Absorption, Distribution and Excretion
Since L-canavalina has shown potential as an antitumor drug, its toxicity was investigated. This natural product showed only mild toxicity in Sprague-Dawley rats after a single subcutaneous injection: the LD50 for adult rats was 5.9 ± 1.8 g/kg, and the LD50 for 10-day-old rats was 5.0 ± 1.0 g/kg. Following a single administration of 2.0 g/kg, the systemic clearance of canavalina in adult rats was 0.114 L/hr, the steady-state volume of distribution was 0.154 L, and the half-life was 1.56 h. After intravenous injection, 48% of the dose was excreted unchanged in the urine; after subcutaneous injection, 16% of the dose was excreted in the urine. The bioavailability of the 2.0 g/kg subcutaneous dose was 72%. A single oral dose of canavalina showed lower toxicity in adult rats than subcutaneous injection. The bioavailability of the 2.0 g/kg oral dose was 43%, with only 1% of cannavarine excreted in the urine. 24 hours after oral administration, 21% of cannavarine remained in the gastrointestinal tract. At 4 or 24 hours post-administration, less than 1% of the L-[guanidino-(14)C]cannavarine at the 2.0 g/kg dose was incorporated into the proteins of both adult and neonatal rats. Repeated subcutaneous injections of cannavarine led to more severe toxicity. Rats receiving daily subcutaneous injections of cannavarine for 7 consecutive days experienced weight loss and hair loss. Food intake in adult rats receiving this regimen decreased by 80%, but returned to normal after discontinuation of cannavarine injections. Histological studies of adult rats treated with 3.0 g/kg cannavarine daily for 6 consecutive days revealed atrophy and fibrosis of pancreatic acinar cells. Following a single subcutaneous injection of 2.0 g/kg cannavarin, serum amylase and lipase levels increased; after three consecutive injections, both serum enzymes decreased. Simultaneously, serum glucose and urea nitrogen levels increased, while cholesterol levels decreased. The most significant change was a marked decrease in serum aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase activities.

