PLGA (poly (lactic-co-glycolic acid)) gel used as injectable long-acting local anesthesia medicine as well as application and preparation method of PLGA gel

Abstract

The invention belongs to the technical field of medicine preparation, and relates to PLGA gel used as an injectable long-acting local anesthesia medicine as well as application and a preparation method of the PLGA gel. The PLGA gel comprises the PLGA gel which is loaded with superparamagnetic Fe3O4 particles and a small molecule local anesthetic, namely, levobupivacaine (Levobupi). The PLGA hydrogel containing Fe3O4 and levobupivacaine can be used as an injectable long-acting local anesthesia medicine, and the hydrogel can be changed from liquid to solid after being injected into a living body, so that the release efficiency of the medicine is slowed down, and the local action time of the hydrogel is prolonged. In addition, the hydrogel can undergo a phase change from a solid state to a liquid state under magnetocaloric therapy (MHT), increase pore diameter, promote release of levobupivacaine, and provide enhanced pain relief in vivo. Compared with the prior art, the invention has the beneficial effects of simple synthesis and easy industrialization. The drug release rate is adjusted through the phase change of the hydrogel, super-long-acting analgesia is realized, and the analgesia time is gt; 20 days. The gel can be loaded with various local anesthesia drugs such as L-bupivacaine, and pain management can be carried out on various pain scenes clinically.

Description

Technical Field

[0001] The invention belongs to the technical field of drug preparation, and relates to a PLGA gel used as an injectable long-acting local anesthetic drug and an application and a preparation method thereof. Background Art

[0002] Neuropathic pain is a severe and persistent pain condition that usually occurs after a nerve is damaged. Unlike acute pain, which is protective, neuropathic pain persists even after the original injury has healed. This pain is extremely challenging to treat, affecting 7-10% of the world's population and causing significant health and economic burdens.

[0003] Local anesthetics, opioids, and cyclooxygenase inhibitors are commonly used for pain relief, and opioids remain the main treatment for postoperative pain. However, opioids are associated with a variety of adverse reactions, including tolerance, addiction, nausea, abuse, itching, and risk of overdose. Although local anesthetics are effective in managing postoperative pain, they have a short duration of action and cannot completely replace opioids. Moreover, their analgesic intensity is difficult to adjust according to the needs of individual patients. Although continuous infusion of local anesthetics through a catheter can provide long-term pain relief, it is generally not recommended for patients taking anticoagulants.

[0004] Near-infrared thermal stimulation has been used to improve pain management in remotely triggered pain management systems, but its effectiveness is limited by the low tissue penetration of near-infrared light (NIR). In contrast, magnetic hyperthermia (MHT) can be used as an effective on-demand pain management method with a controllable trigger-release pattern due to its deep tissue penetration.

[0005] The invention provides a reversible liquid-solid switching magnetic response gel (PLGA gel), which shows stability and biocompatibility both in vitro and in vivo.

[0006] A PLGA gel loaded with superparamagnetic Fe3O4 and small molecule local anesthetic levobupivacaine (Levobupi), namely Levobupivacaine / Fe3O4@PLGA or Levobupi / Fe3O4@PLGA. Levobupi / Fe3O4@PLGA exhibits an ideal magnetocaloric effect after loading superparamagnetic Fe3O4, and at the same time, PLGA can achieve a rapid liquid-solid phase transition process to prolong the release time of levobupivacaine. In addition, Levobupi / Fe3O4@PLGA is able to initiate a reversible solid-liquid phase transition technology to expand the pore size under the action of magnetocaloric effect. The changed pore size can promote the release of levobupivacaine under mild thermal stimulation and provide enhanced pain relief in vivo. The synergistic effect of magnetic field and Levobupi / Fe3O4@PLGA can trigger the release of levobupivacaine by magnetocaloric effect. The experimental results showed that Levobupi / Fe3O4@PLGA combined with MHT could significantly relieve local pain and had adjustable intensity properties. Summary of the invention

[0007] In view of the fact that existing local anesthetics have insufficient action time and cannot achieve controlled drug release, the present invention provides a new injectable long-acting local anesthetic - a PLGA hydrogel containing Fe3O4 and levobupivacaine. The drug can be injected into a living body, and after injection, the drug release efficiency is slowed down by liquid-solid phase transition, and heated under the action of a magnetic field to become liquid again, thereby improving the drug release efficiency and analgesic effect. While reducing dependence on opioids, the effect of pain treatment is improved.

