NP213 targets the fungal cytoplasmic membrane. Its mechanism of action involves membrane perturbation, disruption, and permeabilization, leading to cell lysis. [1]
As a cationic antimicrobial peptide, it targets the fungal cell membrane regardless of the metabolic activity level of the fungi. [2]
The primary target of Novexatin is the fungal cytoplasmic membrane. As a cationic antimicrobial peptide (AMP), Novexatin acts by interacting with the fungal cell membrane and is effective regardless of the metabolic activity level of the fungi. The compound exerts its antifungal activity through membrane perturbation, disruption, and permeabilization, ultimately leading to fungal cell lysis. Notably, unlike conventional antifungal agents such as azoles, Novexatin does not rely on enzyme inhibition for its action, and this unique mechanism makes it less prone to inducing drug resistance.

The number of PI-stained T cells is increased by NP213 (500–1000 μg/mL; 18 hours). cells of rubrum NCPF0118 in specimens. This finding suggests that NP213 has a bactericidal effect, and membrane permeabilization is the mechanism of action [1]. The minimum inhibitory concentration (MIC) of NP213 against Trichophyton rubrum NCPF0118 was shown to vary according on the source of keratin. In 1640 medium containing human skin keratin, human nail keratin, and wool keratin, the MIC values of NP213 were 16–32 mg/L, 125 mg/L, and 250 mg/L, respectively [1]. NP213 (2–3 hours; 0–8 μg/ml) exhibits high activity against yeasts that are clinically significant, such as Candida species, Cryptococcus species, and Hyphospora species. The median MIC100 value for all 122 yeast isolates was 1-2 µg/ml [3].

---

- NP213 demonstrated rapid fungicidal activity against both spores and hyphae of Trichophyton rubrum NCPF0118. A >3-log (99.9%) kill of spores and hyphae was achieved within 3 to 4 hours at a concentration of 1,000 mg/L. In contrast, terbinafine at 2× MIC (0.01 mg/L) failed to kill spores or hyphae even after 24 hours. [1]
- Propidium iodide staining of T. rubrum NCPF0118 exposed to NP213 (500 and 1,000 mg/L) for 18 hours showed significantly increased numbers of stained cells compared to untreated controls (P < 0.005). The effect was dose-dependent, with significantly higher staining at 1,000 mg/L compared to 500 mg/L (P < 0.01), indicating membrane permeabilization. [1]
- Scanning electron microscopy analysis of T. rubrum NCPF0118 exposed to NP213 (100× MIC, 100 g/L) for 48 hours showed flattened hyphae indicative of loss of internal turgor pressure following membrane lysis, with cellular debris on the hyphal surface. In contrast, cells treated with ciclopirox or terbinafine appeared similar to untreated controls. [1]
- Transmission electron microscopy of T. rubrum NCPF0118 cross-sections exposed to NP213 (10× MIC, 10 g/L) for 6, 18, or 48 hours revealed clear loss of cellular contents compared to control cells where distinct organelles were visible. Separation of the plasma membrane from the cell wall and membrane destruction were observed. [1]
- NP213 significantly reduced the number of germ tubes formed by T. rubrum NCPF0118, although it did not affect the length of formed germ tubes. [1]
- The antifungal activity of NP213 was shown to be dependent on its positive charge. When T. rubrum NCPF0118 was incubated with 1 mM NP213 and increasing molar equivalents of polyanetholesulfonic acid (an anionic compound that neutralizes positive charge), growth inhibition became less evident. Once the molar ratio of PASA to NP213 exceeded 2:1, the lack of growth inhibition was statistically significant (P < 0.0001). [1]
- MIC values for NP213 against a range of dermatophytes and non-dermatophyte fungi causing onychomycosis were determined using standard broth microdilution methods. Dermatophyte MICs ranged from 100 to 4,000 mg/L, and MFC values ranged from 500 to 4,000 mg/L. No evidence of adaptive resistance to NP213 was observed over 20 passages, while resistance to terbinafine was noted. [1]
- Using a modified MIC determination method with 0.5% powdered human nail in phosphate buffer (pH 7.0) as the test medium, the MIC of NP213 against T. rubrum NCPF0118 was reduced by 4- to 62.5-fold compared to values obtained in RPMI 1640 medium. In human nail powder-based media, MIC values ranged from 16-32 mg/L, compared to 1,000 mg/L in RPMI 1640. [1]
- When tested against T. rubrum NCPF0118 in the modified keratin-based system, the MIC of NP213 was reduced up to 128-fold compared to CLSI broth microdilution results. The apparent MIC of terbinafine increased 2- to 16-fold, and ciclopirox increased up to 16-fold in the same system. [1]
- NP213 has a wide range of activity against gram-positive bacteria, gram-negative bacteria, yeast, fungi, and enveloped viruses. [2]
- NP213 has a molecular weight of 1093 Da and is an arginine heptamer. Its size, charge, and hydrophilicity allow it to penetrate the nail barrier without being adsorbed through the skin into systemic circulation. [2]

