(1→3)-β-D-glucan synthase

A sharp reduction of the metabolic activity of cells within the biofilm as assessed by the XTT reduction assay was demonstrated when preformed C. albicans 3153A biofilms were exposed to caspofungin (Fig.1). By this method, the 48-h MIC50 of caspofungin for sessile C. albicans 3153A cells within biofilms was 0.0625 μg/ml. Although complete sterility of biofilms was not achieved by treatment with caspofungin, the experiments showed a >97% reduction in the metabolic activity of sessile cells with caspofungin concentrations as low as 0.125 μg/ml. Caspofungin was also active against biofilms formed by all the C. albicans clinical isolates tested (n = 18), with MIC50s for sessile cells ranging between 0.0625 and 0.125 μg/ml, compared to fluconazole MIC50s for sessile cells of ≥64 μg/ml for all isolates. In agreement with the XTT assays, only residual metabolic activity was detected in cells within the caspofungin-treated biofilms, which showed a diffuse green fluorescence pattern characteristic of dead cells (Fig.3B). In confirmation of the SEM results, CLSM demonstrated that caspofungin treatment resulted in biofilms that were less hyphal and also showed minor distortions of the overall biofilm architecture. As shown in Fig.4, coating with caspofungin resulted in significant (up to 60%) reduction of the metabolic activity of adherent cells compared to that of cells in untreated (control) wells. Together these findings indicate that caspofungin displays potent activity against C. albicans biofilms in vitro and merits further investigation for the treatment of biofilm-associated infections. [3]

Caspofungin (1-8 mg/kg; i.p.; daily for 7 days) enters the central nervous system of mice and reaches concentrations that diminish Candida burden in the brain [1]. Caspofungin (0.41-41 μM; i.p.; 5 weeks; male C57BL/6 mice) is a safe antifungal drug with mouse vitreous concentrations ranging from 0.41 to 4.1 μM [2].

The echinocandin MK-0991, formerly L-743,872, is a water-soluble lipopeptide that has been demonstrated in preclinical studies to have potent activity against Candida spp., Aspergillus fumigatus, and Pneumocystis carinii. An extensive in vitro biological evaluation of MK-0991 was performed to better define the potential activities of this novel compound. Susceptibility testing with MK-0991 against approximately 200 clinical isolates of Candida, Cryptococcus neoformans, and Aspergillus isolates was conducted to determine MICs and minimum fungicidal concentrations MF(s). The MFC at which 90% of isolates are inhibited for 40 C. albicans clinical isolates was 0.5 microg/ml. Susceptibility testing with panels of antifungal agent-resistant species of Candida and C. neoformans isolates indicated that the MK-0991 MFCs for these isolates are comparable to those obtained for susceptible isolates. Growth kinetic studies of MK-0991 against Candida albicans and Candida tropicalis isolates showed that the compound exhibited fungicidal activity (i.e., a 99% reduction in viability) within 3 to 7 h at concentrations ranging from 0.06 to 1 microg/ml (0.25 to 4 times the MIC). Drug combination studies with MK-0991 plus amphotericin B found that this combination was not antagonistic against C. albicans, C. neoformans, or A. fumigatus in vitro. Studies with 0 to 50% pooled human or mouse serum established that fungal susceptibility to MK-0991 was not significantly influenced by the presence of human or mouse serum. Results from resistance induction studies suggested that the susceptibility of C. albicans was not altered by repeated exposure (40 passages) to MK-0991. Erythrocyte hemolysis studies with MK-0991 with washed and unwashed human or mouse erythrocytes indicated minimal hemolytic potential with this compound. These favorable results of preclinical studies support further studies with MK-0991 with humans.[4]

Effect of coating the wells of a microtiter plate with caspofungin on C. albicans biofilm formation. A modified assay was used in which the wells of a microtiter plate were directly precoated with caspofungin in order to investigate the drug's ability to prevent biofilm formation. Briefly, 200-μl volumes of caspofungin at different concentrations in sterile PBS were added to selected wells of a microtiter plate and incubated overnight at 4°C. After incubation, excess caspofungin was aspirated and the plates were washed once in sterile PBS. C. albicans 3153A cells were washed in PBS and resuspended at a concentration of 106 cells per ml in RPMI 1640. The 96-well microtiter plates were then seeded with the suspension (100 μl per well) and incubated for 24 h at 37°C to allow biofilm formation. The contents of the wells were aspirated and washed three times in sterile PBS, and the extent of biofilm formation was assessed by the XTT reduction assay and by light microscopy. The inhibitory effect of caspofungin was expressed as the percentage of the optical density (OD) of caspofungin-treated wells compared to that of control (plastic) wells for the XTT assays. Statistical analysis was performed with Student's t test. P values of <0.05 were considered statistically significant. The analyses were performed by using Prism version 3.00 for Window.[3]

Animal/Disease Models: DBA/2N mice deficient in complement component 5 [1]
Doses: 1, 2, 4 and 8 mg/kg
Route of Administration: intraperitoneal (ip) injection; one time/day for 7 days
Experimental Results: diminished concentration of Candida load in the brain.