[0008] To achieve the above object, the technical solution of the present invention is as follows:

[0009] The invention discloses a PLGA gel used as an injectable long-acting local anesthetic, which contains Fe3O4 and levobupivacaine, and is a PLGA gel containing superparamagnetic Fe3O4 particles and small molecule local anesthetic levobupivacaine (Levobupivacaine, referred to as Levobupi).

[0010] The PLGA hydrogel containing Fe3O4 and levobupivacaine can be used as an injectable long-acting local anesthetic. After being injected into a living body, the hydrogel can be changed from liquid to solid, thereby slowing down the release efficiency of the drug and prolonging the local action time of the hydrogel.

[0011] The preparation method of the above-mentioned PLGA hydrogel containing Fe3O4 and levobupivacaine is as follows:

[0012] PLGA (PLGA, molecular weight: 40,000Da, 50:50) was dissolved in N-methylpyrrolidone (NMP) at a mass / volume ratio of 0.5-0.6. The mixture was homogenized in an oscillating incubator at 35°C-45°C overnight to form a uniform gel matrix. 5 mg of levobupivacaine was added to 100 mg of PLGA matrix and mixed well to prepare levobupivacaine@PLGA gel. At a mass ratio of 5-20 wt% Fe3O4, Fe3O4 particles were uniformly dispersed into the levobupivacaine@PLGA matrix by mechanical stirring to prepare Levobupi / Fe3O4@PLGA, i.e., PLGA hydrogel containing Fe3O4 and levobupivacaine.

[0013] Compared with the prior art, the invention has the following beneficial effects: simple synthesis and easy industrialization. The drug release rate is adjusted by the phase change of the hydrogel to achieve ultra-long-acting analgesia, with an analgesic time of >20 days. The gel can be loaded with a variety of local anesthetics such as levobupivacaine, and can be used for pain management in various clinical pain scenarios. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] Figure 1 .Morphological characteristics of Levobupi / Fe3O4@PLGA.

[0015] Figure 2 .Scanning electron microscope image of Levobupi / Fe3O4@PLGA after magnetic field heating.

[0016] Figure 3 .Hysteresis curves of Levobupi / Fe3O4@PLGA and Fe3O4.

[0017] Figure 4 .Biological safety of Levobupi / Fe3O4@PLGA before and after magnetic field heating. Green represents living cells. Red represents dead cells.

[0018] Figure 5 .Evaluation of the retention time of DiR-labeled Levobupi / Fe3O4@PLGA in mice.

[0019] Figure 6 .Results of mechanical allodynia in mice after different treatments.

[0020] Figure 7 Results of mechanical allodynia in mice before and after magnetic field heating. The arrows in the figure represent the time of magnetic field application.

[0021] Figure 8 . Quantitative temperature curves of PBS and levobupivacaine / Fe3O4@PLGA (doped with different mass fractions of Fe3O4).

[0022] Fig. 9 .The corresponding quantitative temperature curves of different volumes of 10% Fe3O4-levobupi / Fe3O4@PLGA.

[0023] Fig.10 .H&E stained sections of muscles after different treatments.

[0024] Fig.11 .Levobupi / Fe3O4@PLGA exhibits liquid-solid phase transition ability and becomes solid after being injected into water.

