(STEE-Cur)-(LF-LMP) quaternary compound for improving curcumin stability

Abstract

The invention discloses a (STECur) (LFLMP) quaternary compound for improving the stability of curcumin, and belongs to the field of food additives and functional ingredients. The preparation method comprises the following steps: by taking LF, LMP, STE and Cur as raw materials, self-assembling to form a quaternary compound (STE Cur) (LFLMP) by utilizing electrostatic and hydrophobic interaction and non-covalent interaction such as hydrogen bonds; by adding the polyelectrolyte compound LFLMP into the binary compound STECur, the Cur can be promoted to enter a deep hydrophobic cavity of STE, and the solubility of the Cur is further enhanced. Meanwhile, the LFLMP can be self-assembled with the STECur to form a quaternary compound (STECur) (LFLMP with a core-shell structure, which takes the STECur as a core and the LFLMP as a shell, so that the light stability, the heat stability and the long-term storage stability of the Cur can be effectively improved. The preparation method is simple, convenient to operate and low in cost, and the obtained product has good stability and functionality.

Description

Technical Field

[0001] The invention relates to a (STE-Cur)-(LF-LMP) quaternary complex for improving the stability of curcumin, and belongs to the field of food additives and functional ingredients. Background Art

[0002] Curcumin (Cur) is a recognized bioactive compound with excellent antioxidant, anti-inflammatory, and antitumor properties. However, as a hydrophobic natural polyphenol, Cur has low solubility in water, poor stability in alkaline and neutral environments, and is easily degraded under conditions such as light and heat, which greatly limits its application in food processing. Biopolymers as delivery systems are an effective means to overcome the challenges of Cur's instability, hydrophobicity, and low bioavailability. Natural active ingredients in food, such as biomacromolecules (proteins, lipids, polysaccharides) and small molecules (peptides, glycosides), have excellent biocompatibility and biodegradability, making them ideal carriers for constructing Cur delivery systems. In addition, the preparation methods of biopolymer delivery systems include self-assembly, antisolvent method, electrospinning, and pH-driven method. In comparison, self-assembly technology is more efficient and environmentally friendly, and provides a better way to develop high-performance encapsulation of bioactive substances.

[0003] Stevioside (STE) is a natural sweetener with biological activity and amphiphilicity, mainly including stevioside and rebaudioside A. The amphiphilicity of STE gives it the ability to act as a surfactant, making it widely used in food and beverage processing. STE can self-assemble into ultrasmall micelles in aqueous solution, thereby enhancing the solubility and stability of hydrophobic polyphenols (such as Cur, capsaicin and resveratrol). For example, Wang et al. constructed micelles and solid dispersions based on STE, significantly improving the solubility of phenanthroline encapsulated in the hydrophobic core of spherical micelles with a diameter of 5 nm; Nguyen et al. used STE to extract Cur from turmeric powder and obtained an aqueous solution of Cur that maintained 80% stability in the pH range of 6.0 to 10.0 and after storage at 25°C for 1 week.

[0004] In the above example, the problem in constructing the Cur delivery system is that the spatial conformation of Cur loaded on a single STE micelle is relatively simple, and due to the weak interfacial interaction between STE and Cur, the formed micelle structure is unstable, resulting in poor light and heat stability and long-term storage stability.

[0005] Therefore, it is necessary to provide a method with simple preparation, diverse structures and good stability to construct a stable Cur complex delivery system.

[0006] Polyelectrolyte complexes based on oppositely charged proteins and polysaccharides can significantly improve the water solubility, stability, and bioavailability of Cur. Cur binds to proteins and polysaccharides through non-covalent interactions such as hydrophobic interactions, hydrogen bonding, and van der Waals forces. Strong interactions between proteins and polysaccharides maintain the steric stability of the protein and may also enhance the stability of encapsulated Cur. Polyelectrolyte complexes formed from various proteins and polysaccharides, including gellan gum / chitosan, chitosan / sodium alginate, and lactoferrin / polysaccharide, have been extensively studied. Multicomponent complexes of lactoferrin and pectin have shown great potential for stabilizing hydrophobic polyphenols, providing new insights into the construction of Cur complex stabilization systems. However, the effects of these reported binary and ternary complexes of proteins and polysaccharides on the photostability and thermal stability of Cur remain limited. To date, no studies have investigated the preparation of self-assembled protein-pectin-stevioside quaternary complexes for encapsulating and stabilizing Cur. Summary of the Invention

[0007] The technical problem to be solved by the present invention is that Cur has low solubility in water, poor stability in alkaline and neutral environments, and is easily degraded under conditions such as light and heat. The spatial conformation of the existing self-assembly system encapsulating Cur is relatively simple, and the structure of the formed self-assembly is unstable, resulting in poor light and heat stability and long-term storage stability of Cur.