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Animal/Disease Models: Male C57BL/6 mice [2] Doses: 0.41, 1.2, 2.5, 4.1 and 41 μM
Route of Administration: intraperitoneal (ip) injection; continued for 5 weeks
Experimental Results: ERG waveform changed from 0.41 μM to 4.1 μM, no significant change .

Absorption, Distribution and Excretion
Following intravenous infusion, 92% of the drug is distributed in tissues within 36–48 hours. After a single intravenous injection of [3H] caspofungin acetate, the excretion of caspofungin and its metabolites in the human body is as follows: 35% in feces and 41% in urine. 12 mL/min [after a single intravenous administration] Elimination routes: Feces: 35% excreted as drug or metabolites. Kidneys: 41% excreted as drug (approximately 1.4% unchanged) or metabolites. Dialysis: Not removed by hemodialysis. Following a single 70 mg dose of radiation, approximately 92% of the radioactive material is distributed in tissues within 36–48 hours. Caspofungin has very low distribution in erythrocytes. Caspofungin can cross the placenta in rats and rabbits and is detectable in fetal plasma of pregnant animals administered caspofungin. Caspofungin is distributed in rat milk; it is unclear whether caspofungin is distributed in human milk. For more complete data on absorption, distribution, and excretion of caspofungin (13 items in total), please visit the HSDB record page. Metabolism/Metabolites Primarily metabolized slowly via hydrolysis and N-acetylation. Caspofungin also undergoes spontaneous chemical degradation, further hydrolyzing into its constituent amino acids and degradation products, including dihydroxyhomotyrosine and N-acetylated dihydroxyhomotyrosine. Caspofungin is slowly metabolized in the liver via hydrolysis and N-acetylation; following a single intravenous injection of the radiolabeled drug, 35% and 41% of the parent drug and metabolites, respectively, are excreted in feces and urine. This study investigated the metabolism, excretion, and pharmacokinetics of caspofungin following a single intravenous injection in mice, rats, rabbits, and monkeys. …Radiation excretion was slow in all studied species, with low levels of radioactivity detected in daily urine and fecal samples over a long collection period. Although urine analysis revealed the presence of multiple metabolites (M0, M1, M2, M3, M4, M5, and M6), most radioactivity was associated with the polar metabolites M1 [4(S)-hydroxy-4-(4-hydroxyphenyl)-L-threonine] and M2 [N-acetyl-4(S)-hydroxy-4-(4-hydroxyphenyl)-L-threonine]. Therefore, caspofungin is primarily eliminated through metabolic transformation; however, its metabolic rate is slow. …
Caspofungin is slowly metabolized through hydrolysis and N-acetylation. Caspofungin also spontaneously degrades into the open-ring peptide compound L-747969. At later time points after administration (≥5 days), the covalent binding level of the radiolabeled substance in plasma was low (≤7 picomoles/mg protein, or ≤1.3% of the administered dose) following a single injection of (3)H caspofungin acetate, likely due to two active intermediates formed during the chemical degradation of caspofungin to L-747969. Furthermore, caspofungin is also hydrolyzed to generate constituent amino acids and their degradation products, including dihydroxyhigh-tyrosine and N-acetyldihydroxyhigh-tyrosine. These two tyrosine derivatives are only present in urine, suggesting rapid renal clearance. Caspofungin acetate… After intravenous infusion of 70 mg (3)HCaspofungin acetate in healthy subjects, excretion of drug-related substances was very slow, with 41% and 35% of the administered radioactive substances recovered from urine and feces, respectively, within 27 days. Plasma and urine samples collected approximately 24 hours after administration contained primarily unmetabolized caspofungin acetate, along with trace amounts of the peptide hydrolysis product M0 (a linear peptide). However, at subsequent sampling time points, M0 became the main circulating component, and the corresponding urine samples mainly contained hydrolytic metabolites M1 and M2, as well as M0 and unmetabolized MK-0991. These substances accounted for 13%, 71%, 1%, and 9% of the total urinary radioactive material excreted in the first 16 days after administration, respectively. The main metabolite M2 is highly polar and extremely unstable under acidic conditions, converting into a less polar product, which was identified as N-acetyl-4(S)-hydroxy-4-(4-hydroxyphenyl)-L-threonine γ-lactone. After derivatization in an aqueous medium, M2 was identified as the corresponding γ-hydroxy acid, namely N-acetyl-4(S)-hydroxy-4-(4-hydroxyphenyl)-L-threonine. Metabolite M1 is extremely polar and eluted after the dead volume of the HPLC column, and was identified as deacetyl-M2 by chemical derivatization. Therefore, the major urinary and plasma metabolites of MK-0991 originate from peptide hydrolysis and/or N-acetylation. /Caspofungin Acetate/