[0025] Fig.12 XRD spectra of PLGA, Fe3O4, Levobupivacaine and Levobupi / Fe3O4@PLGA showed that the crystal structure of the loaded materials did not change, indicating that the encapsulation process maintained the structural integrity of each component. DETAILED DESCRIPTION

[0026] Example 1: Preparation of Levobupi / Fe3O4@PLGA

[0027] PLGA (PLGA, molecular weight: 40,000Da, 50:50) was dissolved in N-methylpyrrolidone (NMP) at a mass / volume ratio of 0.5-0.6. The mixture was homogenized in a shaking incubator at 37°C overnight to form a uniform gel matrix. A predetermined amount of levobupivacaine (5mg) was added to 100mg of PLGA matrix; mixed well to prepare levobupivacaine@PLGA gel. Fe3O4 particles (5-20mg) were uniformly dispersed into the levobupivacaine@PLGA matrix by stirring at a mass ratio of 10wt% Fe3O4 to prepare Levobupi / Fe3O4@PLGA.

[0028] Example 2: Release Assessment

[0029] The gel was encapsulated in a dialysis bag (concentration 5 mg / mL, molecular cutoff 3.5 KDa) and immersed in 20 mL of distilled water and shaken at 160 rpm in a constant temperature shaker to ensure the consistency of the environment and the repeatability of the experiment. In the AMF (alternating magnetic field) exposure experiment, the setting parameters of the AMF generator should be clearly specified, such as frequency 626 kHz, output current 21.6 A, coil number 2 turns, coil length 1 cm, coil diameter 3 cm, and field strength 5.72 K / Am. The accurate recording and execution of these parameters are essential to ensure the accuracy of the experimental results. Each AMF application lasted for 15 minutes, and the whole process was repeated six times to induce mild thermal effects and promote the release of levobupivacaine. 5 mL of release medium samples were collected regularly (e.g., every hour) and the concentration of levobupivacaine was quantitatively analyzed using UV-vis spectroscopy. The passive release curve can be evaluated by comparing the control samples with and without AMF exposure. The cumulative release of all samples will be analyzed by UV-vis absorbance measurement together with the standard calibration curve to calculate the released concentration of levobupivacaine.

[0030] Example 3: Safety Assessment

[0031] Safety evaluation of Levobupi / Fe3O4@PLGA: To visually observe the live and dead status of cells, DRG cells were seeded in a 35 mm diameter confocal culture dish (2×10^5 cells per dish) and co-cultured with Levobupivacaine / Fe3O4@PLGA for 4 hours at room temperature. After AMF treatment, the cells were incubated with calcein-AM (2×10^6μM) and propidium iodide (PI, 4×10^6μM) for 30 minutes and maintained at 37°C with 5% CO2. After incubation, the cells were washed with PBS, and the live and dead cells were observed and evaluated using a laser confocal microscope (LSM 710, Zeiss).

[0032] Example 4: Evaluation of the magnetocaloric effect of Levobupivacaine / Fe3O4@PLGA.

[0033] To ensure an exhaustive evaluation of the magnetothermal response of Levobupivacaine / Fe3O4@PLGA, the study was conducted in saline solution by varying the mass ratio of Fe3O4 nanoparticles (5%, 10%, and 20%) and adjusting the gel volume (50μL, 100μL, and 200μL). The thermal response upon exposure to an alternating magnetic field (AMF) was quantitatively measured using a far-infrared thermometer. The resulting temperature data was captured and analyzed by thermal imaging software.

[0034] Example 5: Establishment of a neuropathic pain model

[0035] First, anesthesia was induced by 3.5% isoflurane, and anesthesia was maintained by 1.2% isoflurane. An incision of approximately 0.5 cm long was made in the left thigh of the mouse to completely expose the sciatic nerve and its main branches. The tibial nerve and common peroneal nerve were then ligated and cut, and the distal 2 to 4 mm segment of each nerve was removed, while the superficial peroneal nerve was left unchanged.