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Metabolism/Metabolites
L-Cannavarin (CAV) is an arginine (ARG) analogue isolated from canavalia ensiformis. CAV can be incorporated into the cellular proteins of MIA PaCa-2 human pancreatic cancer cells, and abnormal cannavarin proteins are not preferentially degraded. The hydrolysis of CAV to canarolin (CAN) and urea is mediated by arginase. CAN is a potent metabolite that inactivates vitamin B6-containing enzymes and may inhibit cell growth. To determine the presence of arginase in MIA PaCa-2 cells and its specificity for ARG and CAV, we employed a radiometric assay that quantifies arginase-mediated hydrolysis of (14)C from L-[guanidino-(14)C]ARG or L-[guanidino-(14)C]CAV. The amount of (14)CO2 released when cells were exposed to [(14)C]CAV or [(14)C]ARG was negligible. The amount of arginase secreted by pancreatic cancer cells was also extremely low. We compared the cytotoxic effects of CAN and CAV on cells treated with different concentrations of ARG. These studies indicate that the cytotoxic effect of CAV on MIA PaCa-2 cells is not due to its conversion to the active metabolite CAN. Reduced hydrolysis of CAV by arginase leads to the persistence of CAV and increases its incorporation into these cellular proteins. The low arginase content in the pancreas makes CAV an ideal candidate for further research in pancreatic cancer. L-carnavanine and its arginase-catalyzed metabolite L-carnaline are two novel anticancer drugs under development. Since immunotoxicity evaluation is crucial during drug development, we evaluated the antiproliferative effects of L-carnavanine and L-carnaline in vitro. Results showed that both L-carnavanine and L-carnaline are cytotoxic to cultured peripheral blood mononuclear cells (PBMCs). Furthermore, PBMCs were simultaneously exposed to L-carnavanine or L-carnaline, along with a series of compounds that may act as metabolic inhibitors of L-carnavanine and L-carnaline (L-arginine, L-ornithine, D-arginine, L-lysine, L-homarginine, putrescine, L-ω-nitroarginine methyl ester, and L-citrulline). The ability of these compounds to overcome the cytotoxic effects of L-carnavanine or L-carnaline was assessed to gain a deeper understanding of the potential biochemical mechanisms underlying the toxicity of these two novel anticancer drugs. The results indicate that the toxicity mechanism of L-carnavanine is mediated through the L-arginine utilization pathway, while its metabolite, L-carnaline, exerts toxicity on human peripheral blood mononuclear cells (PBMCs) by interfering with polyamine biosynthesis. Elucidating the biochemical mechanisms underlying the effects of L-carnavanine and L-carnaline on lymphocyte proliferation will help maximize the efficacy of these novel anticancer drugs and minimize their toxicity. This study investigated the metabolism of L-carnavanine, a non-protein amino acid with significant antitumor activity. 2.0 g/kg of L-carnavanine was mixed with 5 μCi of L-[guanidino-(14)C]carnavanine (58 μCi/μmol) and administered to approximately 200 g female Sprague-Dawley rats via intravenous, subcutaneous, or oral routes. After 24 hours, the recovery rates of 14C in urine were 83%, 68%, and 61% of the administered dose, respectively. Another 5-8% of 14C is excreted as 14CO2. The gastrointestinal tract contains 21% of the orally ingested 14C. Regardless of the route of administration, the 14C content in serum, feces, tissues, and newly synthesized proteins is only a few percent of the initial dose. Analysis of urinary metabolites containing 14C revealed that after intravenous injection, 14C urea accounted for 88% of the urine; after subcutaneous injection, 75%; and after oral administration, 50%. In all routes of administration, 14C guanidine accounted for 5% of the radioactivity in urine, and 14C guanidinoacetic acid accounted for 2%. Serum and urinary amino acid analysis showed a significant increase in ornithine levels. The levels of basic amino acids such as histidine, lysine, and arginine were also high in urine. Plasma ammonia levels were measured after oral administration of 1.0, 2.0, and 4.0 g/kg cannavarin, respectively. A rapid but transient increase in plasma ammonia levels was observed only in the 4.0 g/kg dose group. This suggests that at the drug concentration used in this study, elevated plasma ammonia levels are unlikely to be the cause of cannavarin toxicity. Previously, cannavarin (2-amino-4-guanidinoxybutyrate) was observed to react with amino acid oxidases to form hydroxyguanidine. This study indicates that hydroxyguanidine is generated by a non-enzymatic β,γ-elimination reaction following enzymatic oxidation of the α-carbon, and the removal of the β-hydrogen is generally catalyzed by a base. This elimination reaction requires the presence of an anionic stabilizing group at the α-position—a protonated imino group (imine ion group) or a carbonyl group. The imine ion group is more activating than the carbonyl group. Protonation of the guanidinoxy group further promotes the elimination reaction. Another product generated in the elimination reaction was identified as vinyl glyoxylate (2-oxo-3-butenoate), a highly electrophilic substance. The product of subsequent oxidation and hydrolysis was identified as α-keto-γ-guanidinoxybutyrate (ketocannavarin). The ratio of hydroxyguanidine to ketocanavalina depends on the concentration and alkalinity of the basic catalyst and the pH value. In the presence of aminourea, the elimination reaction is inhibited because the imino group in the aminourea derivative of ketocanavalina is not sufficiently protonated. Incubation of canavalina with 5'-deoxypyridoxal also produces hydroxyguanidine. Since the elimination reaction occurs under mild conditions, in vivo, the elimination reaction can occur when the α-C of L-canavalina (taken or endogenously generated) or other amino acids with a well-defined leaving group at the γ-position (e.g., S-adenosylmethionine, methionine sulfoxide imine, homocysteine, or a cysteine-homocysteine mixed disulfide) is oxidized by an L-amino acid. This process may be catalyzed by L-amino acid oxidases, transaminases, or dehydrogenases. Therefore, ethylene glyoxylic acid is likely a normal metabolite in mammals, and elevated concentrations may lead to in vivo toxicity of canavalina and some of the aforementioned amino acids.

