This compound does not target specific biological receptors but functions through its chelating activity, forming stable water-soluble complexes with various metal ions. NTA acts as a tetradentate ligand, efficiently binding divalent and trivalent metal ions including Ca²⁺, Mg²⁺, Zn²⁺, Cu²⁺, and Fe³⁺ through its three carboxylate groups and one nitrogen atom . Through chelation, NTA can modulate metal ion availability in biological systems, indirectly affecting metal-dependent enzymatic reactions, cell signaling, and metabolic processes.

In cell-free systems, trisodium NTA exhibits potent metal-chelating capability, forming stable complexes with various metal ions. In bacterial and yeast mutagenicity tests, NTA itself did not induce gene reversions in five strains of Salmonella typhimurium (TA1535, TA1537, TA1538, TA98, and TA100), did not induce forward gene mutations in Schizosaccharomyces pombe P1, and did not induce mitotic gene conversions at two loci in Saccharomyces cerevisiae D4, indicating a lack of intrinsic genotoxicity . However, NTA significantly increases the solubility and mutagenic activity of certain insoluble chromium compounds such as PbCrO₄, markedly enhancing their mutagenicity in the Salmonella/microsome assay and increasing chromosome-damaging activity in CHO cell sister chromatid exchange assays . Additionally, trisodium nitrilotriacetate monohydrate inhibits gap-junctional intercellular communication between Chinese hamster lung fibroblasts, which may be relevant to its tumor-promoting mechanism .

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For life science-related study, nitriletriacetic acid trisodium salt is a biochemical reagent that can be utilized as an organic substance or biological material.

In rat models, ingestion of trisodium NTA causes dose-dependent nephrotoxicity. A 4-week rat feeding study showed that animals fed Na₃NTA·H₂O exhibited a significantly increased kidney-to-body weight ratio, and this renal toxicity could not be reproduced by equivalent sodium intake from sodium acetate, indicating that NTA's nephrotoxicity is not solely attributable to sodium load . All forms of NTA (except Zn(K/Na)NTA) increased fecal excretion of major monovalent (Na, K) and divalent (Ca, Mg) cations while reducing fecal Zn content; only Mg showed a decrease in apparent carcass retention after a 3-day balance period . In a 2-year chronic feeding study in rats, NTA caused a dose-related increase in the incidence and severity of chronic interstitial nephritis and nephrosis, but no difference in tumor types was observed across dietary groups compared to controls . In animal models, trisodium NTA is used as an inducer of bladder and kidney tumors in tumorigenesis studies .

Cell-free assays for trisodium NTA typically involve determining its chelating activity or assessing its effects in the presence of metal ions. A typical procedure involves mixing the test metal compound (e.g., PbCrO₄) with NTA at various concentrations (e.g., 10⁻⁴ M to 10⁻² M) in an appropriate buffer, incubating for a defined period, and then measuring soluble metal ion concentration using atomic absorption spectroscopy or other analytical methods to assess NTA's solubilization effect on metal compounds. In Salmonella/microsome mutagenicity assays, NTA is mixed with the test strain (e.g., TA100) and S9 metabolic activation system, and revertant colonies are counted to evaluate the genotoxicity of NTA and its metal complexes .

In a study using human EUE cell lines, the mutagenicity of trisodium NTA was evaluated using diphtheria toxin resistance selection. The protocol involved culturing EUE cells in suitable medium and treating them with various concentrations of NTA (2×10⁻² M to 2×10⁻⁸ M) for 24 hours. After the treatment period, cells were plated in selective medium containing diphtheria toxin, and surviving resistant clones were counted to assess mutation frequency. Results showed that NTA effectively induced diphtheria toxin-resistant mutants at all concentrations tested except 2×10⁻⁶ M . In CHO cell sister chromatid exchange assays, cells were co-treated with NTA and chromium compounds, and metaphase spreads were harvested for chromosome analysis to assess chromosome damage .

In vivo toxicity studies in rats typically use dietary administration of trisodium NTA. In a 2-year chronic toxicity/carcinogenicity study, NTA trisodium salt was mixed into feed at concentrations of 0.03%, 0.15%, and 0.5% and administered continuously to Sprague-Dawley rats starting from weaning. Body weight, survival, and food consumption were monitored periodically throughout the study. At 6, 12, 19, and 24 months, 5 animals per group were randomly selected for metabolic and histological studies. At study termination, necropsy was performed, major organ weights were recorded, and detailed histopathological examination of kidney and other tissues was conducted to assess the incidence and severity of tubular degeneration, chronic interstitial nephritis, and nephrosis . In rat cation balance experiments, weanling rats were fed diets containing various forms of NTA for 4 weeks, feces and urine were collected for cation analysis, and kidney-to-body weight ratios were calculated .

Absorption, Distribution and Excretion
Trisodium nitrotriacetic acid (AMT) is well absorbed in rats and dogs and excreted unchanged in the urine. AMT (8 μg/g bone) bound to calcium ions (Ca2+) and incorporated into rat bones accounts for only 0.007% of the tissue's 24-hour calcium turnover, and is therefore considered unlikely to have adverse effects on development. Rats were fed diets supplemented with 0.05%, 0.1%, 0.3%, 0.5%, 0.75%, or 2% trisodium nitrotriacetic acid for 42 and 30 days, respectively. Even when urinary NTA concentrations were equal to or higher than plasma concentrations by 200 times, NTA accumulation in bladder tissue was not greater than in the heart and liver.
In a whole-body autoradiography study, NMRI albino mice were intravenously injected with 0.93 Mg (1-(14)C-acetate) NTA trisodium salt and C57BI mice were orally administered the same dose of NTA; a large amount of radioactivity was observed in the bones and lasted for 48 hours, which was the longest time interval studied.

