P-450 3A3. Demethylation to 1-methylxanthine appears to be catalyzed by cytochrome P-450 1A2 or closely related cytochromes. The N-demethylation pathway is absent in newborns, and the hydroxylation pathway is also significantly underfunctional. The activity of these pathways increases slowly, reaching its maximum at 1 year of age. /Theophylline/

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Biological Half-Life
7–9 hours
In 15 infants aged 3–23 months, the clearance and half-life of theophylline were measured at least 24 hours after intravenous infusion of aminophylline. The mean half-life was 4.4 ± 2.2 hours. The half-life varied tenfold, indicating that individualization of theophylline dosage is particularly important for infants to avoid undertreatment and toxicity.

Toxicity Summary
Identification and Uses: Aminophylline is a white or slightly yellow granule or powder. It is prepared from theophylline and an aqueous solution of ethylenediamine and is used as a bronchodilator. Human Exposure and Toxicity: Aminophylline can induce seizures in patients without a known history of underlying epilepsy or without risk factors for seizure exacerbation. Most of these seizures are difficult to control, and their harmfulness is often underestimated compared to other drug toxicities. Despite the long clinical history of aminophylline-induced seizures, little is known about the underlying molecular mechanisms leading to methylxanthine-induced seizures. Adult deaths typically occur during or after intravenous administration of high doses of aminophylline in patients with renal, hepatic, or cardiovascular complications. In other patients, the rate of injection, rather than the dose, appears to be a more significant factor in inducing acute hypotension, seizures, coma, cardiac arrest, ventricular fibrillation, and death. Therefore, intravenous administration of aminophylline or theophylline should be slow. Childhood deaths are usually due to overdose and significant sensitivity to theophylline's central nervous system stimulation. Some reports on aminophylline hypersensitivity reactions show that most cases in English literature are delayed-type reactions. However, most Japanese cases are immediate-type reactions. Acetylation is the main metabolic pathway of ethylenediamine. On the other hand, most Japanese are rapid or moderate acetylated individuals. Caucasians have a 50% probability of being slow acetylated individuals. This difference suggests that the incidence of moderate- and delayed-type reactions to aminophylline hypersensitivity differs between Japanese and Caucasians. Aminophylline treatment may be related to elevated cardiac enzyme levels. In vitro experiments show that aminophylline protects MRC-5 cells from apoptosis by inhibiting the activity of caspase 3 and caspase 8. Animal experiments: Aminophylline (100-250 mg/kg) can sustainably induce seizures and post-seizure death in mice, while conventional anticonvulsants and adenosine receptor agonists are not effectively antagonistic to these effects. Biochemical analysis of brain homogenates showed that aminophylline-induced epilepsy was associated with elevated levels of malondialdehyde and nitric oxide metabolites in brain tissue, while superoxide dismutase activity was decreased. Pretreatment with melatonin and L-NAME mitigated these changes. Aminophylline induced seizures in rats in a dose-dependent manner, with the incidence of seizures and death reaching its maximum at a dose of 300 mg/kg, and a significant increase in free radical production. Antioxidant pretreatment showed varying degrees of attenuation of aminophylline-induced free radical production, but had little antagonistic effect on aminophylline-induced seizures and post-epileptic mortality. In pregnant rabbits, aminophylline treatment did not accelerate overall lung anatomy, as evidenced by the alveolar volume to lung tissue weight ratio. Under similar experimental conditions, caffeine treatment in doe rabbits had no effect on the fetal lungs. In another experiment, pregnant doe rabbits were intravenously injected with aminophylline (6 mg/kg/day) starting on day 25 post-mating, and delivered the fetus via uterine slit on day 28. One group of newborn rabbits breathed air, while the other group breathed 100% oxygen. Lung mechanics was assessed during spontaneous or artificial respiration in the newborn rabbits, and histological studies of the lungs were performed, with particular attention to alveolar volume density. The aminophylline-treated group showed increased body weight, improved survival rate, and elevated phosphatidylglycerol levels in the bronchoalveolar lavage fluid. Animals in the aminophylline-treated group breathing air showed increased respiratory rate, but lung compliance data indicated no significant difference between the treatment and control groups. The conclusion is that the beneficial effects of aminophylline are primarily attributed to the combined effects of accelerating fetal growth and improving postnatal respiratory regulation, rather than specific effects on lung biochemistry and functional maturation. Aminophylline exacerbates seizure damage in the developing brain of rats.