[0008] The present invention provides a (STE-Cur)-(LF-LMP) quaternary complex for improving the stability of Cur. The invention uses LF (lactoferrin), LMP (low methyl pectin), STE (stevioside) and Cur as raw materials. Cur and STE are co-assembled to form a STE-Cur binary complex, and LF and LMP are co-assembled to form a LF-LMP binary complex through non-covalent interactions such as electrostatic interaction, hydrophobic interaction and hydrogen bonding. Then, STE-Cur and LF-LMP are self-assembled through non-covalent interactions to form a (STE-Cur)-(LF-LMP) quaternary complex. The quaternary complex can efficiently and stably encapsulate Cur and significantly improve its light stability and long-term storage stability.

[0009] The method for preparing the (STE-Cur)-(LF-LMP) quaternary complex comprises the steps of:

[0010] (1) Prepare LF, LMP, and STE stock solutions separately using 10 mM pH 5.0 acetate buffer and store at 4°C overnight to allow them to fully dissolve. Dissolve Cur in anhydrous ethanol to prepare a Cur stock solution with a concentration of 10-200 mM and store it at 4°C in the dark for later use. Add 0.5 mL of Cur stock solution dropwise to 4.5 mL of STE stock solution and stir at 25°C for 2-24 h to prepare STE-Cur complex; mix LF and LMP stock solutions in equal volumes and stir at 25°C for 2-24 h to prepare LF-LMP polyelectrolyte complex.

[0011] (2) Under magnetic stirring, 5 mL of STE-Cur mother solution was slowly added to 5 mL of LF-LMP mother solution, and the mixture was stirred at 25 °C for 2-24 h to obtain the (STE-Cur)-(LF-LMP) quaternary complex.

[0012] The (STE-Cur)-(LF-LMP) quaternary complex is a core-shell structure with the STE-Cur binary complex as the core and the LF-LMP binary complex as the shell.

[0013] The esterification degree of the LMP is 30%-45%.

[0014] The final concentration of STE is 0.5%-5% (w / v).

[0015] The final concentration of ethanol in the quaternary complex system is 1%-5% (v / v), and the final concentration of Cur is 0.5-10 mM. It should be noted that the ethanol concentration in the system is only 1%-5%, so the denaturation of LF induced by ethanol can be ignored.

[0016] The final concentration of LF in the quaternary complex system is 0.1%-2% (w / v), and the final concentration of LMP is 0.5%-5% (w / v).

[0017] In an optional embodiment of the present invention, the quaternary complex is constructed under conditions of a stirring frequency of 50-300 r / min, a stirring time of 2-24 h, and a constant temperature of 25° C.

[0018] The present invention provides a (STE-Cur)-(LF-LMP) quaternary complex that improves the stability of Cur. First, STE can form ultrasmall micelles in an aqueous solution, solubilizing Cur in the hydrophobic cavity of the micelles to form a binary complex STE-Cur. Then, the polyelectrolyte complex LF-LMP is added to STE-Cur. The LF-LMP complex can promote Cur to enter the deep hydrophobic cavity of STE, forming a (STE-Cur)-(LF-LMP) quaternary complex with a tighter structure, thereby effectively improving the stability of the system and the encapsulation and delivery efficiency of Cur. Finally, the obtained (STE-Cur)-(LF-LMP) quaternary complex has a core-shell structure with STE-Cur as the core and LF-LMP as the shell, which can efficiently and stably encapsulate Cur, and significantly improve the stability of Cur through the double-layer protection of STE and LF-LMP.

[0019] The (STE-Cur)-(LF-LMP) quaternary complex constructed by the layer-by-layer self-assembly method of the present invention has the advantages of good biocompatibility, simple preparation method, high delivery efficiency, etc., and can effectively improve the light and heat stability and long-term storage stability of Cur. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 Fluorescence spectra of LF in LF-Cur (A), LF-STE-Cur (B), LF-LMP-Cur (C) and (STE-Cur)-(LF-LMP) quaternary complex (D) under different Cur concentrations.