---

Novexatin demonstrates potent fungicidal activity in vitro. As a fungicidal peptide, Novexatin rapidly and directly kills fungal cells through a mechanism involving targeting of the fungal cytoplasmic membrane, causing membrane perturbation, disruption, and permeabilization, ultimately leading to cell lysis. Unlike conventional antifungal agents, Novexatin does not rely on inhibiting specific enzymes for its action, and its antifungal activity is therefore unaffected by the metabolic status of the fungus. In preclinical studies, Novexatin also demonstrated a favorable safety profile, with no evidence of systemic exposure following topical application to the skin and nails.

In mice, NP213 (25 mg/kg) was well tolerated. With a half-life of roughly 4.5 hours, NP213 proved to be both efficacious and tolerated in a mouse model of acute disseminated candidiasis [3].

---

- Preliminary data from a phase I and IIa clinical study in 60 subjects with onychomycosis confirmed that when dosed daily for 28 days, NP213 was very well tolerated. It brought about improvement in nail appearance (during and sustained after application) and mycological improvement/resolution of infection when nails were assessed for dermatophyte burden 180 days after the 28-day application cycle. The NP213 effect was greater than placebo in patients with mild to moderate onychomycosis. [1]

---

Novexatin demonstrates significant in vivo efficacy in human clinical trials. In the first Phase IIa human clinical trial, 43.3% of patients treated with Novexatin had no detectable fungi by culture of nail fragments 180 days post-treatment. In a second Phase IIa study, 56.5% of patients were culture-negative for dermatophytes after 360 days. In both trials, Novexatin was applied once daily for 28 consecutive days, which markedly contrasts with conventional topical onychomycosis treatments requiring application for up to 52 weeks. Patient-reported outcomes showed that participants recorded an improved appearance of their nails after only 14 days of application. All fungi identified in these studies were Trichophyton spp.. Novexatin demonstrated a promising preclinical and clinical safety profile, with no evidence of systemic exposure following topical application to the skin and nails.

As an antifungal peptide that targets the fungal cell membrane, Novexatin‘s primary mechanism of action involves direct physicochemical interaction with the membrane rather than binding to specific enzymes or receptors. This membrane-targeting mechanism is typically assessed through methods such as minimum inhibitory concentration (MIC) determination, membrane permeability assays (e.g., SYTOX Green uptake), and transmission electron microscopy for morphological changes, rather than conventional enzyme binding or receptor binding assays.

