An anionic surfactant-chitosan oligosaccharide complex assembly and its application

Abstract

This invention discloses an anionic surfactant-chitosan oligosaccharide composite assembly and its application, belonging to the field of mouthwash technology. The composite assembly provided by this invention is formed by the co-assembly of chitosan oligosaccharide and anionic surfactant through electrostatic interactions. The positive charge of the chitosan oligosaccharide can disrupt bacterial cell membranes, while the hydrophobic chain of the anionic surfactant can penetrate bacterial lipid membranes. The two synergistically significantly enhance broad-spectrum antibacterial activity against Gram-positive / negative bacteria and exhibit inflammation-induced pH-responsive antibacterial activity: reducing antibacterial strength in healthy oral environments, thereby reducing drug side effects; and significantly enhancing antibacterial ability in the slightly acidic, sub-healthy oral environment, thus effectively killing bacteria. Simultaneously, this invention avoids the use of pH adjusters, making it more suitable for the actual needs of mouthwashes in the oral care field for highly efficient and stable antibacterial agents.

Description

Technical Field

[0001] This invention relates to the field of mouthwash technology, and in particular to an anionic surfactant-chitosan oligosaccharide complex assembly and its application. Background Technology

[0002] Mouthwash, as a common oral care product, has become increasingly important in the field of oral health in recent years. It effectively cleans the mouth, prevents tooth decay, inhibits plaque, and improves bad breath, and is widely used in home, personal care, and professional dental settings. However, many mouthwash products on the market currently have significantly insufficient antibacterial properties.

[0003] On the one hand, some mouthwashes have insufficient antibacterial effects and poor long-lasting effects. They can only inhibit oral bacteria for a short period of time, and bacteria can easily regenerate over time, making it difficult to maintain oral health in the long term. For example, some traditional mouthwashes mainly rely on ingredients such as alcohol and fragrances to achieve temporary fresh breath, but their ability to kill oral pathogens is limited, and they cannot fundamentally solve the problem of oral bacteria.

[0004] On the other hand, the antibacterial effect of existing mouthwash products is unstable under different pH conditions. Oral pH fluctuates due to factors such as diet and oral diseases; the normal oral pH is generally between 6.5 and 7.5. Many mouthwashes have acceptable antibacterial properties at near-neutral or slightly alkaline pH levels, but their antibacterial effect decreases significantly once the pH deviates from this range, especially in acidic environments. This makes them difficult to adapt to the dynamic changes in oral pH, limiting their application effectiveness.

[0005] Against this backdrop, developing an assembly with highly effective antibacterial properties at a specific pH value and suitable for mouthwash is of significant practical importance. Chitosan oligosaccharides, as a natural marine bioactive substance, possess good biocompatibility, biodegradability, and certain antibacterial activity, and have attracted widespread attention in the pharmaceutical, food, and cosmetic fields in recent years. Its antibacterial mechanism mainly stems from the interaction between the amino and hydroxyl groups on the chitosan molecular chain and the negative charge on the bacterial cell wall surface, thereby disrupting the integrity of the bacterial cell wall. However, the antibacterial effect of chitosan oligosaccharides used alone is often limited, making it difficult to meet the demand for highly effective antibacterial properties in mouthwash.

[0006] Several technologies have been developed to prepare chitosan oligosaccharide composite materials and mouthwashes. For example, Chinese patent CN116726193A discloses a method for preparing an attapulgite nano-hybrid antibacterial material modified with Isatis indigotica-chitosan oligosaccharide. The antibacterial material prepared by this method has high stability, good biocompatibility, and exhibits strong antibacterial activity against both Gram-positive and Gram-negative bacteria. However, its preparation process is complex, increasing production costs.

[0007] Chinese patent CN118304247A discloses a method for preparing a chitosan oligosaccharide antibacterial and moisturizing waterless hand sanitizer. This method utilizes a composite chitosan oligosaccharide solution, thickener, emollient, skin-enhancing factor, toner, surfactant, mixed traditional Chinese medicine extract, essential oil, and other raw materials to prepare the hand sanitizer. However, the chitosan oligosaccharide polycations are only simply incorporated, and their effect is very limited.

[0008] Chinese patent CN119424303A discloses a mouthwash composition containing effective ingredients such as Lactobacillus salivarius, domiphene, pH adjuster, stabilizer, and preservative. Domiphene can achieve highly effective bactericidal effects, but it only works under neutral conditions; therefore, a pH adjuster needs to be added to adjust the pH of the mouthwash composition to 6.8-7.2.