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Use during pregnancy and lactation
◉ Overview of use during lactation
Expert opinion considers theophylline use during lactation acceptable. Maternal use of theophylline may sometimes cause excitement, irritability, and restless sleep in infants. Newborns, especially premature infants, are most susceptible due to slow theophylline clearance and low serum protein binding. There is no need to avoid theophylline; however, maternal serum theophylline concentrations should be maintained at the lower limit of the therapeutic range, and the infant should be monitored for theophylline side effects. Infant serum theophylline concentrations help determine whether irritability is caused by theophylline. Avoiding breastfeeding for 2 hours after intravenous theophylline injection or 4 hours after oral immediate-release theophylline can reduce the dose ingested by breastfed infants. When using oral sustained-release theophylline preparations, the timing of breastfeeding is not significantly related to the timing of administration or offers no benefit.
◉ Effects on Breastfed Infants
A 3-day-old breastfed infant experienced irritability and restlessness while the mother was taking 200 mg of aminophylline every 6 hours. These effects disappeared after discontinuation but recurred upon re-administration over the next 9 months. These effects were likely caused by theophylline in breast milk. Five other infants reported in this study did not experience adverse reactions after their mothers took theophylline. Because newborns and preterm infants clear theophylline more slowly, the accumulation of theophylline in infant serum appears to be most likely to occur in them.
◉ Effects on Lactation and Breast Milk
As of the revision date, no relevant published information was found.
◉ Summary of Medication Use During Lactation
The expert panel considers the use of aminophylline during lactation to be acceptable. Maternal use of aminophylline may sometimes cause excitement, irritability, and restlessness in infants. Newborns, especially premature infants, are most susceptible due to their slower theophylline clearance and lower serum protein binding rates. There is no need to avoid the use of aminophylline products; however, maternal serum theophylline concentrations should be maintained at the lower limit of the therapeutic range, and the infant should be monitored for theophylline side effects. Infant serum theophylline concentrations can help determine whether the agitation is caused by theophylline. Avoiding breastfeeding within 2 hours after intravenous administration of aminophylline or within 4 hours after oral immediate-release aminophylline can reduce the dose ingested by breastfed infants.
◉ Effects on Breastfed Infants
A 3-day-old breastfed infant experienced irritability and restlessness while the mother was taking 200 mg of aminophylline every 6 hours. These symptoms disappeared after discontinuation of the drug, but recurred upon re-administration within the next 9 months. These symptoms may be caused by theophylline in breast milk. Five other infants reported in this study did not experience adverse reactions after their mothers took theophylline. Theophylline accumulation in infant serum appears to be most likely to occur in newborns and preterm infants, as they clear theophylline more slowly.
◉ Effects on Lactation and Breast Milk
As of the revision date, no relevant published information was found.