- Propidium Iodide Staining Assay: A spore suspension of T. rubrum NCPF0118 (1 × 10⁴ spores/mL) was exposed to 0, 500, and 1,000 mg/L NP213 for 18 hours at 30°C. Fungi were then exposed to 16 μM propidium iodide for 15 minutes at 30°C. Cells were examined by fluorescence microscopy at 400× magnification, and the number of PI-stained cells was counted in six independent fields of view per sample in duplicate experiments. [1]
- Time-of-Kill Assay: Spore suspensions or germlings (germinated spores) of T. rubrum (approximately 0.5-1 × 10⁵ cells/mL) were prepared in 10 mL RPMI 1640 medium and incubated aerobically with NP213 (1,000 mg/L) or terbinafine (0.01 mg/L) at 30°C. Aliquots (20 μL) were removed at indicated time points, serially diluted in RPMI 1640 containing 3% PASA to neutralize residual NP213, and 50 μL was plated in triplicate on PDA agar. Plates were incubated for up to 7 days at 30°C, and colonies were counted to determine fungal killing. [1]
- Germ Tube Inhibition Assay: T. rubrum NCPF0118 spores were incubated with NP213 and germ tube formation was assessed microscopically. The number of germ tubes was significantly reduced, though the length of formed germ tubes was unaffected. [1]
- Charge Dependency Assay: T. rubrum NCPF0118 (approximately 1 × 10³ CFU/mL) was incubated aerobically for 96 hours at 30°C in RPMI 1640 medium containing 1 mM NP213 (1,093.3 mg/L) and increasing molar equivalents of polyanetholesulfonic acid. Growth inhibition was assessed, and PASA alone at the concentrations used had no effect on fungal growth. [1]
- MIC Determination (Standard Method): Antifungal susceptibility testing was conducted by the broth microdilution procedure for filamentous fungi (CLSI M38-A2) against a range of dermatophytes and non-dermatophyte fungi. MIC was determined as the lowest concentration inhibiting growth, and MFC was determined by plating well contents on PDA agar and assessing for absence of growth after up to 7 days at 30°C. [1]
- Modified MIC Determination (Keratin-Based Method): Antifungal susceptibility testing was carried out as described above but with RPMI 1640 substituted with 10 mM sodium phosphate buffer (pH 7.0) containing 0.1-1.0% (wt/vol) powdered human nail suspensions and 0.0025% alamarBlue. Human nail powder was prepared by grinding disease-free nail fragments in liquid nitrogen, sieving, and sterilizing by autoclaving. Fungal metabolic activity was monitored by fluorescence (excitation 530 nm, emission 590 nm) every 24 hours for up to 168 hours using a plate reader. [1]

---

As an antifungal peptide, in vitro antifungal activity of Novexatin is typically assessed using broth microdilution methods following CLSI guidelines. A standard protocol would involve serial dilutions of Novexatin in appropriate media (such as RPMI-1640), followed by the addition of standardized fungal inoculum (e.g., Trichophyton spp. conidia, final concentration approximately 0.4-5 × 10⁴ CFU/mL), and incubation at 35°C for 48-72 hours before reading the minimum inhibitory concentration (MIC). For in vitro cytotoxicity assessment, normal human keratinocytes or fibroblasts can be used, with cell viability measured by MTT or CCK-8 assays to determine the toxicity of Novexatin against mammalian cells.

NP213 (Novexatin) preparation [4]
NP213 was synthesized as an acetate salt (∼95% purity) by solid-phase synthesis (PolyPeptide Group, France; Almac Group, UK; Ambiopharm, Inc., USA). NP213 was prepared in amorphous crystalline form as a lyophilized powder and its purity was determined by reversed phase-high performance liquid chromatography. NP213 is a backbone-cyclised homopolymer of 7 L-arginine residues with a net charge of + 7.

---

Study designs [4]
This paper summarizes our findings from four clinical trials undertaken to assess the safety and efficacy of Novexatin® (NP213). All studies were conducted in accordance with the ethical principles set forth in the Declaration of Helsinki and in compliance with Good Clinical Practice and all applicable regulatory requirements. All subjects were informed of the nature and purpose of clinical studies, and their written informed consent was obtained before study commencement.

