Preparation of Cross-Linked Sodium Alginate Microspheres with Different Metal Ions Using the Microfluidic Electrospray Technology

Hydrogen microspheres are characterized by good compatibility, design flexibility, and a high degree of customization. They have a broad application protector in biomedical fields such as therapeutic drug delivery and the construction of TC repair discoveries. In this study, we prepared hydrogen microspheres using a simple microfluidic device.

Recently we have been trying to use hydrogen microspheres loaded with some small molecules from traditional Chinese medicine to promote all the healing. In this study, we found that zinc and copper ion cross-linked sodium alginate microspheres have good anti-microbial properties, and thus copper ALGMS and zinc ALGMS are suitable for the treatment of bacterial infectious diseases. To being, mix ALG with ultrapure water in a water bath tempered at 50 degrees Celsius.

Separately prepare mass fraction with 5%solutions of calcium chloride, ferric chloride, zinc sulfate, and copper sulfate in ultrapure water. Pour each solution separately into a collecting dish. Place a glass capillary tube over a blowtorch flame to soften it and stretch it to various diameters.

With a glass cutter, cut off the fine parts and gently polish the tip port with silicon carbide sandpaper. Connect the thick end of the capillary tube to a long tubing and the other end to a dispensing needle and a five-milliliter syringe. Attach the syringe to the microfluidic syringe pump and the capillary tubing at one end to the holder.

Connect the red high-voltage end clip of the high-voltage power supply to the dispensing needle of the syringe and place the silver clip in the collection fluid. This configuration creates a simple microfluidic electrospray device. Place the collection dishes containing different metal ions directly under the capillary tube.

Turn on the microfluidic syringe pump, select fast forward to prime the tube with the ALG, and set the flow rate to two milliliters per hour. Switch on the high-voltage power switch and turn the knob to set the voltage to five to seven kilovolts. Collect ALG microspheres cross-linked with different metal ions.

After the formation of ALG hydrogel microspheres, transfer them to individual labeled 1.5 milliliters centrifuge tubes. The micrographs of ALG microspheres showed good sphericity, smooth surface, and uniform particle size distribution. Scanning electron microscopy showed that the microspheres were generally spherical, with well-defined roundness.

To begin, prepare zinc, calcium, copper, and ferric alginate hydrogel microspheres. For the antimicrobial performance test, add 10 milliliters of each of the two bacterial solutions into a 15 milliliter sterile centrifuge tube. Then incubate the cultures with 0.5 milliliters of hydrogel microspheres in a constant temperature shaker.

Dilute each bacterial solution ten to the power of five times with sterile water. Using sterile glass beads, evenly spread 200 microliters of bacterial solution on an LB solid medium. Place the plates in an incubator at 37 degrees Celsius for 12 hours to assess colony formation.

Add one milligram per milliliter of bovine serum albumin or BSA suspension to 500 microliters of each microsphere and incubate for 24 hours. Add each saturated sample to one milliliter PBS and agitate for 15 minutes at 37 degrees Celsius, with continuous shaking at 80 revolutions per minute. After aliquoting, replace the medium with a fresh one at 1, 3, 6, 12, 24, 36, and 48 hours.

Quantify the supernatant protein concentrations at specific times. Add the prepared ALG microspheres to PBS and incubate them at 37 degrees Celsius for 24 hours. Obtain whole blood from healthy mice in anticoagulation tubes containing sodium citrate.

After vortexing, centrifuge the tube for 15 minutes at 1, 500 revolutions per minute to obtain red blood cells. After flushing the cells, prepare a 10%red blood cell solution with PBS and add 20 microliters of blood cell solution to the microsphere experimental and control groups. Incubate the samples at 37 degrees Celsius for four hours, and centrifuge them at 1, 500 RPM for 15 minutes.

After keeping the supernatant aside, photograph the red blood cells in each group following hemolysis. Finally, measure the absorbance values of the supernatants in each group and compute the hemolysis rate using the formula. Different microspheres exhibited antibacterial activity against staphylococcus aureus and escherichia coli, with copper and zinc ALG microspheres showing the strongest antibacterial properties.

The drug release assay revealed that the release rate of the iron microsphere was relatively faster than that of the other three ions. Biocompatibility assessment showed that the red blood cells in the suspension remained intact upon contact with different microspheres, indicating minimal hemolysis by the microspheres.