[0009] Surfactants, with their advantages of high surface activity, low irritation, and easy degradation, have broad application prospects in cleaning products. However, while anionic surfactants offer advantages in biocompatibility, their antibacterial spectrum is narrow, and their action mainly relies on disrupting microbial cell membranes, resulting in limited effectiveness against certain Gram-negative bacteria or drug-resistant bacteria. Furthermore, surfactants are easily rinsed or diluted, making it difficult to maintain long-lasting antibacterial effects. The need for high concentrations of surfactants to reach an effective bactericidal threshold when used alone limits their application in biomedical settings.

[0010] Currently, antibacterial technologies based on anion / cation co-assembly have attracted widespread attention in various fields such as materials science and biomedicine, with relevant research and application examples already existing in areas such as nanomaterial preparation and drug delivery system construction. For example, the co-assembly of cationic antimicrobial peptides with anionic nanomaterials can effectively improve the stability and antimicrobial activity of antimicrobial peptides, while simultaneously achieving targeted killing of bacteria. Anion / cation co-assembly technology exhibits unique advantages in the field of antibacterial applications. Through principles such as electrostatic interactions, it can achieve synergistic effects of multiple antimicrobial components, enhancing antimicrobial efficacy while reducing the development of bacterial resistance.

[0011] Based on this, this invention proposes an inflammation-induced anionic surfactant-chitosan oligosaccharide anionic / cationic synergistic antibacterial assembly, aiming to optimize its pH-responsive antibacterial properties under inflammation-induced conditions: reducing antibacterial strength in a healthy oral cavity to minimize drug side effects; and significantly enhancing antibacterial ability in a slightly acidic, sub-healthy oral environment to achieve effective sterilization. This invention avoids the use of pH adjusters, making it more suitable for the actual needs of mouthwashes in the oral care field for highly effective and stable antibacterial agents. Summary of the Invention

[0012] The purpose of this invention is to provide an anionic surfactant-chitosan oligosaccharide complex assembly and its application, which can effectively enhance the antibacterial efficacy of mouthwash and avoid the use of pH adjusters, thus having broad market application prospects.

[0013] To achieve the above objectives, on the one hand, the present invention provides an anionic surfactant-chitosan oligosaccharide composite assembly, wherein the anionic surfactant and chitosan oligosaccharide are electrostatically co-assembled into a composite assembly.

[0014] Preferably, the anionic surfactant includes one of sodium dodecylbenzenesulfonate, sodium dodecyl sulfate, sodium lauroyl glutamate, and sodium laurylate.

[0015] On the other hand, the present invention provides a method for preparing the above-mentioned anionic surfactant-chitosan oligosaccharide complex assembly: it is prepared by mixing 4 mg / mL chitosan oligosaccharide aqueous solution and 4 mg / mL sodium dodecylbenzenesulfonate aqueous solution in a molar ratio of 1-2:1-2 and stirring until homogeneous.

[0016] Preferably, it is prepared by mixing 4 mg / mL chitosan oligosaccharide aqueous solution and 4 mg / mL sodium dodecylbenzenesulfonate aqueous solution in a molar ratio of 1:1 and stirring until homogeneous.

[0017] On the other hand, the present invention provides the application of an anionic surfactant-chitosan oligosaccharide complex assembly as described above in the preparation of mouthwash.

[0018] Therefore, the anionic surfactant-chitosan oligosaccharide complex assembly and its application of the present invention have the following beneficial effects:

[0019] (1) Cationic chitosan oligosaccharide and anionic surfactant form a stable composite structure through electrostatic co-assembly. The positive charge of chitosan oligosaccharide can destroy bacterial cell membranes, while the hydrophobic chain of surfactant can effectively penetrate lipid membranes. The two work together to significantly enhance the broad-spectrum antibacterial activity against Gram-positive / negative bacteria.

[0020] (2) pH-responsive in vitro antibacterial test of anionic surfactant-chitosan oligosaccharide complex assembly showed that the anionic surfactant-chitosan oligosaccharide complex assembly exhibited significant antibacterial activity against both Gram-negative and Gram-positive bacteria under inflammatory-induced acidic conditions, while remaining highly stable in healthy oral cavity, thereby reducing drug loss and side effects.