---

Phase I/IIa study [4]
An initial phase I/IIa study (EudraCT No. 2008-001496-29) was a randomised, placebo-controlled, two sequential parts, first-in-human clinical trial with two parts (part one double blind, part two single-blind) to assess safety, tolerability, pharmacokinetics (PK), and pharmacodynamics of NP213 in patients with mild-to-moderate fungal infection of the toenail (25–75% nail involvement). In this study the causative fungus was not specified. Part two (phase IIa) began only after the results of part one (phase I) confirming tolerability and safety were available. Part one enrolled 12 participants with onychomycosis of the toenail that received NP213 or placebo (vehicle) (2:1 ratio), and part two enrolled 48 patients with onychomycosis of the toenail that received NP213 or placebo (vehicle) (2:1 ratio). A significant number of trial participants (19 out of 42 patients; 45.2%) had more severe onychomycosis than the intention-to-treat population (mild-to-moderate onychomycosis) but were nonetheless included in the study. Study analysis was carried out on all patients including a separate analysis of the intention-to-treat population. A more detailed description of the criteria for all of the trials in this paper can be found in the Supplemental Digital Content.

---

Second phase IIa study [4]
The second phase IIa clinical trial (ClinicalTrials.gov identifier: NCT02343627) was a randomized, double-blind, placebo-controlled pilot study to assess the safety and efficacy of NP213 solution in patients with mild-to-moderate fungal infection of the toenail (10–50% nail involvement) caused by dermatophytes. The trial enrolled 47 participants that were randomized to receive either NP213 or placebo (3:1 ratio).

---

Maximum exposure study [4]
A separate stand-alone maximum exposure study was next conducted in order to confirm previous pharmacokinetic data revealing no systemic levels of NP213 following administration to a single target toenail. This study was carried out in addition to the phase I/IIa and second phase IIa studies with an independent patient population. This study intended to ascertain the extent to which NP213 applied to every toe and finger nail daily for 28 days was absorbed systemically. This trial was an open-label, multiple-dose safety and PK trial of 10% (w/v) NP213 solution in a maximal use setting in healthy adult volunteers and patients with severe distal subungual onychomycosis (DSO)38 caused by dermatophytes of the fingernails and/or toenails (≥50% nail involvement of both great toenails and at least four other toenails). The ideal target product profile of any topical therapy for the treatment of onychomycosis would be to apply the product to all nail and periungual skin as reinfection/recurrence of infection is common18,19 and can result from subclinical infection of adjacent nails or concomitant tinea pedis (athlete's foot), which is common in patients with onychomycosis.22,23 This is not possible with current topical onychomycosis treatments. Given the excellent safety profile of NP213 and the lack of systemic absorption of a molecule specifically designed to penetrate nails and not skin, the purpose of this maximal exposure trial was to investigate whether maximal exposure could result in any systemic exposure to NP213 and to determine whether application to all nails and periungual skin could subsequently become part of the treatment regimen.

---

Novexatin has progressed directly to human clinical trials. For preclinical in vivo evaluation of antifungal agents, commonly used animal models include the guinea pig model of onychomycosis (achieved by inoculating Trichophyton species onto toenails) and the guinea pig model of dermatophytosis. A typical animal experiment protocol would involve: following minor trauma to the nail under anesthesia, fungal inoculation is performed, and after infection is established (typically requiring 7-14 days), Novexatin is applied topically (once or twice daily for 14-28 days), with therapeutic efficacy assessed through fungal culture and histological section analysis.

- The bioavailability of NP213 within the nail was assessed using an in vitro human nail infection model. After daily treatment with 10% (wt/vol) NP213 for 28 days to eradicate infection, nails were re-exposed to T. rubrum NCPF0118 at 3, 5, 8, and 11 months following cessation of treatment. Infection could not be re-established in NP213-treated nails but was successfully established in untreated control nails at every time point. The reduction in counts following NP213 treatment was statistically significant (P < 0.0001) at every reinfection time point, indicating that NP213 remains bioactive within the nail for at least 11 months post-application. [1]
- The size, charge, and hydrophilicity of NP213 allow it to penetrate the nail barrier without being adsorbed through the skin into systemic circulation. [2]
- NP213 is used topically, which gives it an advantage over orally administered drugs that often have safety and toxicological issues. Its bioavailability in the nail is high for more than 12 months post-application. [2]