Toxicity Data
LC50 (Rat) > 5,000 mg/m³/4h

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Oral administration of trisodium nitrotriacetic acid (at concentrations of 0.01%, 0.10%, or 1% wt/v in drinking water) did not enhance the pathological effects of lead on the renal system in rats; however, rats given 0.01% or 0.10% trisodium nitrotriacetic acid (with or without concurrent lead administration) developed hyperglycemia.
Effects of renal tumor promoters: In male Wistar rats undergoing unilateral nephrectomy, the effects of β-cyclodextrin, DL-serine, basic lead acetate, trisodium hyponitrotriacetic acid monohydrate, potassium bromate, and diethylene glycol (as a negative control) on the early development of renal cell carcinoma were investigated after administration of N-ethyl-N-hydroxyethylnitrosamine (N-ETHEN). Male Wistar rats were fed a diet containing 1000 ppm N-ETHEN for 2 weeks. In week 3, the left kidney was removed, and the animals were then divided into 7 groups of 15 rats each. Each group received the following treatments: starting from week 3, they were fed a diet containing 1000 ppm lead acetate, 10000 ppm trisodium nitrotriacetic acid monohydrate, or 500 ppm potassium bromate for 18 weeks; subcutaneous injection of β-cyclodextrin 45 mg/100 g body weight/day for 7 consecutive days; subcutaneous injection of DL-serine 100 mg/100 g body weight every two weeks for 6 weeks; and 5% diethylene glycol was added to the drinking water as a negative control for 2 days. Five rats from each group were sacrificed at weeks 8, 12, and 20, and their kidneys were histologically examined. At week 20, the mean number of adenomatous hyperplasia (precancerous lesions) observed in the β-C group, DL-serine group, lead acetate group, trisodium nitrotriacetic acid monohydrate group, and potassium bromate group was significantly higher than that in the diethylene glycol group and the control group. Therefore, the carcinogenic effect of the chemical substance, manifested as a significant increase in adenomatous hyperplasia, could be detected in this model within a relatively short period of 20 weeks. This study investigated the dose-dependent effect of trisodium nitrotriacetic acid monohydrate as a carcinogen in a two-stage carcinogenesis process in the bladder of male Wistar rats. First, carcinogenesis was induced by adding 0.05% N-butyl-N-(4-hydroxybutyl)nitrosamine to drinking water for 4 weeks. Subsequently, 1%, 0.5%, and 0.3% trisodium nitrotriacetic acid monohydrate were added to the diet for 28 weeks. Rats were sacrificed at week 32. The results showed that with increasing concentrations of trisodium nitrotriacetic acid (TNA) monohydrate, the incidence and number of precancerous lesions (papillary or nodular hyperplasia, PN hyperplasia) gradually increased in rats treated with 0.3% to 1% TNA monohydrate. In rats treated with 1% and 0.5% TNA monohydrate in their diet, the incidence of papillomas was significantly higher than in rats treated with N-butyl-N-(4-hydroxybutyl)nitrosamine alone; in rats treated with 1% TNA monohydrate in their diet, the incidence of transitional cell carcinoma of the bladder was also significantly higher than in rats treated with N-butyl-N-(4-hydroxybutyl)nitrosamine alone (P<0.05). Administration of different doses of TNA monohydrate alone (without N-butyl-N-(4-hydroxybutyl)nitrosamine) did not induce any histological changes in the bladder (papillary or nodular hyperplasia, papillomas, or transitional cell carcinoma). These results indicate that trisodium nitrotriacetic acid monohydrate is a potent promoter of N-butyl-N-(4-hydroxybutyl)nitrosamine-induced bladder cancer in rats, and its effect is dose-dependent. /Trisodium Nitrotriacetic Acid Monohydrate/
For more complete data on interactions of trisodium nitrotriacetic acid (9 in total), please visit the HSDB record page.

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Non-human toxicity values
Mouse intraperitoneal injection LC50: 680 mg/kg body weight
Mouse intraperitoneal injection LD50: 300 mg/kg body weight
Rat intraperitoneal injection LD50: 254-444 mg/kg body weight
Rat inhalation LC50: >5 mg/L /4 hours
For more complete data on non-human toxicity values of trisodium nitrotriacetic acid (15 in total), please visit the HSDB record page.

[1]. https://pubchem.ncbi.nlm.nih.gov/compound/21152

Sodium hypotriacetate is an organosodium salt composed of sodium ions and hypotriacetate ions in a 3:1 ratio. It is carcinogenic and nephrotoxic. It contains one hypotriacetate (3-) ion. It is a derivative of acetic acid, with the chemical formula N(CH₂COOH)₃. It is a complexing agent (chelating agent) that can form stable complexes with Zn²⁺. (Excerpt from Mill's Dictionary of Chemistry, 5th edition)

C6H6NNA3O6

257.08

256.988

5064-31-3

Nitrilotriacetic acid;139-13-9;Nitrilotriacetic acid disodium salt;15467-20-6

21152

White to off-white solid powder

0.56

498.2ºC at 760mmHg

69-71 °C(lit.)

255.1ºC

0

7

3

16

171

0

C(C(=O)[O-])N(CC(=O)[O-])CC(=O)[O-].[Na+].[Na+].[Na+]

DZCAZXAJPZCSCU-UHFFFAOYSA-K

InChI=1S/C6H9NO6.3Na/c8-4(9)1-7(2-5(10)11)3-6(12)13;;;/h1-3H2,(H,8,9)(H,10,11)(H,12,13);;;/q;3*+1/p-3

trisodium;2-[bis(carboxylatomethyl)amino]acetate

TRISODIUM NITRILOTRIACETATE; 5064-31-3; Nitrilotriacetic acid trisodium salt; Trisodium NTA; Hampshire NTA;

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.)