Stable Aqueous Suspensions of Manganese Ferrite Clusters with Tunable Nanoscale Dimension and Composition

In this video, we present the synthesis, of super paramagnetic manganese ferrite nano clusters. We report a one pot hydrothermal synthesis of manganese ferrite clusters or MFCs that offers independent control over both primary nanocrystal and cluster dimension, as well as the iron to manganese ratio. Magnetic separation allows for rapid sample purification while service functionalization using a sulfonated polymer ensures that the materials are non-aggregating, even in biologically relevant aqueous solutions.

The resulting products are well positioned for applications in biotechnology and medicine. Wash and thoroughly dry all glassware to be used in the synthesis. The amount of water in the synthesis impacts the dimensions of the MFCs.

So it’s crucial to ensure the glassware has no residual water in it. To wash the glassware, rinse with water and detergent and scrub with a FLAS brush to remove debris. Thoroughly rinse to remove all detergent and finish with the rinse of deionized water.

Rinse the polyphenol lined reactors with 37%hydrochloric acid to remove any debris from previous use. To do this, place the reactors and their caps in a large beaker and fill with hydrochloric acid until the reactors are completely submerged. Let this sit for 30 minutes before pouring out the hydrochloric acid.

Continuously rinse the beaker with the reactors with water for one to two minutes, then place the reactors in the oven to dry. Use an automatic pipette to transfer 20 milliliters of ethylene glyco into a 50 milliliter beaker with a magnetic stir bar. Weight out the required amount of iron chloride to achieve a final concentration of 1.3 millimolar and add it to the beaker.

Put the beaker on a stir plate and turn it on at 480 RPM to begin continuous stirring of the beaker. Weigh 250 milligrams of polyacrylic acid and add it to the beaker. After the addition of PAA, the solution becomes opaque and slightly lighter in color.

Weigh 1.2 grams of Urea and add it to the beaker. Using a pipette, add 0.7 millimolar manganese chloride to the beaker. Finally, using the pipette, add the required amount of ultra pure water to the beaker.

Let the solution stir for thirty minutes and notice the color change. It’ll present as a translucent dark orange color. Transfer the reaction mixture into the PPL reactor.

Note that after the solution has stirred, some solids may have accumulated on the sides of the beaker. Use a magnet to drag the stir bar around the walls of the beaker to ensure any solids that have accumulated on the sides are dispersed into the reaction solution. Once the solution is mixed and ready, transfer it into the 50 milliliter PPL lined reactor.

Use a clamp and lever to seal the reactor in the stainless steel autoclave as tightly as possible. Clamp the reactor vessel to a stable surface and using a rod, insert it into the cap as a lever, push the reactor to seal. Note that the sealed reactor should not be able to be opened by hand.

This is crucial as the high pressure environment of the oven requires a tight seal on the reactor. Place a reactor into an oven for 20 hours at 215 degrees Celsius. After the hydrothermal reaction is done, remove the reactor from the oven and allow it to cool down to room temperature.

The pressure of the oven will enable the reactor to be opened by hand. Note that at this point, the reactor will contain the MFC product dispersed in ethylene glycol with other impurities, such as unreacted polymer. And will be opaque black solution.

The product isolated in the following steps. Place 200 milligrams of steel wool into a glass vial. Fill the glass vial halfway with the reaction mixture from the reactor.

Fill the rest of the vial with acetone and shake well. Note that the steel wool increases the magnetic field strength in the vial and will help magnetic separation of the nano clusters from solution. Place the vial on a magnet for magnetic collection to occur.

The result will be a translucent solution with precipitate at the bottom. Pour off the supernatant solution while the MFCs are magnetically trapped by the steel wool by holding the magnet to the bottom of the vial while pouring. Ethylene glycol will mostly be removed in this step.

Start washing with the low ratio of acetone to water and increase the ratio in subsequent washes until pure. Do this three to four times. Remove the vial from the magnet and fill it with water.

Shake well to dissolve the MFCs. Now the product will be fully dispersed in water. Repeat the previous two steps several times until the aqueous solution of the MFCs produces no bubbles when shaken.