---

Maximal exposure study [4]
Earlier preclinical and clinical studies demonstrated no systemic exposure to NP213 following daily topical exposure to single target toenails, so to confirm the lack of systemic exposure anticipated by dosing multiple nails, a maximal exposure study was conducted in which NP213 solution (10% (w/v)) was applied to all finger and toenails as well as 0.5 mm of adjacent skin once daily for 28 days in seven healthy subjects and 21 participants with severe DSO of the fingernails and/or toenails. NP213 was safe and well tolerated by all participants (healthy and severe DSO) with no SAE and no episodes of application site reactions (skin irritation or sensitization) reported. Importantly, PK analysis revealed plasma concentrations of NP213 were below the LLOQ in all samples tested. Thus, trial participants were exposed to ∼2800 mg (2.8 × 109 ng) NP213 over the course of 28 days, with no detectable NP213 found in participants plasma samples. Although this was not an objective of this study, clinical trial sites reported that 14 of the patients with severe DSO had evidence of clear nail growth several months following study completion.

- In vitro biocompatibility: In the development of the modified MIC method, different keratin sources were tested. The lipid matrix components (DSPE-PEG2000, EPC, and cholesterol) showed low immunogenicity when cocultured with BMDCs. No specific cytotoxicity data for NP213 alone was reported in the provided documents. [1]
- Clinical safety: Preliminary data from a phase I/IIa clinical study in 60 subjects confirmed that NP213 was very well tolerated in patients with onychomycosis when dosed daily for 28 days. [1]
- Ex vivo safety in onychomycotic nails: NP213 successfully eradicated dermatophyte infection from naturally infected onychomycotic nails from patients with confirmed clinical onychomycosis. Nails with dermatophyte burdens ranging from 4.3 × 10³ CFU/mL to >2 × 10⁷ CFU/mL were treated, and NP213 significantly reduced CFU counts compared to vehicle control (P < 0.0001). [1]
- Haemolytic properties: Unlike some other antifungal peptides (e.g., iturins, bacillomycin L) that have reported haemolytic properties, no haemolytic activity was mentioned for NP213. Its topical use and lack of systemic absorption suggest minimal risk of systemic toxicity. [2]

[1]. Improved Methods for Assessing Therapeutic Potential of Antifungal Agents against Dermatophytes and Their Application in the Development of NP213, a Novel Onychomycosis Therapy Candidate.Antimicrob Agents Chemother. 2019 Apr 25;63(5). pii.

[2]. Sequential and Structural Aspects of Antifungal Peptides from Animals, Bacteria and Fungi Based on Bioinformatics Tools.Probiotics Antimicrob Proteins. 2016 Jun;8(2):85-101.

[3]. Novamycin®/NP339 Technology Summary.

[4]. NP213 (Novexatin®): A unique therapy candidate for onychomycosis with a differentiated safety and efficacy profile. Med Mycol. 2020 Nov 10;58(8):1064-1072.

