Surfactant

The magnolol-loaded mixed micelles (MMs) and magnolol nanosuspensions (MNs) were prepared to use film hydration and antisolvent methods, respectively. The optimal MMs and MNs formulations were prepared to use magnolol, Soluplus®, and Poloxamer 188 in ratios of 1:12:5 and 2:1:1, respectively. The average particle size of MMs was 111.8 ± 14.6, and MNs was 78.53 ± 5.4 nm. The entrapment and drug loading efficiency for MMs were 89.58 ± 2.54% and 5.46 ± 0.65%, correspondingly. The drug loading efficiency of MNs was 42.50 ± 1.57%. In the in vitro release study, MMs showed a slow drug release while that of MNs was fast. The results of the Caco-2 transcellular transport study indicated that both MMs and MNs increased the permeation of magnolol. MMs and MNs markedly promoted gastrointestinal drug absorption by 2.85 and 2.27-fold, respectively, as shown in the pharmacokinetics study [1].

Traumatic Brain Injury (TBI), the main contributor to morbidity and mortality worldwide, can disrupt the cell membrane integrity of the vascular endothelial system, endangering blood–brain barrier function and threatening cellular subsistence. Protection of the vascular endothelial system might enhance clinical outcomes after TBI. Poloxamer 188 (P188) has been shown to improve neuronal function after ischemia/reperfusion (I/R) injury as well as after TBI. We aimed to establish an in vitro compression-type TBI model, comparing mild-to-moderate and severe injury, to observe the direct effects of P188 on Mouse Brain Microvascular Endothelial Cells (MBEC). Confluent MBEC were exposed to normoxic or hypoxic conditions for either 5 or 15 h (hours). 1 h compression was added, and P188 was administered during 2 h reoxygenation. A direct effect of P188 on MBEC was tested by assessing cell number/viability, cytotoxicity/membrane damage, metabolic activity, and total nitric oxide production (tNOp). While P188 enhanced cell number/viability, metabolic activity, and tNOp, an increase in cytotoxicity/membrane damage after mild-to-moderate injury was prevented. In severely injured MBEC, P188 improved metabolic activity only. P188, present during reoxygenation, influenced MBEC function directly in simulated I/R and compression-type TBI [2].

Metabolic Activity[2]
The metabolic activity was determined using the CellTiter 96 AQueous One Solution Cell Proliferation Assay. The tetrazolium compound (3- (4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt; MTS) used in this assay is bio-reduced by metabolically active cells forming a colored formazan product, soluble in culture medium. After thawing for 90 min at 20–25 °C, 20 µL of CellTiter 96 AQueous One Solution Reagent was added to each well containing 100 µL of media (achieving a concentration of approximately 16.67%). The plates were incubated in the cell culture incubator at 37 °C for 1–4 h in humified air with 5% CO2. The abs were read in the plate reader at 490 nm. Lastly, the blank abs were subtracted from the total abs.[2]

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Total NO Production[2]
The tNOp was assayed using the Cayman’s Nitrate/Nitrite Colorimetric Assay Kit. Nitric Oxide (NO), a highly reactive, short-lived free radical, is synthesized in biological systems by enzymes of the NO synthase (NOS) family, including endothelial NOS (eNOS), neuronal NOS (nNOS), and inducible NOS (iNOS). Different cell types can produce NO, including endothelial cells. The main effect of NO is paracrine activation of guanylate cyclase, which leads to an intracellular increase in cyclic guanosine monophosphate causing smooth muscle cells to relax. This assay measures the total production of NO by adding the amount of nitrite (NO2−) and the amount of nitrate (NO3−), which both represent the final products of NO. First the NO3− is exposed to NO3− reductase, converting it to NO2−. In a second step, the Griess Reagents are used to convert NO2− to an azo chromophore. Measurement of the resulting abs can determine the NO2− concentration. Reagents and samples were prepared and the assay was performed as instructed in the assay protocol. In short, 10 µL of enzyme cofactor mixture and nitrate reductase mixture were added to up to 80 µL samples (resulting concentrations: 10% enzyme cofactor mixture, 10% nitrate reductase mixture, 80% sample). After the required incubation time, 50 µL Griess Reagent R1 followed by 50 µL Griess Reagent R2 were supplemented (a concentration of 25% of each result). The abs were measured at 540 nm using a plate reader. A standard curve was prepared, following the instructions in the assay protocol, whenever the assay was performed.