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Protein Binding
60%

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Interaction
This study aimed to describe the pharmacodynamic interaction between propofol and aminophylline. Nine beagle dogs were randomly assigned to three propofol infusion groups at infusion rates of 0.75 (Group A), 1.00 (Group B), and 1.25 (Group C) mg/kg/min. In the first phase, propofol was administered only; in the second phase, aminophylline was administered only, at infusion rates of 0.69 (Group A), 1.37 (Group B), and 2.62 (Group C) mg/kg/hr, respectively. In phases three through five, both drugs were administered concurrently. The infusion rates of aminophylline were 0.69 (Phase 3), 1.37 (Phase 4), and 2.62 (Phase 5) mg/kg/hr, respectively. The infusion time for aminophylline was 0–30 hours, and for propofol, it was 24 hours over 20 minutes. Blood samples and electroencephalograms (EEGs) were collected at pre-defined time intervals. The linear regression slope between the logarithmically transformed dose of aminophylline and AUC_inf was 0.6976 (95% CI 0.5242–0.8710). The pharmacokinetics of aminophylline best conformed to a one-compartment model (including enzyme autoinduction). The pharmacokinetics and pharmacodynamics of propofol best fit the three-compartment model and the S-type Emax model, respectively. The estimated pharmacodynamic parameters for propofol are: k(e0) = 0.805/min, E0 = 0.76, Emax = 0.398, Ce(50 na) = 2.38 μg/mL (without exposure to aminophylline), C(e50 wa) = 4.49 μg/mL (with exposure to aminophylline), γ = 2.21. The potency of propofol is reduced when used concomitantly with aminophylline. The pharmacodynamic antagonism between aminophylline and propofol's sedative effects may not be dose-dependent, but rather an all-or-nothing response. Animal studies have shown that concomitant use of β-adrenergic agonists (e.g., isoproterenol) and theophylline derivatives (e.g., aminophylline) may increase cardiotoxicity. Although this interaction has not been confirmed in humans, some reports suggest that this combination may have the potential to induce arrhythmias. Further clinical data are needed to determine if this potential interaction exists in humans. Theophylline can interact with a variety of drugs. This interaction may be pharmacodynamic, meaning it alters the therapeutic response to theophylline or other drugs, or causes adverse reactions without changing serum theophylline concentrations. However, more common are pharmacokinetic interactions, where theophylline clearance is affected by other drugs, leading to increases or decreases in serum theophylline concentrations. Theophylline rarely alters the pharmacokinetics of other drugs. The drugs listed in the table may have clinically significant pharmacodynamic or pharmacokinetic interactions with theophylline. The information in the "Effects" column of the table assumes that the interacting drug was added to a theophylline steady-state treatment regimen. If a patient is taking drugs that inhibit theophylline clearance (e.g., cimetidine, erythromycin), a lower dose of theophylline will be required to achieve therapeutic serum theophylline concentrations when starting theophylline. Conversely, if a patient is taking drugs that promote theophylline clearance (e.g., rifampin), a higher dose of theophylline will be required to achieve therapeutic serum theophylline concentrations when starting theophylline. Discontinuing concomitant medications that promote theophylline clearance can lead to theophylline accumulation at potentially toxic levels unless the theophylline dose is appropriately reduced. Discontinuing concomitant medications that inhibit theophylline clearance can lead to decreased serum theophylline concentrations unless the theophylline dose is appropriately increased. /Theophylline/
Table: Clinically Significant Drug Interactions of Theophylline [Table #844]

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Non-human Toxicity Values
Mice Intravenous LD50 146 mg/kg
Mice Subcutaneous LD50 186 mg/kg
Mice Intraperitoneal LD50 217 mg/kg
Mice Oral LD50 150 mg/kg
For more complete non-human toxicity data for aminophylline (9 types), please visit the HSDB records page.

[1]. Luteolin, a non-selective competitive inhibitor of phosphodiesterases 1-5, displaced [3H]-rolipram from high-affinity rolipram binding sites and reversed xylazine/ketamine-induced anesthesia. ur J Pharmacol. 2010 Feb 10;627(1-3):269-75.

[2]. Response of blood gas tensions to aminophylline and isoprenaline in patients with asthma. Thorax. 1967 Nov;22(6):543-9.

[3]. Adenosine receptors: development of selective agonists and antagonists. Prog Clin Biol Res. 1987;230:41-63.

Therapeutic Uses
Bronchodilator; Cardiotonic agent; Phosphodiesterase inhibitor; Purinergic P1 receptor antagonist. ClinicalTrials.gov is a registry and results database that lists human clinical studies funded by public and private institutions worldwide. The website is maintained by the National Library of Medicine (NLM) and the National Institutes of Health (NIH). Each record on ClinicalTrials.gov includes a summary of the study protocol, including: the disease or condition; the intervention (e.g., the medical product, behavior, or procedure under investigation); the title, description, and design of the study; participation requirements (eligibility criteria); the location of the study; contact information for the study location; and links to other relevant health websites, such as the NLM's MedlinePlus (for patient health information) and PubMed (for citations and abstracts of academic articles in the medical field). Aminophylline is listed in the database. Intravenous theophylline (usually in the form of aminophylline) has been used to relieve periodic apnea in patients with Cheyne-Schönlein respiration and to increase arterial blood pH. /Not included in US product label/
Veterinary Drugs: Aminophylline is indicated for the control of reversible airway constriction, prevention of bronchoconstriction, and as an adjunct therapy for the treatment of other respiratory diseases. Because it is a salt form of theophylline, its uses are similar to those of theophylline. It is used to treat inflammatory airway diseases in cats (feline asthma), dogs, and horses. In dogs, its uses include tracheal collapse, bronchitis, and other airway diseases. It is ineffective for respiratory diseases in cattle.
For more complete data on the therapeutic uses of aminophylline (10 in total), please visit the HSDB record page.
Drug Warnings