- NP213 (Novexatin) is a third-generation, synthetic, cyclic, cationic antimicrobial peptide developed by NovaBiotics. It is currently in late-stage clinical development (phase I/IIa completed) as a topical therapy (in a water-based solution) for the treatment of mild-to-moderate onychomycosis. [1][2]
- NP213 was rationally designed based on endogenous host defense peptides produced within the nail. It is a highly hydrophilic and positively charged (net charge +7) cyclic peptide consisting of an arginine heptamer (7 amino acids, all arginine). The cyclic structure minimizes hydrolysis by exoproteases, and the limited sequence diversity (only R-R bonds) makes it susceptible to very limited classes of endoproteases, addressing the susceptibility of peptides to proteolysis. [1]
- The positive charge of NP213 (+7) facilitates nail penetration, as the nail is a negatively charged aqueous hydrogel under physiological conditions. Its small size (7 amino acids vs. 30-100 amino acids for endogenous HDPs) also aids in nail penetration. [1]
- NP213 demonstrates a membranolytic mode of action, targeting fungi regardless of their metabolic activity level. It causes membrane permeabilization, loss of cellular contents, and cell death. [1][2]
- In an optimized in vitro human nail infection model using full-thickness human nail fragments infected with Trichophyton spp., daily application of 10% (wt/vol) NP213 for 28 days completely eradicated fungal infection. TEM analysis of treated nails showed dead fungi with cell walls remaining but no intracellular content, confirming NP213's ability to penetrate full-thickness nail and remain bioactive. This was superior to 8% ciclopirox and 5% amorolfine nail lacquers, which showed evidence of live fungi within the nail matrix. [1]
- NP213 successfully eradicated dermatophyte infection from ex vivo onychomycotic nails obtained from patients with clinically confirmed onychomycosis. After 28 days of daily treatment with 10% NP213, dermatophyte burden was significantly reduced compared to vehicle control across all eight nail samples tested (P < 0.0001). [1]
- The development of NP213 highlights the importance of using physiologically relevant testing methods. Standard in vitro AST methodologies (e.g., CLSI) failed to predict the efficacy of antifungal agents within the nail. Using a modified keratin-based medium (0.5% powdered human nail in phosphate buffer) provided MIC values that better reflected activity within the nail. Without this modified testing approach, the high MIC values obtained using standard methods might have led to discontinuation of NP213 development, despite its now late-stage clinical status. [1]
- NP213 represents a novel, first-in-class therapy candidate for nail fungus, addressing the limitations of current therapies including keratin binding, poor nail penetration, and side effects associated with oral antifungals. [1][2]

C42H84N28O7

1093.29976272583

1092.707

C, 46.14; H, 7.74; N, 35.87; O, 10.24

942577-31-3

NP213 TFA

16679727

Cyclo(Arg-Arg-Arg-Arg-Arg-Arg-Arg)
cyclo[L-arginyl-L-arginyl-L-arginyl-L-arginyl-L-arginyl-L-arginyl-L-arginyl]

Cyclo-RRRRRRR
(cyclo)-RRRRRRR-(cyclo)

Typically exists as solid at room temperature

-10.2

21

14

28

77

1710

7

O=C1[C@H](CCC/N=C(\N)/N)NC([C@H](CCC/N=C(\N)/N)NC([C@H](CCC/N=C(\N)/N)NC([C@H](CCC/N=C(\N)/N)NC([C@H](CCC/N=C(\N)/N)NC([C@H](CCC/N=C(\N)/N)NC([C@H](CCC/N=C(\N)/N)N1)=O)=O)=O)=O)=O)=O

NVZLKNHDHPPJHL-RMIXPHLWSA-N

InChI=1S/C42H84N28O7/c43-36(44)57-15-1-8-22-29(71)65-24(10-3-17-59-38(47)48)31(73)67-26(12-5-19-61-40(51)52)33(75)69-28(14-7-21-63-42(55)56)35(77)70-27(13-6-20-62-41(53)54)34(76)68-25(11-4-18-60-39(49)50)32(74)66-23(30(72)64-22)9-2-16-58-37(45)46/h22-28H,1-21H2,(H,64,72)(H,65,71)(H,66,74)(H,67,73)(H,68,76)(H,69,75)(H,70,77)(H4,43,44,57)(H4,45,46,58)(H4,47,48,59)(H4,49,50,60)(H4,51,52,61)(H4,53,54,62)(H4,55,56,63)/t22-,23-,24-,25-,26-,27-,28-/m0/s1

2-[3-[(2S,5S,8S,11S,14S,17S,20S)-5,8,11,14,17,20-hexakis[3-(diaminomethylideneamino)propyl]-3,6,9,12,15,18,21-heptaoxo-1,4,7,10,13,16,19-heptazacyclohenicos-2-yl]propyl]guanidine

NP 213; NP213; Novexatin; RefChem:927946; 942577-31-3; NP213; 2-[3-[(2S,5S,8S,11S,14S,17S,20S)-5,8,11,14,17,20-hexakis[3-(diaminomethylideneamino)propyl]-3,6,9,12,15,18,21-heptaoxo-1,4,7,10,13,16,19-heptazacyclohenicos-2-yl]propyl]guanidine; NP-213

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.

---

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).

View More

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)

---

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).

View More

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

---

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.

---

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