Pierce Lactate Dehydrogenase (LDH) Cytotoxicity Assay Kit (Thermo Fisher Scientific; Waltham, MA, USA) was used to determine cytotoxicity/membrane damage. In healthy cells, LDH is a cytosolic enzyme. Following injury of the plasma membrane, LDH is released into the medium. We used a colorimetric method quantifying cellular cytotoxicity by measuring extracellular LDH, using a coupled enzymatic reaction. LDH catalyzes the reaction from lactate to pyruvate leading to a reduction of nicotinamide adenine dinucleotide from its oxidized (NAD+) to its reduced (NADH) form. The tetrazolium salt iodonitrotetrazolium was reduced to a red formazan product by diaphorase using NADH. The formazan formation is directly proportional to the amount of LDH release. This can be used to indicate the level of cytotoxicity/membrane disruption. The reagent was prepared and stored following the manufacturer’s instructions. Briefly, the vial containing the substrate mix (lyophilizate) was diluted with 11.4 mL ultrapure water. The assay buffer was thawed, while shielded from light. The reaction mix consists of 0.6 mL assay buffer (5%) and 11.4 mL substrate mix (95%). Firstly, 50 µL of the media of each well were transferred to a not pre-coated, clear 96-well plate. Afterward, 50 µL of the reaction mix (achieving a concentration of 50% reaction mix) was added, the plates were tapped gently and protected from light using aluminum foil. The plates were incubated at room temperature. After 30 min, 50 µL stop solution (concentration of approximately 33%) were added. Then, 10 min later, the absorbance within the media was measured at 490 nm using a plate reader (Synergy H1, BioTek Instruments Inc.). In a second step, 5 µL of lysis buffer (10X) were added to the original plate, containing the cells and 50 µL of residual media. The plates were incubated at 37 °C for 60 min and the assay was performed as described above. The absorbance (abs) was calculated: abs(media)/(abs[media] + abs[lysed cells]) [2].

Animal/Disease Models: TourniquetInduced Ischemia-Reperfusion Injury in Rats [2]
Doses: 150 mg/kg
Route of Administration: intravenous/i.v.
Experimental Results: dramatically decreased the elevated TBARS but not to control levels and SOD activity was at control levels.

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Male Sprague-Dawley rats underwent 180 minutes of tourniquet-induced ischemia. Five minutes before tourniquet release, rats received either a bolus of (1) Poloxamer-188 (P-188) (150 mg/kg; P-188 group) or (2) vehicle (Vehicle group) via a jugular catheter (n=10 per group). After 240 minutes reperfusion, both groups received a second bolus of either Poloxamer-188 (P-188) or vehicle (Vehicle) via a tail vein catheter. Sixteen hours later, rats were killed; muscle weights were determined, infarct size (2,3,5-triphenyltetrazolium chloride method), and blinded histologic analysis (hematoxylin and eosin) were performed on the gastrocnemius and tibialis anterior muscles, as well as indices of antioxidant status. [3]