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Biological Half-Life
9–11 hours
Initial: 9 to 11 hours (β phase). Supplementation: 40 to 50 hours (γ phase).
A long terminal elimination half-life (11.7 to 59.7 hours) was observed in all preclinical animals following a single intravenous injection of caspofungin.

Effects During Pregnancy and Lactation
◉ Overview of Use During Lactation There is currently no information regarding the use of caspofungin during lactation. Due to its high plasma protein binding rate (up to 97%) and low oral bioavailability, caspofungin is unlikely to enter breast milk and be absorbed by the infant. Caspofungin can be safely administered intravenously to infants 3 months and older. The amount absorbed into breast milk is likely to be far lower than the infant's dose. If the mother needs to use caspofungin, this is not a reason to discontinue breastfeeding. ◉ Effects on Breastfed Infants As of the revision date, no relevant published information was found. ◉ Effects on Lactation and Breast Milk As of the revision date, no relevant published information was found. Protein Binding 97% Interaction …This study investigated the efficacy of caspofungin and meropenem, alone and in combination, in treating mice with disseminated candidiasis. Immunomodulatory mice were infected with 2 × 10⁶ CFU of Candida albicans via intravenous injection. Intraperitoneal administration began 24 hours post-infection and continued for 7 days. Treatment groups included those receiving caspofungin (0.5, 1.25, and 5 mg/kg/day), meropenem (20 mg/kg/day), and a combination of both drugs. Renal colony-forming unit (CFU) counts showed lower residual bacterial loads in mice receiving the combination therapy. Caspofungin was effective at doses of 0.5, 1.25, and 5 mg/kg compared to the untreated infection control group. In vitro studies showed that the minimum inhibitory concentrations (MICs) of caspofungin and meropenem were <0.075 μg/mL and >64 μg/mL, respectively. A synergistic effect was observed with the combination therapy. Histopathological examination showed a 25% reduction in inflammation and a smaller area of renal tubular necrosis in the combination therapy group compared to the monotherapy group. These results suggest that combination therapy with caspofungin and meropenem may be beneficial.
Concomitant use with tacrolimus may lead to a decrease in tacrolimus plasma concentrations; monitoring of tacrolimus concentrations is recommended, and dose adjustment should be made as necessary.
Potential pharmacokinetic interactions (decreased caspofungin plasma concentrations). Concomitant use of caspofungin with drug clearance inducers or mixed inducers/inhibitors (such as efavirenz, nelfinavir, nevirapine, phenytoin sodium, rifampin, dexamethasone, or carbamazepine) may result in a clinically significant decrease in caspofungin plasma concentrations. …
Two parallel-group studies evaluated the potential interactions between caspofungin and nelfinavir or rifampin. In Study A, healthy subjects received caspofungin monotherapy (50 mg, once daily intravenously) for 14 days (n = 10), or in combination with nelfinavir (1250 mg, twice daily orally) (n = 9), or in combination with rifampin (600 mg, once daily orally) (n = 10). In Study B, 14 subjects received rifampin (600 mg orally once daily) for 28 days, combined with caspofungin (50 mg intravenously once daily) for the last 14 days; another 12 subjects received caspofungin monotherapy (50 mg intravenously once daily) for 14 days. The geometric mean ratios of the areas under the curve [AUC(0–24)] within 24 hours after caspofungin administration (values in parentheses are 90% confidence intervals [CI]) were as follows: 1.08 (0.93–1.26) in the nelfinavir group, 1.12 (0.97–1.30) in the rifampin group (Study A), and 1.01 (0.91–1.11) in the rifampin group (Study B). Rifampin altered the shape of the caspofungin plasma concentration curve, resulting in a 14%–31% decrease in the trough concentration (C(24h)) at 24 hours after administration, consistent with the net induction effect at steady state. In Study A, both AUC and C(24hr) increased initially with rifampin in combination therapy (61% and 170% increases, respectively, on day 1), but no increase was observed in Study B, consistent with the transient net inhibition before complete induction. The geometric mean ratio of rifampin AUC(0–24) for combination therapy/monotherapy on day 14 was 1.07 (90% CI, 0.83–1.38). Nefernavir had no significant effect on the pharmacokinetics of caspofungin. Rifampin both inhibits and induces the metabolism of caspofungin, leading to a decrease in C(24hr) at steady state. When caspofungin is used in combination with rifampin, increasing the caspofungin dose to 70 mg daily should be considered. For more complete data on interactions of caspofungin (9 drugs), please visit the HSDB records page.