[0036] Example 6: In vivo mechanical hyperalgesia analysis.

[0037] A mechanical hyperalgesia tester was used to evaluate the responsiveness of mice to non-noxious mechanical stimuli. First, the mice were placed on a wire mesh floor in a transparent cage to adapt to the test environment, and this process lasted at least 30 minutes. During the test, a probe was used to touch the dorsal side of each mouse's hind paw with gradually increasing force. The force when the mouse retracted its paw was recorded, and the force value was recorded as the paw mechanical threshold (PWT). To ensure the reliability of the results, this operation was repeated five times for each mouse, and the average PWT of each animal was then calculated for detailed analysis.

[0038] Formula table of different ratios of PLGA and Fe3O4.

[0039]

[0040] Table 1.Various formulations of Levobupivacaine / Fe3O4@PLGA gels with optimal injectability.

[0041] Control experiments with saline solutions containing no magnetic material showed negligible temperature changes. Samples containing 10% and 20% mass fractions of Fe3O4 reached temperatures of approximately 45°C, with the 20% concentration sample clearly exceeding 55°C in the surrounding water bath. In contrast, the 5% concentration did not reach these temperatures, indicating a threshold effect in the magnetic heating capability. The increase in volume of 10% Zolpidem / Fe3O4@PLGA enhanced the magnetocaloric effect, however, the 200 μL sample showed a rapid temperature rise exceeding 55°C, which is considered unsuitable for mild magnetic hyperthermia for pain management ( Fig. 9 The optimal configuration for further experiments was determined to be 100 μL of Levobupi / 10%-Fe3O4@PLGA, which reliably maintained a temperature of 45°C. This setup was tested in vitro on perineurial tissue to mimic treatment conditions without damaging adjacent healthy tissue.

Claims

1. A PLGA gel for use as an injectable long-acting local anesthetic, characterized in that: The invention contains Fe3O4 and levobupivacaine, and comprises a PLGA gel loaded with superparamagnetic Fe3O4 particles and a small molecule local anesthetic levobupivacaine Levobupi.

2. The use of the PLGA hydrogel as an injectable long-acting local anesthetic according to claim 1, characterized in that: After being injected into the body, the hydrogel changes from liquid to solid, thereby slowing down the release efficiency of the drug and prolonging the local action time of the hydrogel.

3. The method for preparing the PLGA hydrogel as an injectable long-acting local anesthetic according to claim 1, characterized in that it comprises the following steps: PLGA (PLGA, molecular weight: 40,000Da, 50:50) was dissolved in N-methylpyrrolidone (NMP) at a mass / volume ratio of 0.5-0.6; the mixture was homogenized in an oscillating incubator at 35°C-45°C overnight to form a uniform gel matrix; 5 mg of levobupivacaine was added to 100 mg of the PLGA matrix and mixed evenly to prepare levobupivacaine@PLGA gel; in the case of a mass ratio of 5-20wt% Fe3O4, Fe3O4 particles were evenly dispersed in the levobupivacaine@PLGA matrix by stirring to prepare Levobupi / Fe3O4@PLGA, i.e., PLGA hydrogel containing Fe3O4 and levobupivacaine.

4. The method for preparing the PLGA hydrogel as an injectable long-acting local anesthetic according to claim 1, characterized in that: PLGA (PLGA, molecular weight: 40,000Da, 50:50) was dissolved in N-methylpyrrolidone (NMP) at a mass / volume ratio of 0.5-0.6; the mixture was homogenized in an oscillating incubator at 37°C overnight to form a uniform gel matrix; 5 mg of levobupivacaine was added to 100 mg of PLGA matrix; mixed evenly to prepare levobupivacaine@PLGA gel; at a mass ratio of 10 wt% Fe3O4, Fe3O4 particles (5-20 mg) were uniformly dispersed in the levobupivacaine@PLGA matrix by stirring to prepare Levobupi / Fe3O4@PLGA.

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