Toxicity Summary
Identification and Uses: L-Carnavanine is a solid. It is a potentially toxic L-arginine antimetabolite found in many legumes. It exhibits significant antitumor activity against various animal tumor-bearing and cancer cell lines. L-Carnavanine has been used as an experimental drug. Human Exposure and Toxicity: L-Carnavanine is a naturally occurring L-amino acid that, due to its structural similarity to L-arginine, interferes with enzymes that utilize L-arginine. In macrophages and polymorphonuclear leukocytes expressing inducible nitric oxide synthase (iNOS), L-Carnavanine inhibits the synthesis of L-arginine-derived NO. The effect of L-Carnavanine on human platelets is concentration-dependent: at low concentrations, it exerts an antiplatelet aggregation effect through NOS-independent inhibition; at high concentrations, it inhibits NO synthesis and ceases to exert its antiplatelet aggregation effect. L-Carnavanine is cytotoxic to cultured human peripheral blood mononuclear cells (PBMCs). These results indicate that the toxicity mechanism of L-canavalina is mediated through the L-arginine utilization pathway, and that its metabolite, L-canavalin, exerts toxicity on human PBMCs by interfering with polyamine biosynthesis. Animal experiments: Following a single subcutaneous injection, L-canavalina exhibits low toxicity in rats: the LD50 for adult rats is 5.9 ± 1.8 g/kg, and the LD50 for 10-day-old rats is 5.0 ± 1.0 g/kg. Repeated subcutaneous injections of canavalina lead to more severe toxicity. Rats receiving daily subcutaneous injections of canavalina for 7 consecutive days experienced weight loss and hair loss. Adult rats receiving this dosing regimen showed an 80% reduction in food intake, which returned to normal after discontinuation of canavalina injections. Histological studies of adult rats treated with 3.0 g/kg canavalina daily for 6 consecutive days revealed pancreatic acinar cell atrophy and fibrosis. Following a single subcutaneous injection of 2.0 g/kg cannavarin, serum amylase and lipase levels increased; after three consecutive days of injections, both enzymes decreased. Simultaneously, serum glucose and urea nitrogen levels increased, while cholesterol levels decreased. The most significant changes were a marked decrease in serum aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase activities. From day 84 to day 477 postnatally, 18 female mice were fed a diet containing 1.56% cannavarin sulfate (1% base), while another 18 mice were fed a control diet. From day 84 to day 164 postnatally, each mouse was fed 4 g/day of diet, followed by 5 g/day. Of the 10 mice fed cannavarin, only 6 (compared to 5 in the control group) successfully gave birth before day 17 of gestation. Counts of the corpus luteum, embryo, and resorption sites suggested that these significant effects on pregnancy were likely due to implantation failure. The survival rate of control mice was only 50%, while the survival rate of mice fed with cannavanin was as high as 89%. These results suggest that cannavanin may prolong the lifespan of mice but interfere with their reproduction. To elucidate the cytotoxic mechanism of each compound, we investigated the mutagenic activity of l-cannavanin and its metabolite l-canarolin against Salmonella typhimurium TA100 and Bacillus subtilis h17 rec+ & M45 rec-. Neither compound nor its metabolite extracted from rat liver homogenate induced DNA base pair substitutions or frameshift mutations. Clearly, these compounds do not act directly on DNA, but affect DNA metabolism through other mechanisms, such as the formation of l-cannavanin protein. Cannavanin significantly inhibited the growth of colon cancer in rats.