[0021] (3) This invention avoids the use of pH adjusters, making it more in line with the actual needs of mouthwash in the field of oral care for efficient and stable antibacterial agents.

[0022] The technical solution of the present invention will be further described in detail below with reference to the accompanying drawings and embodiments. Attached Figure Description

[0023] To more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings used in the description of the embodiments of the present invention will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0024] Figure 1 Particle size analysis of different anionic surfactant / chitosan oligosaccharide assemblies (molar ratio 1:1) prepared in Examples 1-4;

[0025] Figure 2 Zeta potential analysis of different anionic surfactants in Examples 1-4;

[0026] Figure 3 Zeta potential analysis of different anionic surfactant / chitosan oligosaccharide assemblies (molar ratio 1:1) prepared in Examples 1-4;

[0027] Figure 4 Fourier transform infrared spectra of sodium dodecylbenzenesulfonate (SDBS), chitosan oligosaccharide (COS), and the sodium dodecylbenzenesulfonate / chitosan oligosaccharide (SDBS / COS) assembly (molar ratio 1:1) in Example 1;

[0028] Figure 5 The inhibition rates of sodium dodecylbenzenesulfonate (SDBS), chitosan oligosaccharide (COS), and sodium dodecylbenzenesulfonate / chitosan oligosaccharide (SDBS / COS) assemblies against Escherichia coli at different pH values ​​were determined.

[0029] Figure 6 The inhibition rates of sodium dodecylbenzenesulfonate (SDBS), chitosan oligosaccharide (COS), and sodium dodecylbenzenesulfonate / chitosan oligosaccharide (SDBS / COS) assemblies against Staphylococcus aureus at different pH values ​​were determined.

[0030] Figure 7 The kinetic curves of SDBS / COS against Staphylococcus aureus at pH 6.5 are shown. Detailed Implementation

[0031] The technical solution of the present invention will be further described below with reference to the accompanying drawings and embodiments.

[0032] To make the objectives, technical solutions, and advantages of this application clearer, more thorough, and more complete, the technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings and embodiments. The following detailed descriptions are all illustrations of embodiments, intended to provide further detailed explanation of the present invention. Unless otherwise specified, all technical terms used in this invention have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains.

[0033] A method for preparing an anionic surfactant-chitosan oligosaccharide complex assembly includes: mixing 4 mg / mL chitosan oligosaccharide aqueous solution and 4 mg / mL sodium dodecylbenzenesulfonate aqueous solution in a molar ratio of 1-2:1-2 and stirring until homogeneous.

[0034] The examples tested the antibacterial effect of the anionic surfactant-chitosan oligosaccharide complex assembly at different pH values ​​by adjusting the pH of the chitosan oligosaccharide aqueous solution and the sodium dodecylbenzenesulfonate aqueous solution.

[0035] The instruments, equipment, reagents, and materials used in the examples were all obtained commercially.

[0036] Example 1

[0037] An anionic surfactant-chitosan oligosaccharide complex assembly is prepared by the following steps:

[0038] S1. Add 0.2g of chitosan oligosaccharide (COS) and 0.2g of sodium dodecylbenzenesulfonate (SDBS) to 50mL of deionized water, stir evenly, and sonicate for 1h to fully dissolve them to obtain chitosan oligosaccharide mother liquor and sodium dodecylbenzenesulfonate mother liquor, and adjust them to pH=6.5 respectively.

[0039] S2. Mix the chitosan oligosaccharide mother liquor with the sodium dodecylbenzenesulfonate mother liquor at a molar ratio of 2:1, 1:1, or 1:2. Vortex the mixed solution for 5 minutes to obtain the sodium dodecylbenzenesulfonate / chitosan oligosaccharide assembly (SDBS / COS).

[0040] The structural formula of chitosan oligosaccharide is as follows:

[0041] .

[0042] The structural formula of sodium dodecylbenzenesulfonate is as follows:

[0043] .

[0044] Example 2

[0045] An anionic surfactant-chitosan oligosaccharide composite assembly was prepared using the same method as in Example 1, except that sodium dodecyl sulfate (SDS) was selected as the anionic surfactant, ultimately yielding a sodium dodecyl sulfate / chitosan oligosaccharide assembly (SDS / COS).

[0046] The structural formula of sodium dodecyl sulfate is shown below:

[0047] .

