Lacto-N-neotetraose (LNnT) is an endogenously produced metabolite. Lacto-N-neotetraose inhibits TNF-α-induced IL-8 secretion in immature epithelial cells. Lacto-N-neotetraose exhibits anti-inflammatory effect and may improve wound healing.

Physicochemical Properties

Molecular Formula	C26H45NO21
Molecular Weight	707.63
Exact Mass	707.248
CAS #	13007-32-4
Appearance	White to off-white solid powder
Density	1.72g/cm3
Boiling Point	1173.6ºC at 760mmHg
Flash Point	663.6ºC
Vapour Pressure	0mmHg at 25°C
Index of Refraction	1.668
HS Tariff Code	2934.99.9001
Storage	Powder-20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: This product requires protection from light (avoid light exposure) during transportation and storage.
Shipping Condition	Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Biological Activity

Targets	Endogenous Metabolite[1], TNFR1[2].
ln Vitro	T84 cells secrete IL-8 when exposed to 5 mg/mL of lacto-N-neotetraose for 24 hours[2]. In FHs 74 Int cells, lacto-N-neotetraose (5 mg/mL, 24 h) decreases TNF-α-induced IL-8-secretion by 38%[2]. With a Kd value of 900 ± 660 nM, lacto-N-neotetraose binds tumor necrosis factor receptor 1 (TNFR1)[2]. Lacto-N-neotetraose (5 mg/mL, 24 h) reduces inflammation caused by TNF-α in FHs 74 Int cells by shedding the TNFR1 ectodomain[2].
ln Vivo	In the lungs of pneumonia-stricken rabbits, lacto-N-neotetraose (100 μM, administered intratracheally for 24 hours) decreases the amount of Streptococcus pneumonia[3]. On the seventh day after surgery, the intradermal injection of 100/200 μg lacto-N-neotetraose enhances the rate of wound closure[4].
Animal Protocol	Animal/Disease Models: Rabbit model of pneumonia[3] Doses: 100 μM Route of Administration: Intratracheal administration for 24 h Experimental Results: diminished by ~2 logs the bacterial load in the lung at 48 h after challenge. Eliminated the extensive, edematous right middle lobe lesion evident at 48 h in control animals. Animal/Disease Models: Mice with symmetric full-thickness wounds Doses: 100, 200 μg Route of Administration: Intradermal injection, at 3, 7, 14, and 21 days post-surgery. Experimental Results: demonstrated better healing score, follicle formation, and lower epidermal thickness index (H&E staining).
References	[1]. CRISPRi-Guided Multiplexed Fine-Tuning of Metabolic Flux for Enhanced Lacto- N-neotetraose Production in Bacillus subtilis. J Agric Food Chem. 2020 Feb 26;68(8):2477-2484. [2]. The Human Milk Oligosaccharides 3-FL, Lacto-N-Neotetraose, and LDFT Attenuate Tumor Necrosis Factor-α Induced Inflammation in Fetal Intestinal Epithelial Cells In Vitro through Shedding or Interacting with Tumor Necrosis Factor Rece. [3]. Oligosaccharides interfere with the establishment and progression of experimental pneumococcal pneumonia. J Infect Dis. 1997 Sep;176(3):704-12. [4]. The in vivo effect of Lacto-N-neotetraose (LNnT) on the expression of type 2 immune response involved genes in the wound healing process. Sci Rep. 2020 Jan 22;10(1):997.

Solubility Data

Solubility (In Vitro)

H2O : 125 mg/mL (176.65 mM)

Solubility (In Vivo)

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 Formulations (e.g. IP/IV/IM/SC) 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). 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 Formulations 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). 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.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	1.4132 mL	7.0658 mL	14.1317 mL
	5 mM	0.2826 mL	1.4132 mL	2.8263 mL
	10 mM	0.1413 mL	0.7066 mL	1.4132 mL

*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.