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Interaction
The growth and development of final-instar larvae of the tobacco hawk moth (Manduca sexta, Sphingidae) fed a diet containing 2.5 mmol l-cannavanin were significantly disrupted. When these organisms were fed a diet containing cannavanin and supplemented with L-arginine, L-citrulline, L-ornithine, or L-2,4-diaminobutyric acid in a molar ratio of 1:10, the cannavanin-mediated larval growth inhibition was significantly enhanced; larval hemolymph volume increased (edema), and mass mortality occurred due to incomplete larval-pupa molting. This study also investigated the regulatory effects of the non-selective endothelin receptor antagonist bosentan and the relatively selective inducible nitric oxide synthase inhibitors aminoguanidine and L-cannavanin on reduced mesenteric blood flow and liver and spleen damage caused by endotoxemia. Swiss albino mice (20-40 g) were intraperitoneally injected with bosentan (3, 10, or 30 mg/kg), aminoguanidine (15 mg/kg), or L-cannavanin (20 or 100 mg/kg) 10 minutes before receiving saline or E. coli endotoxin (10 mg/kg). Four hours later, mice were anesthetized, and mesenteric blood flow, spleen and liver weight/body weight ratios were measured, followed by histopathological examination. Endotoxin reduced mesenteric blood flow (mL/min): 3.0 ± 0.2 in the saline group; and increased liver weight (47.5 ± 2.0 g/kg body weight; 60.8 ± 1.9 g/kg body weight; n=10, p<0.05) and spleen weight (3.9 ± 0.5 g/kg body weight; 8.6 ± 0.9 g/kg body weight; n=10, p<0.01) in the endotoxin group, causing significant histopathological damage to both organs. Bosentan was ineffective at a dose of 3 mg/kg, but completely eliminated all the harmful effects of endotoxin at doses of 10 mg/kg and 30 mg/kg. Aminoguanidine blocked most of the effects of endotoxin, except for its effect on the spleen. In contrast, L-carnavanine only blocked endotoxin-induced liver weight gain, but it itself increased spleen weight and failed to block any other effects of endotoxin. Therefore, it can be inferred that the beneficial effects of aminoguanidine are not primarily produced through a selective inducible nitric oxide synthase inhibition mechanism, as L-carnavanine is not entirely effective. The beneficial effects of using bosentan to inhibit endothelin in the treatment of endotoxemia could be further used to understand and treat sepsis-related syndromes. This study investigated the effects of L-carnavanine (a nitric oxide synthase inhibitor) on endotoxin-induced shock in pentobarbital-anesthetized rats. Endotoxin infusion (2.5 mg kg⁻¹ hr⁻¹, over 6 hours) resulted in progressive and significant hypotension and hypoglycemia. Electron microscopy revealed significant changes in the kidneys, including severe endothelial cell destruction and platelet aggregation in the vessels. Significant aggregation of polymorphonuclear leukocytes in small vessels with endothelial cell destruction was observed in the lungs. Treatment with L-carnavanin (administered as a bolus injection of 10 mg/kg per hour, starting 70 minutes after endotoxin or saline infusion) significantly reduced endotoxin-induced hypotension without affecting hypoglycemia. This treatment significantly reduced endotoxin-induced renal and pulmonary lesions observed under electron microscopy. Although L-carnavanin, like L-NAME, inhibited the activity of inducible nitric oxide synthase in the cerebellum and spleen in vitro, unlike L-NAME, it did not alter arterial blood pressure or carotid blood flow in control rats. The data suggest that L-carnavanin is at least a selective inhibitor of inducible nitric oxide synthase in vivo, suggesting that inhibition of this enzyme may be beneficial for endotoxin-induced shock. We analyzed the effects of L-carnavanin and cadmium on the composition of ribonucleoproteins in HeLa S3 cells. Both chemicals induced similar changes in the structure of different RNPs and in RNA and protein synthesis. Pulse-tracking autoradiography experiments showed that both cannavarin and cadmium preferentially inhibited nucleolar RNA synthesis and slowed the transport or processing of nucleolar and extranuclear RNA. The nucleolus became rounded and dense. Perichondrial granules and fibers accumulated, interchromatin fibers decreased, and some intranuclear structures appeared, which appeared to be involved in the morphogenesis of the perichromatin granules accumulated during processing. The appearance of 29-35 nm granule clusters may be related to assembly defects of the components of perichromatin granules. The effects of different transcription inhibitors on the accumulation of perichromatin granules suggest that these granules represent a specific subset of hnRNP.
Based on several physiological properties of L-cannavarin, we validated the prediction that this arginine analog could enhance the cytotoxic effects of gamma rays on mammalian cells. We used the human colon tumor cell line HT-29 to conduct time-dose studies in logarithmic growth phase culture to determine the conditions that could maximize the incorporation of L-cannavarin into cellular proteins while maintaining most cells viable for subsequent gamma-ray viability measurements. At an L-carnavanine to arginine ratio of 2.5, this analogue exerted cytotoxic effects on cells for at least 6 days following a single cell division. During the first 12 hours of treatment in the logarithmic growth phase, even at an L-carnavanine to arginine ratio of 20, cell death due to clonogenic ability was minimal (less than 20%). However, 24-hour exposure led to an exponential decrease in cell viability with increasing L-carnavanine concentration. Based on these findings, we investigated the effects of L-carnavanine treatment and gamma-ray injury on cell viability, examining different conditions and durations. The results showed that 48 hours of treatment with this analogue at an L-carnavanine to arginine ratio of 10 maximally enhanced X-ray-induced cytotoxicity (assessed by loss of clonogenic ability). A significant increase in radiosensitivity was observed when L-carnavanine was administered before or after cell irradiation. This enhancement was observed at all radiation doses in both regimens. Combined with our previous findings on the antitumor activity of L-canavalina in L1210 mouse leukemia, these results suggest that this amino acid analog has potential applications in cancer treatment.