The result will be a dark opaque ferrofluid that will respond strongly to magnets. In order to keep our clusters stable, we modify them with a co-polymer, PAA-co-AMPS-co-PEG, which provides both steric and electrostatic repulsion. The sulfonate group of the AMPS units will provide charge stabilization while the PEG unit will stericly hinder the inter cluster aggregation.

Overall, the modified clusters will remain stable even in various types of harsh conditions. Combine 10 milliliters of purified nanoparticles in a 20 milliliter vial, with 10 milliliters of saturated Nitra dopamine solution. Wait for five minutes.

Wash the Nitra dopamine coated MFCs using magnetic separation. Pour out the pale yellow supernatant. Add water and shake vigorously.

Then pour out water using the magnet to retain the product. Repeat this washing several times, leaving the dark brown collection in the vial. Mix one milliliter of EDC solution, one milliliter of MES buffer, and three milliliters of polymer solution.

Lightly stir by swirling the mixture and let it sit for approximately five minutes. It should be a clear and colorless solution when fully combined. Add this mixture to the MFC collection and place the vial in an ice bath.

Lower the probe sonicator into the solution and then turn it on. After a five minute sonication treatment, add roughly five milliliters of ultra pure water to the vial while the sonicator is still running. Continue monitoring the vessel to ensure that no product spills.

Maintain the ice in the ice water mixture, as some of the initial ice will melt due to the intensity and heat of the sonication. Allow the mixture to sonicate for an additional 25 minutes for a total of 30 minutes. Place the vial on top of a magnet to separate the MFCs and pour out the supernatant solution.

Wash the modified MFCs with deionized water several times. Fill the vial with the MFCs with ultra pure water. Pipette this fluid into a vacuum filtration system with a 0.1 micron polyether sulfate membrane filter to remove any irreversibly aggregated MFCs.

Make sure to flush the walls of the funnel to minimize any loss of product. Vacuum filter the solution. Repeat this process two to three times.

The result will be a purified aqueous solution of mono disperse MFCs. MFCs isolated when the magnetic separation method have higher mono dispersity than those separated with ultracentrifugation, as shown here. Here, we see TEM images of purified nano clusters, in order of increasing average cluster diameter.

Denoted as DC.The amount of water added in the initial reaction mixture determines the diameter of the nano clusters. Adding more water in the reaction results in nano clusters with smaller diameters, while less water increases their diameters. In this way, the experimenter has control over the size of the nano cluster product.

Here we see TEM images of nano clusters in order of increasing manganese to iron molar ratio. The ratio of manganese to iron precursors in the initial reaction mixture determines the molar ratio of the metals in the cluster product. Increasing the manganese to iron ratio in the synthesis will increase this ratio in the clusters, and vice versa.

In contrast the following TEM images depict samples with irregular morphologies. As shown in the left image, the out of shape spilled looking cluster was produced with the exclusion of any additional water. This hinders the dynamic assembly of the primary nano crystals that have yet to form clusters.

The sample in the image on the right had insufficient reaction time, which was not enough for primary nano crystal growth and cluster ripening. These poor results demonstrate that an appropriate amount of the reactant as well as reaction time is necessary to achieve consistently successful results. Here, we place a sample of the original PAA coded clusters in PBS buffer on the left.

On the right, we do the same with an equivalent amount of the modified PAA co-AMPS-co-PEG coded clusters. Notice the fast aggregation of the PAA coded clusters, while the modified clusters remain stable for a long time. This suggests the improved colloidal stability as a result of the copolymer coding.

In conclusion, our synthesis allows for the quick and efficient production of manganese ferrite clusters. The synthesis creates independently tunable cluster dimension and composition by simply controlling the addition of water and manganese to iron precursor ratio. We can easily modify this method to achieve different but predictable magnetic nanomaterials.

Further, the magnetic separation and service functionalization techniques achieve high mono dispersity and strong stability in biological media, respectively. Our method allows for greater accessibility in the clusters production and widespread application across a variety of fields.