[1]. Flattery AM, et, al. Efficacy of caspofungin in a juvenile mouse model of central nervous system candidiasis. Antimicrob Agents Chemother. 2011 Jul;55(7):3491-7.
[2]. Mojumder DK, et, al. Evaluating retinal toxicity of intravitreal caspofungin in the mouse eye. Invest Ophthalmol Vis Sci. 2010 Nov;51(11):5796-803.
[3]. Antimicrob Agents Chemother. 2002 Nov; 46(11): 3591–3596.
[4]. Antimicrob Agents Chemother.1997 Nov;41(11):2326-32

Caspofungin (global brand name: Cancidas) is an antifungal drug and the first member of the echinocandin class named by Merck. It is usually administered intravenously. Caspofungin is effective against Aspergillus and Candida infections, and its mechanism of action is the inhibition of β(1,3)-D-glucan synthesis in the fungal cell wall. Caspofungin is an antifungal echinocandin lipopeptide, semi-synthesized from the fermentation products of Glarea lozoyensis. Caspofungin inhibits 1,3-β-glucan synthase, leading to reduced synthesis of β(1,3)-D-glucan (an important component of the fungal cell wall), thereby weakening the fungal cell wall and ultimately causing its rupture. It is effective against Aspergillus and Candida fungi. A cyclic lipopeptide echinocandin and β-(1,3)-D-glucan synthase inhibitor used to treat visceral or systemic fungal infections.
See also: Caspofungin (note moved here).

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Drug Indications
For the treatment of esophageal candidiasis and invasive aspergillosis in patients who are unresponsive to or intolerant of other therapies.
FDA Label
For the treatment of invasive candidiasis in adults or children; for the treatment of invasive aspergillosis in adults or children who are unresponsive to or intolerant of amphotericin B, liposome amphotericin B, and/or itraconazole. Unresponsive is defined as infection progression or failure to improve after at least 7 days of effective antifungal therapy; empirical treatment of suspected fungal infection (such as Candida or Aspergillus) in adults or children with fever and neutropenia.
For the treatment of invasive candidiasis in adults or children. For the treatment of invasive aspergillosis in adults or children who are resistant to or intolerant of amphotericin B, liposome amphotericin B, and/or itraconazole. Resistance is defined as infection progression or failure to improve after at least 7 days of effective antifungal therapy. This is used for empirical treatment of fever and neutropenia in adult or pediatric patients with suspected fungal infections (such as Candida or Aspergillus).

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Mechanism of Action
Caspofungin inhibits the synthesis of β-(1,3)-D-glucan, an important component of the cell walls of Aspergillus and Candida. β-(1,3)-D-glucan is absent in mammalian cells.
The primary target is β-(1,3)-glucan synthase.
Caspofungin inhibits the synthesis of β-(1,3)-D-glucan, an important component of fungal cell walls, which is absent in mammalian cells.
Caspofungin acetate… belongs to the echinocandins class of drugs, which inhibit the formation of β-(1,3)-D-glucan in fungal cell walls. Mutations in the FKS1 gene, which encodes the large subunit of β-(1,3)-glucan synthase, lead to drug resistance. Caspofungin acetate is the active ingredient of caspofungin, which inhibits the synthesis of α-(1,3)-D-glucan, an important component of the cell walls of susceptible Aspergillus and Candida species. β-(1,3)-D-glucan is not present in mammalian cells. Caspofungin has been shown to have activity against Candida species, as well as activity against the active growth regions of Aspergillus fumigatus hyphae. /Caspofungin Acetate/

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Therapeutic Use
Caspofungin is indicated for the empirical treatment of suspected fungal infections in patients with fever and neutropenia. /US Product Label Contains/
Caspofungin is indicated for the treatment of candidemia and the following candidiasis infections: esophageal, intra-abdominal and abscess, peritonitis, and pleural cavity infections. /Included in US Product Label/
Caspofungin is indicated for the treatment of patients with invasive aspergillosis who are unresponsive to or intolerant of other therapies, including amphotericin B (liposomal and non-liposomal formulations) and/or itraconazole. /Included in US Product Label/
/Expl Ther/ ... Azole-resistant Candida albicans isolates remain sensitive to caspofungin…Caspofungin acetate is effective against Candida albicans, Aspergillus fumigatus, Pneumocystis carinii, and Histoplasma capsulatum infections in laboratory animals. Clinical trials of intravenous caspofungin are currently underway…for the treatment of deep candidiasis, neutropenia, and fever in patients unresponsive to antimicrobial therapy. /Caspofungin Acetate/
For more complete data on the therapeutic uses of caspofungin (8 formulations), please visit the HSDB record page.