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Non-human toxicity values
LD50 rat subcutaneous injection (10 days old) 5.0 ± 1.0 g/kg
LD50 rat subcutaneous injection (adult) 5.9 ± 1.8 g/kg

L-Carnavanine is a non-protein L-α-amino acid formed by replacing the oxygen atom of L-homoserine with a guanidinium group (carbamoimide group). Although its structure is related to L-arginine, it is not protein-derived. It is a plant-derived insecticide and plant metabolite. Its function is related to L-homoserine. It is the conjugate base of L-carnavanine (1+). It is the zwitterion tautomer of L-carnavanine. L-Carnavanine has been reported to be found in Poissonia orbicularis, Sesbania herbacea, and several other organisms with relevant data. Therapeutic Uses /EXPL THER/ L-Carnavanine is a selectively inducible nitric oxide synthase (NOS) inhibitor with therapeutic effects on circulatory failure in rats with endotoxin shock. To investigate the direct relationship between these beneficial effects and the inhibition of NO production by L-carnavanin in rats under endotoxin shock, we used electron spin resonance (ESR) to detect changes in venous nitrosylhemoglobin (NO-hemoglobin) levels. Anesthetized rats were intravenously injected with lipopolysaccharide (10 mg/kg). One hour after lipopolysaccharide injection, the rats were divided into four groups: the lipopolysaccharide group received 0.3 mL of normal saline intravenously every hour; the L-carnavanin 10 and L-carnavanin 20 groups received 10 mg/kg or 20 mg/kg of L-carnavanin intravenously every hour, respectively; the L-NAME group received 15 mg/kg of NG-nitro-L-arginine methyl ester (L-NAME) intravenously first, followed by 10 mg/kg intravenously every hour. The sham-operated group received normal saline instead of lipopolysaccharide, and the L-carnavanin group received 20 mg/kg of L-carnavanin intravenously every hour one hour after normal saline injection. Five hours after injection of lipopolysaccharide (LPS) or saline, the pressor response to norepinephrine (1 μg/kg, intravenously) was assessed. In the LPS group, LPS caused a progressive decrease in mean arterial pressure and impaired the pressor response to norepinephrine. Administration of L-canavalina or L-NAME alleviated endotoxin-induced hypotension and vascular hyporesponsiveness to norepinephrine. In the L-canavalina group, L-canavalina did not alter mean arterial pressure or the pressor response to norepinephrine. L-canavalina or L-NAME significantly inhibited endotoxin-induced elevation of venous NO-hemoglobin levels. These data suggest that the beneficial hemodynamic effects of L-canavalina are associated with enhanced NO production by inducible nitric oxide synthase (iNOS) in a rat model of endotoxin shock. L-canavalina is a potential treatment for endotoxin shock. Cardiovascular failure in sepsis may be due to increased nitric oxide biosynthesis caused by diffuse expression of inducible nitric oxide synthase (iNOS). Nitric oxide synthase inhibitors may have therapeutic value in this context, but their use has been reported to produce harmful side effects, possibly related to the blockade of constitutive nitric oxide synthase (iNOS). Therefore, the use of selective inhibitors of inducible nitric oxide synthase (iNOS) may be more appropriate. This study aimed to evaluate the role of L-carnavanine (a potential selective inhibitor of inducible nitric oxide synthase (iNOS)) in an animal model of septic shock. Anesthetized rats were challenged by intravenous injection of 10 mg/kg lipopolysaccharide (LPS). One hour later, mice were randomly assigned to receive either L-carnavanine (20 mg/hr/kg, n=15), nitro-L-arginine methyl ester (5 mg/hr/kg, n=13), or 0.9% sodium chloride solution (2 mL/hr/kg, n=21) via intravenous infusion over 5 hours. Lipopolysaccharide (LPS) induces a progressive decrease in blood pressure and cardiac index, accompanied by significant lactic acidosis and a significant increase in plasma nitrate levels. L-carnavanine significantly alleviated all these changes and improved the tolerance of endotoxemia animals to acute hypovolemic episodes. Furthermore, L-carnavanine significantly improved the survival