Study on the Free Radical Degradation of Hydroxypropyl Methyl Cellulose in Vitro

Abstract: The Fenton-AH—Hz02 system was used to simulate the reactive oxygen species radicals (ROS) in the body to degrade the cosmetic filling auxiliary material hydroxypropyl methylcellulose (HPMC), and the changes in the molecular weight and dynamic viscosity of HPMC during the degradation process were studied. . It was found that during the degradation process, the molecular weights of E3-HPMC and E50-HPMC decreased by 90%, the dynamic viscosity of E4M-HPMC decreased by 75%, and the dynamic viscosity of E10M-HPMC decreased by 40%. It indicated that ROS had a significant degradation effect on HPMC.

Key words:hydroxypropyl methylcellulose; reactive oxygen species; free radical degradation

Hydroxypropyl methylcellulose (HPMC) is a cellulose ether formed after partial hydroxymethylation and hydroxypropylation of natural cellulose molecules. It is a commonly used pharmaceutical excipient and is generally used as a dispersant, Thickeners, ophthalmic implants, cosmetic filling materials, etc. Domestic hydroxypropyl methylcellulose-sodium hyaluronate solution (Yimei, EME) utilizes the hydroxyl groups in the HPMC molecule to interact with the hydroxyl and carboxyl groups in hyaluronic acid to improve their usability as skin filling materials. It fills the blank of my country's biocomposite materials in the field of medical cosmetology, and its degradation process in the body has therefore attracted extensive attention from researchers.

Reactive oXygen species (ROS) is an important free radical in the body and plays an important role in the body. A large number of ultraviolet (UV) chromophores in the skin are in an excited state after absorbing UV photon energy, and undergo a photodynamic reaction with oxygen molecules in the skin. With the participation of reductases or transition metals, ROS will be generated, and a large amount of ROS will Cause skin aging, produce wrinkles, spots and so on. ROS mainly includes four kinds of free radicals, superoxide anion radical, hydroxyl radical, hydrogen peroxide and singlet oxygen, among which OH has the strongest effect. It has been reported that HPMC can absorb ROS inherent in the body.

HPMC will be degraded under the action of ROS, resulting in the decrease of molecular weight and viscoelasticity of HPMC. The author used the Fenton-AH-H2O2 system to mediate the generation of hydroxyl radicals, simulated the degradation of HPMC by ROS, and characterized the molecular weight and dynamic viscosity changes of HPMC during the degradation process to study the degradation mechanism of HPMC in vivo.

1. Experiment

1.1 Materials, reagents and instruments

Hydroxypropyl methylcellulose [Grade: E3, E50, E4M, E10M, representing the mass fraction of 2% HPMC aqueous solution with dynamic viscosity (mPa·s) of 3, 50, 4000, 10000 respectively], KIMA CHEMICAL CO. , LTD; L-ascorbic acid (AH), phosphate buffer solution with a pH value of 3, Tianjin Guangfu Fine Chemical Factory; 30% hydrogen peroxide, Tianjin Chemical Reagent No. 6 Factory.

Aqueous Gel Permeation Chromatography, Waters Company; NDJ-1 Rotational Viscometer, Shanghai Sendi Scientific Instrument Equipment Co., Ltd.

1.2 Method

Generally, the Fenton reaction caused by the Haber-Weiss reaction is the main way to generate ·OH in vitro.

1.2.1 Detecting the molecular weight change of HPMC during the degradation process

Dissolve 0.5 g HPMC (E3, E50) and 0.125 g L-ascorbic acid in 22.5 mL pH=3 phosphate buffer solution, add 2.5 mL H2O2 with a mass fraction of 3%, place in In a constant temperature oscillator in a water bath at 37°C; sample 0.08 mL regularly, add 0.02 mL of 0.1 mol L-1 NaOH solution to destroy the free radical environment, and measure the molecular weight of the solution by aqueous gel permeation chromatography.

Blank experiment: Dissolve 0.5 g HPMC (E3, E50) and 0.125 gL-ascorbic acid in 25 mL pH=3 phosphate buffer solution; regularly sample 0.08 mL, add 0.02 mL 0.1 m01. The NaOH solution of L-1 was placed in a water bath constant temperature oscillator at 37°C, and the molecular weight of the solution was measured by an aqueous gel permeation chromatography.

1.2.2 Detection of the dynamic viscosity change of HPMC during the degradation process

Dissolve 2 g of HPMC (E4M, E10M) and 0.5 g of L-ascorbic acid in 90 ml, pH=3 phosphate buffer solution, add 10 mL of H2Oz solution with a mass fraction of 3%, place in a constant temperature water bath at 37 C, and use A rotational viscometer measures the viscosity of the solution.

Blank experiment: Dissolve 2 g HPMC (E4M, E10M) and 0.5 g L-ascorbic acid in 100ml phosphate buffer solution with pH=3, place it in a 37C constant temperature water bath, and measure the viscosity of the solution with a rotational viscometer.

2. Results and Discussion

2.1 Changes in molecular weight of HPMC

From the molecular weight change of HPMC with lower dynamic viscosity (E3-HPMC, E50-HPMC) during the degradation process, it can be seen that the average molecular weight of E3-HPMC is 19.4 kDa, and the average molecular weight of E50-HPMC is 110.0 kDa.

From the curves of E3-HPMC and E50-HPMC molecular weight with degradation time, it can be seen that the molecular weight of HPMC decreased rapidly in the first 100 min when H2O2 was added to the HPMC-AH-PBS system. This is due to the rapid reaction between H2O2 and AH in the system, which produces a large amount of OH, which causes the rapid degradation of HPMC and the rapid decrease of molecular weight; when the degradation reaction lasts for about 200 min, due to the large consumption of H2Oz, the concentration of OH in the system Decrease, and gradually lose the degradation effect on HPMC, so in the later stage of the experiment, the molecular weight of HPMC tends to be stable. At the end of the degradation, the final molecular weight of E3-HPMC was stable at about 1 kDa, which was 90% lower than that before degradation; the final molecular weight of E50-HPMC was stable at about 10 kDa, which was also 90% lower than that before degradation.

2.2 Changes in dynamic viscosity of HPMC

For E4M-HPMC and E10M-HPMC with high dynamic viscosity, because the molecular weight is too large, it is difficult to track the degradation process with the molecular weight as a reference, so the degradation process is tracked by measuring the dynamic viscosity of the solution.

It can be seen that the dynamic viscosities of E4M-HPMC and E10M-HPMC tend to be stable after 240 min. At the end of degradation, the final viscosity of E4M-HPMC was about 1000 mPa·s, which was 75% lower than that before degradation; the final viscosity of E10M-HPMC was about 6000 mPa·s, which was 40% lower than that before degradation. Its degradation process is similar to that of E3-HPMC and E50-HPMC.

3.Conclusion

By simulating the reactive oxygen species (ROS) conditions in vivo, the degradation process of HPMC was studied in vitro using the Fenton-AH-H2O2 system. The molecular weight and dynamic viscosity of HPMC were detected by aqueous gel permeation chromatography and rotational viscometer. The results showed that the molecular weight of E3-HPMC and E50-HPMC decreased by 90%, and the dynamic viscosity of E4M-HPMC decreased by 75%. The dynamic viscosity of E10M-HPMC decreased by 40%. It shows that ROS has a significant degradation effect on HPMC, and also shows that HPMC can absorb free radicals in the body and reduce the content of free radicals in the body.