[0048] Example 3

[0049] An anionic surfactant-chitosan oligosaccharide complex assembly was prepared using the same method as in Example 1, except that sodium lauroyl glutamate (SLG) was selected as the anionic surfactant, ultimately yielding a sodium lauroyl glutamate / chitosan oligosaccharide assembly (SLG / COS).

[0050] The structural formula of sodium lauroyl glutamate is shown below:

[0051] .

[0052] Example 4

[0053] An anionic surfactant-chitosan oligosaccharide complex assembly was prepared using the same method as in Example 1, except that sodium laurate (SL) was selected as the anionic surfactant, ultimately yielding a sodium laurate / chitosan oligosaccharide assembly (SL / COS).

[0054] The structural formula of sodium lauryl nitrate is shown below:

[0055] .

[0056] Example 5

[0057] An anionic surfactant-chitosan oligosaccharide composite assembly was prepared using the same method as in Example 1, except that the chitosan oligosaccharide mother liquor and sodium dodecylbenzenesulfonate mother liquor were used and adjusted to pH 5.5 respectively.

[0058] Example 6

[0059] An anionic surfactant-chitosan oligosaccharide composite assembly was prepared using the same method as in Example 1, except that the chitosan oligosaccharide mother liquor and sodium dodecylbenzenesulfonate mother liquor were used and adjusted to pH 7.5 respectively.

[0060] Example 7

[0061] An anionic surfactant-chitosan oligosaccharide composite assembly was prepared using the same method as in Example 1, except that the chitosan oligosaccharide mother liquor and sodium dodecylbenzenesulfonate mother liquor were used and adjusted to pH=8.5 respectively.

[0062] Test Example 1

[0063] The particle size of the different anionic surfactant / chitosan oligosaccharide assemblies (molar ratio 1:1) prepared in Examples 1-4 was determined by dynamic light scattering (DLS), and the results are as follows: Figure 1 As shown.

[0064] The results showed that the particle sizes of the sodium dodecylbenzenesulfonate / chitosan oligosaccharide (SDBS / COS), sodium dodecyl sulfate / chitosan oligosaccharide (SDS / COS), sodium lauroyl glutamate / chitosan oligosaccharide (SLG / COS) assemblies, and sodium laurylate / chitosan oligosaccharide assemblies (SL / COS) were 142 nm, 220 nm, 712 nm, and 295 nm, respectively. Even at concentrations far below the critical micelle concentration of the anionic surfactant, they still exhibited large particle size distributions, indicating that the anionic surfactant and chitosan oligosaccharide can obtain assemblies with larger particle sizes through electrostatic interactions.

[0065] Test Example 2

[0066] The Zeta potentials of chitosan oligosaccharide, different anionic surfactants in Examples 1-4, and different anionic surfactant / chitosan oligosaccharide assemblies prepared in Examples 1-4 (molar ratio 1:1) were determined, and the results are as follows: Figures 2-3 As shown.

[0067] Depend on Figure 2 It is known that anionic surfactants all exhibit a negative charge. Analysis Figures 2-3 It can be seen that the charge of the anionic surfactant and chitosan oligosaccharide increases after assembly, indicating that the assembly process can achieve better antibacterial effect.

[0068] Test Example 3

[0069] Fourier transform infrared spectra of sodium dodecylbenzenesulfonate (SDBS), chitosan oligosaccharide (COS), and the sodium dodecylbenzenesulfonate / chitosan oligosaccharide (SDBS / COS) assembly (molar ratio 1:1) in Example 1 were determined, and the results are as follows: Figure 4 As shown. Analysis reveals that COS values ​​of 1541 and 1404 cm... -1 The blue peaks representing amide bonds shifted to 1549 and 1409 cm⁻¹. -1 Location; 1021cm -1 The peak representing the glycosidic bond shifts to 1032 cm⁻¹. -1 This may be because the spatial position and orientation of the amide group (-CONH2) and glycosidic bond in COS may change after electrostatic interaction with SDBS, resulting in a change in vibrational frequency.

[0070] Test Example 4

[0071] The in vitro antibacterial effects of sodium dodecylbenzenesulfonate (SDBS), chitosan oligosaccharide (COS), and sodium dodecylbenzenesulfonate / chitosan oligosaccharide (SDBS / COS) assemblies (molar ratios 1:1 and 1:2) at different pH values ​​were determined in Examples 1, 5-7.