rate of mice challenged with a lethal dose of LPS. In contrast to L-carnavanine, nitro-L-arginine methyl ester, while raising blood pressure, caused a severe decrease in cardiac index and significantly exacerbated lactic acidosis. This drug did not improve the survival rate of endotoxemia mice. In subsequent experiments, we found that the pressor effect of L-carnavanine in late endotoxemia (4 hours) could be reversed by L-arginine, confirming its association with nitric oxide synthase inhibition. Conversely, in early endotoxemia (first hour) or in the absence of LPS exposure, L-carnavanine had no effect on blood pressure, indicating that the drug does not have constitutive nitric oxide synthase inhibitory activity. In summary, L-carnavanine has beneficial hemodynamic and metabolic effects on endotoxin shock in rodents and can improve their survival rate. The effect of L-carnavanine is related to the selective inhibition of inducible nitric oxide synthase (NOS), which is distinctly different from the adverse effects produced by the non-selective NOS inhibitor nitro-L-arginine methyl ester under similar conditions.
/EXPL THER/ Injection of lipopolysaccharide (LPS) into anesthetized rats resulted in a decrease in mean arterial pressure, an increase in heart rate, and death within 4–6 hours. Intravenous infusion of nitro-L-arginine methyl ester (50 mg/kg, a constitutive and inducible NOS inhibitor) 60 minutes after LPS injection caused an immediate increase in blood pressure, followed by a sharp decrease, ultimately leading to death. Conversely, intravenous infusion of L-carnavanine (100 mg/kg, an in vitro selectively inducible NOS inhibitor) at 60 and 180 minutes after LPS stimulation has been reported to increase mean arterial pressure and reverse LPS-induced hypotension. However, in animals where L-carnavanine administration 60 minutes after LPS stimulation prevented hypotension, intravenous infusion of NG-nitro-L-arginine methyl ester 180 minutes after stimulation resulted in an immediate increase in mean arterial pressure, followed by a rapid decline in blood pressure and heart rate, ultimately leading to sudden death. In contrast, re-administration of L-carnavanine 180 minutes after stimulation maintained blood pressure throughout the experiment. These results indicate that inhibition of both constitutive and inducible nitric oxide synthase during endotoxemia leads to death. However, the use of selective inhibitors of inducible nitric oxide synthase restored mean arterial pressure to baseline levels and provides a treatment option for hypotension induced by shock.
/EXPL THER/ There is an urgent need for drugs with novel mechanisms of action to provide new approaches to the treatment of pancreatic cancer. Due to its structural similarity to L-arginine, L-carnavanine (a β-oxa analog of L-arginine) is a substrate for arginyl-tRNA synthetase and is incorporated into nascent proteins, replacing L-arginine. Although L-arginine and L-carnavanine are structurally similar, the oxyguanidinium group of L-carnavanine is much less basic than that of L-arginine. Therefore, L-carnavanine protein lacks the ability to form key ion interactions, leading to alterations in protein structure and function, ultimately resulting in cell death. Because L-carnavanine can be selectively isolated by the pancreas, it may be particularly suitable for adjuvant therapy in pancreatic cancer. This novel cytotoxic mechanism forms the basis of the anticancer activity of L-carnavanine; therefore, arginyl-tRNA synthetase may represent a new target for the development of such therapeutics. For more complete data on the therapeutic uses of (L)-carnavanine (a total of 8 types), please visit the HSDB record page.

C5H12N4O3

176.17

176.091

543-38-4

L-Canavanine sulfate;2219-31-0

439202

Crystals from absolute alcohol

1.61g/cm3

431.2ºC at 760mmHg

184ºC

214.6ºC

1.602

0.094

4

5

5

12

178

1

C(CONC(=N)N)[C@@H](C(=O)O)N

FSBIGDSBMBYOPN-VKHMYHEASA-N

InChI=1S/C5H12N4O3/c6-3(4(10)11)1-2-12-9-5(7)8/h3H,1-2,6H2,(H,10,11)(H4,7,8,9)/t3-/m0/s1

(2S)-2-amino-4-(diaminomethylideneamino)oxybutanoic acid

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.)