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Experimental Procedures and Monitoring [3]
Rats were randomly assigned to Poloxamer-188 (P-188) (P-188 + vehicle) or Vehicle (vehicle only) groups (n = 10 per group). Rats were weighed and anesthetized using 1.5% to 2.5% isoflurane anesthesia and analgesia was administered (buprenorphine; 0.1 mg/kg intraperitoneally). Both hind limbs were shaved and animals were instrumented with a lubricated rectal temperature probe inserted 5 cm beyond the rectal sphincter. Animals were then placed supine on a warm water flow temperature-regulated bed and core temperature was maintained at 37°C ± 1°C. Blood draining of the experimental leg was performed by elevation above the level of the heart for 5 minutes before tourniquet inflation. A pneumatic tourniquet was then applied to the proximal aspect of the elevated hind limb and inflated to a pressure of 250 mm Hg. All procedures have been detailed previously.16 Tourniquets were left in place for 180 minutes. A detailed description of the experiment is shown in Figure 1, which details all procedures including periods of anesthesia, recovery, catheterizations, and administration of drug treatments. Rats were returned to their cages and allowed ad libitum access to water and food during the periods between administering anesthesia. No resuscitation fluids were administered during the experimental period.

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Administration of Poloxamer-188 (P-188) [3]
Sterile P-188 solution contained 150 mg/mL highly purified P-188, 3.08 mg/mL sodium chloride, 2.38 mg/mL sodium citrate, and 0.366 mg/mL citric acid. The placebo solution contained the same ingredients with the exception of Poloxamer-188 (P-188). Doses consisted of 1.0 mL/kg body weight of P-188 solution or vehicle. Injections of P-188 or vehicle were administered via tail vein. Two injections were administered at (1) 5 minutes before tourniquet release and (2) 240 minutes after tourniquet release. Rats were lightly anesthetized (1.5% isoflurane) before the second injection. This administration schedule was designed to maximize the bioavailability of P-188.

[1]. Enhanced Oral Bioavailability of Magnolol via Mixed Micelles and Nanosuspensions Based on Soluplus ®-Poloxamer 188. Drug Deliv. 2020 Dec;27(1):1010-1017.

[2]. Poloxamer 188 Exerts Direct Protective Effects on Mouse Brain Microvascular Endothelial Cells in an In Vitro Traumatic Brain Injury Model. Biomedicines. 2021 Aug 19;9(8):1043.

[3]. Poloxamer-188 reduces muscular edema after tourniquet-induced ischemia-reperfusion injury in rats. J Trauma. 2011 May;70(5):1192-7.

[4]. The effect of poloxamer 188 on nanoparticle morphology, size, cancer cell uptake, and cytotoxicity. Nanomedicine. 2010 Feb;6(1):170-8.

Poloxamer is an epoxide.
Poloxamer is a non-ionic triblock copolymer comprised of a hydrophobic core of polyoxypropylene flanked by two hydrophilic side chains of polyoxyethylene, that may be used as a fixative and as solubilizer, emulsifier and stabilizer in drug delivery systems.
See also: Poloxalene (annotation moved to); Poloxamer 188 (annotation moved to); Poloxamer 331 (annotation moved to) ...

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Traumatic Brain Injury (TBI), the main contributor to morbidity and mortality worldwide, can disrupt the cell membrane integrity of the vascular endothelial system, endangering blood-brain barrier function and threatening cellular subsistence. Protection of the vascular endothelial system might enhance clinical outcomes after TBI. Poloxamer 188 (P188) has been shown to improve neuronal function after ischemia/reperfusion (I/R) injury as well as after TBI. We aimed to establish an in vitro compression-type TBI model, comparing mild-to-moderate and severe injury, to observe the direct effects of P188 on Mouse Brain Microvascular Endothelial Cells (MBEC). Confluent MBEC were exposed to normoxic or hypoxic conditions for either 5 or 15 h (hours). 1 h compression was added, and P188 was administered during 2 h reoxygenation. A direct effect of P188 on MBEC was tested by assessing cell number/viability, cytotoxicity/membrane damage, metabolic activity, and total nitric oxide production (tNOp). While P188 enhanced cell number/viability, metabolic activity, and tNOp, an increase in cytotoxicity/membrane damage after mild-to-moderate injury was prevented. In severely injured MBEC, P188 improved metabolic activity only. P188, present during reoxygenation, influenced MBEC function directly in simulated I/R and compression-type TBI.[1]

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Background: Skeletal muscle injury can result in significant edema, which can in turn lead to the development of acute extremity compartment syndrome (CS). Poloxamer-188 (P-188), a multiblock copolymer surfactant, has been shown to decrease edema by sealing damaged membranes in a number of tissues after a variety of injury modalities. The objective is to determine whether the administration of P-188 significantly reduces skeletal muscle edema associated with ischemia/reperfusion injury (I-R).