[0072] Escherichia coli ( Escherichia coli ) and Staphylococcus aureus ( Staphalococcus aureus Purchased from the China General Microbiological Culture Collection Center (CGMCC) website. Streaked on agar nutrient medium and incubated overnight at 37°C. Single clones of *Escherichia coli* and *Staphylococcus aureus* were picked and inoculated into broth culture medium, and incubated at 37°C and 220 rpm for 8 hours. Afterwards, the cultured *E. coli* and *Staphylococcus aureus* were cultured in broth until 10⁻⁶ mmol / L. 6 The concentration of CFU / mL was used for in vitro antibacterial experiments.

[0073] COS, SDBS, and SDBS / COS assemblies at different pH values ​​from Examples 1, 5-7 were respectively mixed with 10 7 Equal volumes of Escherichia coli and Staphylococcus aureus at a concentration of CFU / mL were mixed and incubated together at 37°C and 220 rpm for 2 hours.

[0074] The bacterial suspension, after co-incubation, was diluted with sterile water and plated onto nutrient agar plates. The plates were then incubated at 37°C for 20 hours, and the colony count was performed. The measurements were conducted three times (n=3).

[0075] Figure 5 The inhibition rates of COS, SDBS, and SDBS / COS assemblies against Escherichia coli under different pH conditions were shown. It can be seen that the antibacterial effect of the SDBS / COS assembly is significantly higher than that of COS and SDBS alone. Furthermore, the antibacterial efficiency at a slightly acidic pH of 6.5 is higher than that at 7.5 and 8.5. At pH 6.5, the antibacterial effect of the SDBS / COS assembly prepared with a COS / SDBS molar ratio of 1:1 is significantly higher than that of the SDBS / COS assembly prepared with a COS / SDBS molar ratio of 1:2.

[0076] Figure 6 The inhibition rates of COS, SDBS, and SDBS / COS assemblies against Staphylococcus aureus under different pH conditions were shown. It can be seen that the antibacterial effect of the SDBS / COS assembly is significantly higher than that of COS and SDBS alone, and the antibacterial efficiency at a slightly acidic pH of 6.5 is higher than that at 7.5 and 8.5. At pH 6.5, the antibacterial effect of the SDBS / COS assembly prepared with a COS / SDBS molar ratio of 1:1 is significantly higher than that of the SDBS / COS assembly prepared with a COS / SDBS molar ratio of 1:2.

[0077] Figure 7 The image shows the bactericidal kinetics curve of SDBS / COS against Staphylococcus aureus at pH 6.5. At a concentration of 4×MIC, it can achieve 100% antibacterial rate in 15 minutes, which shows rapid bactericidal efficiency.

[0078] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and not to limit them. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can still be made to the technical solutions of the present invention, and these modifications or equivalent substitutions cannot cause the modified technical solutions to deviate from the spirit and scope of the technical solutions of the present invention.

Claims

1. An anionic surfactant-chitooligosaccharide complex assembly, characterized by: Anionic surfactant and chitooligosaccharide are electrostatically co-assembled into a complex assembly; The anionic surfactant is sodium dodecyl benzene sulfonate; The anionic surfactant-chitooligosaccharide complex assembly is prepared by mixing 4 mg / mL chitooligosaccharide aqueous solution and 4 mg / mL sodium dodecyl benzene sulfonate aqueous solution at a molar ratio of 1-2:1-2 and then stirring uniformly.

2. A method of preparing an anionic surfactant-chitosan oligosaccharide complex assembly according to claim 1, characterized by: The anionic surfactant-chitooligosaccharide complex assembly is prepared by mixing 4 mg / mL chitooligosaccharide aqueous solution and 4 mg / mL sodium dodecyl benzene sulfonate aqueous solution at a molar ratio of 1-2:1-2 and then stirring uniformly.

3. The method of preparing an anionic surfactant-chitosan oligosaccharide complex assembly according to claim 2, characterized in that: The anionic surfactant-chitooligosaccharide complex assembly is prepared by mixing 4 mg / mL chitooligosaccharide aqueous solution and 4 mg / mL sodium dodecyl benzene sulfonate aqueous solution at a molar ratio of 1-2:1-2 and then stirring uniformly.

4. Use of the anionic surfactant-chitooligosaccharide complex assembly according to claim 1 in the preparation of a mouthwash.

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