Adult death commonly occurs during or after intravenous administration of high doses of aminophylline in patients with renal, hepatic, or cardiovascular complications. In other patients, the rate of administration, rather than the dose, appears to be a more significant factor in causing acute hypotension, seizures, coma, cardiac arrest, ventricular fibrillation, and death. Therefore, intravenous administration of aminophylline or theophylline should be slow. Childhood death is usually due to overdose and significant sensitivity to the central nervous system stimulation of theophylline.
When administered rectally as a suppository (a dosage form no longer sold in the United States), theophylline can cause rectal irritation and inflammation. Rapid intravenous injection of aminophylline may cause dizziness, fainting, lightheadedness, palpitations, syncope, precordial pain, flushing, severe bradycardia, ventricular premature beats (VPC, PVC), severe hypotension, or cardiac arrest. Intramuscular injection of aminophylline can cause severe local pain and tissue shedding…
Theophylline may also cause transient urinary frequency, dehydration, finger and hand twitching, shortness of breath, and elevated serum AST (SGOT) levels. Hypersensitivity reactions characterized by urticaria, generalized pruritus, and angioedema have been reported after aminophylline administration. Other reports indicate that allergy to the ethylenediamine component of aminophylline can cause contact dermatitis. There are also reports of bone marrow suppression, leukopenia, thrombocytopenia, and bleeding tendency after taking theophylline, but the association between these adverse reactions and theophylline treatment remains questionable. Other adverse reactions of theophylline include proteinuria, increased excretion of renal tubular cells and red blood cells in urine, hyperglycemia, and syndrome of inappropriate antidiuretic hormone secretion (SIADH). /Theophylline/
For more drug warnings (complete) data on aminophylline (22 in total), please visit the HSDB record page.

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Pharmacodynamics
Aminophylline is the ethylenediamine salt of theophylline. Theophylline can stimulate the central nervous system, skeletal muscle, and myocardium. It can relax certain smooth muscles in the bronchi, produce a diuretic effect, and increase gastric juice secretion.

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Aminophylline is a compound preparation of theophylline and ethylenediamine, which can improve the solubility of theophylline, making it easier to administer parenterally. [2]
Its mechanism of action in treating asthma is mainly twofold: one is to inhibit phosphodiesterase, thereby increasing the level of intracellular cyclic adenosine monophosphate (cAMP) and relaxing bronchial smooth muscle; the other is to antagonize adenosine receptors, thereby preventing adenosine-induced bronchoconstriction. [2]
At the time of the study (1967), aminophylline was a first-line drug for treating acute asthma attacks due to its ability to rapidly dilate the bronchi and improve gas exchange in the lungs. [2]

C16H24N10O4

420.43

420.198

317-34-0

58-55-9 (free);317-34-0 (EDA);

9433

White to off-white solid powder

454.1ºC at 760mmHg

269-270 °C

4

8

1

30

273

0

FQPFAHBPWDRTLU-UHFFFAOYSA-N

InChI=1S/2C7H8N4O2.C2H8N2/c2*1-10-5-4(8-3-9-5)6(12)11(2)7(10)13;3-1-2-4/h2*3H,1-2H3,(H,8,9);1-4H2

1,3-dimethyl-3,7-dihydro-1H-purine-2,6-dione compound with ethane-1,2-diamine (2:1)

Cardophyllin; Aminophyllin; Aminophylline; Theophyllamine; Phyllocontin; Euphyllin; Truphylline; Minomal R 175 mg tab; Minomal R 350 mg tab; Minomal SR 600 mg tab

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 (e.g. under nitrogen), avoid exposure to moisture and light.

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

Solubility in Formulation 1: 1.43 mg/mL (3.40 mM) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 14.3 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: ≥ 1.43 mg/mL (3.40 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (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 14.3 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: ≥ 1.43 mg/mL (3.40 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 14.3 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 4: 22 mg/mL (52.33 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication.

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