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Drug Warning
In an open-label, uncontrolled clinical study, adverse reactions occurred in 2% or more of patients with invasive aspergillosis treated with caspofungin acetate, including fever, intravenous infusion complications, nausea, vomiting, or flushing. Adverse reactions reported in clinical studies for uses other than aspergillosis include fever, phlebitis/thrombophlebitis, intravenous infusion complications, headache, nausea, pain (unspecified), rash, anemia, abdominal pain, diarrhea, vomiting, facial edema, flu-like symptoms, myalgia, paresthesia, induration, chills, and pruritus.
Post-marketing reported adverse events include: Hepatobiliary system: rare cases of clinically significant hepatic dysfunction; Cardiovascular system: swelling and peripheral edema; Metabolic system: hypercalcemia.
Symptoms possibly mediated by histamine have been reported, including rash, facial swelling, pruritus, fever, or bronchospasm. Hypersensitivity reactions have been reported during CANCIDAS use.
Caspofungin is distributed into breast milk in rats; it is unknown whether caspofungin is distributed into breast milk in humans. Caution should be exercised when using caspofungin during lactation.
For more complete data on drug warnings for caspofungin (6 of 6), please visit the HSDB record page.

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Pharmacodynamics
Caspofungin is an antifungal drug belonging to the echinocandins class. It is used to treat Aspergillus and Candida infections, and its mechanism of action is the inhibition of cell wall synthesis. Echinocandins inhibit the synthesis of glucan in the cell wall by inhibiting 1,3-β-glucan synthase. Although the possibility of resistance exists, in vitro resistance to caspofungin in Aspergillus has not been studied.

C52H88N10O15

1093.31

1091.65

C, 57.13; H, 8.11; N, 12.81; O, 21.95

162808-62-0

Caspofungin diacetate;179463-17-3;Caspofungin-d4;1131958-73-0; 162808-62-0

2826718

Typically exists as solid at room temperature

1.36g/cm3

1408.1ºC at 760mmHg

805.4ºC

0mmHg at 25°C

1.623

0.761

16

18

23

77

1900

14

CCC(CC(CCCCCCCCC(NC1CC(O)C(NC(C2C(O)CCN2C(C(NC(C(NC(C3CC(O)CN3C(C(NC1=O)C(O)C)=O)=O)C(O)C(O)C4=CC=C(O)C=C4)=O)C(O)CCN)=O)=O)NCCN)=O)C)C

JYIKNQVWKBUSNH-WVDDFWQHSA-N

InChI=1S/C52H88N10O15/c1-5-28(2)24-29(3)12-10-8-6-7-9-11-13-39(69)56-34-26-38(68)46(55-22-21-54)60-50(75)43-37(67)19-23-61(43)52(77)41(36(66)18-20-53)58-49(74)42(45(71)44(70)31-14-16-32(64)17-15-31)59-48(73)35-25-33(65)27-62(35)51(76)40(30(4)63)57-47(34)72/h14-17,28-30,33-38,40-46,55,63-68,70-71H,5-13,18-27,53-54H2,1-4H3,(H,56,69)(H,57,72)(H,58,74)(H,59,73)(H,60,75)/t28-,29+,30+,33+,34-,35-,36+,37-,38+,40-,41-,42-,43-,44-,45-,46-/m0/s1

(10R,12S)-N-((2R,6S,9S,11R,12S,14aS,15S,20S,23S,25aS)-20-((R)-3-amino-1-hydroxypropyl)-12-((2-aminoethyl)amino)-23-((1S,2S)-1,2-dihydroxy-2-(4-hydroxyphenyl)ethyl)-2,11,15-trihydroxy-6-((R)-1-hydroxyethyl)-5,8,14,19,22,25-hexaoxotetracosahydro-1H-dipyrrolo[2,1-c:2',1'-l][1,4,7,10,13,16]hexaazacyclohenicosin-9-yl)-10,12-dimethyltetradecanamide

L 743872; MK0991; L743872; MK 0991; L-743872; MK-0991

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.)