Methods: Male Sprague-Dawley rats underwent 180 minutes of tourniquet-induced ischemia. Five minutes before tourniquet release, rats received either a bolus of (1) P-188 (150 mg/kg; P-188 group) or (2) vehicle (Vehicle group) via a jugular catheter (n=10 per group). After 240 minutes reperfusion, both groups received a second bolus of either P-188 (P-188) or vehicle (Vehicle) via a tail vein catheter. Sixteen hours later, rats were killed; muscle weights were determined, infarct size (2,3,5-triphenyltetrazolium chloride method), and blinded histologic analysis (hematoxylin and eosin) were performed on the gastrocnemius and tibialis anterior muscles, as well as indices of antioxidant status.

Results: P-188 resulted in significantly less edema (wet weight) and reduced an index of lipid peroxidation compared with Vehicle (p<0.05). Wet:dry weight ratios were less in the P-188 group (indicating less edema). Muscle viability as indicated by 2,3,5-triphenyltetrazolium chloride staining or routine histology did not reveal statistically significant differences between groups.

Conclusion: P-188 significantly reduced ischemia-reperfusion-related muscle edema and lipid peroxidation but did not impact muscle viability. Excess edema can lead to acute extremity CS, which is associated with significant morbidity and mortality. P-188 may provide a potential adjunctive treatment for the reduction of CS.[2]

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The aim of this work was to investigate the effect of triblock copolymer poloxamer 188 on nanoparticle morphology, size, cancer cell uptake, and cytotoxicity. Docetaxel-loaded nanoparticles were prepared by oil-in-water emulsion/solvent evaporation technique using biodegradable poly(lactic-co-glycolic acid) (PLGA) with or without addition of poloxamer 188, respectively. The resulting nanoparticles were found to be spherical with a rough and porous surface. The nanoparticles had an average size of around 200 nm with a narrow size distribution. The in vitro drug-release profile of both nanoparticle formulations showed a biphasic release pattern. An increased level of uptake of PLGA/poloxamer 188 nanoparticles in the docetaxel-resistant MCF-7 TAX30 human breast cancer cell line could be found in comparison with that of PLGA nanoparticles. In addition, the docetaxel-loaded PLGA/poloxamer 188 nanoparticles achieved a significantly higher level of cytotoxicity than that of docetaxel-loaded PLGA nanoparticles and Taxotere (P < .05). In conclusion, the results showed advantages of docetaxel-loaded PLGA nanoparticles incorporated with poloxamer 188 compared with the nanoparticles without incorporation of poloxamer 188 in terms of sustainable release and efficacy in breast cancer chemotherapy. [3]

8800 (Average)

304.115

9003-11-6

691397-13-4;126925-06-2;697765-47-2;9003-11-6;106392-12-5

24751

White to yellow solid

1.2±0.1 g/cm3

370.7±37.0 °C at 760 mmHg

60-50ºC

160.5±26.5 °C

0.0±0.8 mmHg at 25°C

1.452

0

0

2

0

7

36.7

0

CC(COCCO)OCCO

OQNWUUGFAWNUME-UHFFFAOYSA-N

InChI=1S/C7H16O4/c1-7(11-5-3-9)6-10-4-2-8/h7-9H,2-6H2,1H3

2-[2-(2-hydroxyethoxy)propoxy]ethanol

Pluronic F-68; Pluronic F-127; 2-[2-(2-hydroxyethoxy)propoxy]ethanol; 721929-01-7; Therabloat (TN); ...; CAS-9003-11-6;

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)

DMSO: ~100 mg/mL
Water: ~100 mg/mL
Ethanol: ~100 mg/mL

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.)