[0021] Figure 2 UV-visible scanning spectra of Cur (5% EtOH), STE-Cur complex and (STE-Cur)-(LF-LMP) quaternary complex.

[0022] Figure 3 XRD patterns (A), DSC curves (B), TGA curves (C), and DTG curves (D) of Cur, STE-Cur binary complex, LF-LMP binary complex, and (STE-Cur)-(LF-LMP) quaternary complex.

[0023] Figure 4 Scanning electron microscopy (SEM) images (A) and transmission electron microscopy (TEM) images (B) of the STE-Cur binary complex, LF-LMP binary complex and (STE-Cur)-(LF-LMP) quaternary complex.

[0024] Figure 5 Retention rate of Cur in different complexes under UV irradiation conditions (A) and first-order kinetic model of photodegradation (B).

[0025] Figure 6 Retention rate of Cur in different complexes under heating conditions at 80°C (A) and first-order kinetic curve of Cur thermal degradation (B). The final concentrations of LF, LMP, and STE were 0.5%, 1.5%, and 1.5% (w / v), respectively.

[0026] Figure 7 Retention rate of Cur in different complexes under heating conditions at 80°C (A) and first-order kinetic curve of Cur thermal degradation (B). The final concentrations of LF, LMP, and STE were 2%, 5%, and 5% (w / v), respectively.

[0027] Figure 8 Average particle size (C) and PDI (D) values ​​of the (STE-Cur)-(LF-LMP) quaternary complex after storage in the dark at 25°C for 30 days. Values ​​are mean ± SD, n = 3. Different letters indicate significant differences (p < 0.05). DETAILED DESCRIPTION

[0028] Analytical methods:

[0029] 1. Determination of Cur Fluorescence: Cur was dissolved in acetate buffer containing 5% (v / v) ethanol and pure ethanol, abbreviated as Cur (5% EtOH) and Cur (100% EtOH), respectively. Fluorescence spectra of Cur (5% EtOH) and Cur (100% EtOH), as well as the fluorescence spectra of Cur in STE-Cur and (STE-Cur)-(LF-LMP), were measured. The excitation wavelength was 420 nm, the excitation slit width was 5 nm, and the emission slit width was 10 nm. A blank was prepared using acetate buffer containing 5% (v / v) ethanol.

[0030] 2. XRD, DSC and TGA determination of the composite: The XRD spectra of Cur, STE-Cur, LF-LMP and (STE-Cur)-(LF-LMP) were measured by X-ray diffractometer, and the samples were scanned in the 2θ range of 10-40° at a speed of 2° / min. The thermal stability of the samples was determined by thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC). DSC analysis used nitrogen as the protective gas, a heating rate of 10°C / min, a test range of 25-200°C, and a sample mass of 5-10 mg. Under nitrogen protection, the sample was heated from 25°C to 800°C at a heating rate of 10°C / min, and the TGA of the sample was measured. The sample mass was 5 mg.

[0031] 3. Morphology and structural analysis of the complex: The freeze-dried sample was placed in liquid nitrogen and frozen, and a portion was cut off. This portion of the sample was fixed to the sample stage with conductive tape and then coated with gold. Then, the surface morphology and structure of the sample were observed using a scanning electron microscope (SEM); the morphology and particle size of the aggregates were characterized by a transmission electron microscope (TEM). First, the sample was diluted 50 times, and the copper mesh was placed in the diluted sample solution for 2 seconds, and then transferred to liquid nitrogen for freezing. It was then freeze-dried for 24 hours. Finally, the sample was observed under a 200kV transmission electron microscope.

[0032] 4. Determination of UV stability of Cur in different complexes: Place samples of Cur (5% EtOH), STE-Cur, LF-LMP-Cur, (STE-Cur)-(LF-LMP) in a sealed and transparent container and irradiate them with a 12W UV lamp at 25°C in the dark for 6h, 12h, 18h and 24h respectively. Calculate the retention rate of Cur under different irradiation times. Calculate the first-order kinetic degradation constant (k) and half-life (t) of Cur according to formula (1) and formula (2). 1 / 2 ):

[0033] ln(C / C0)=-kt (1)

[0034] t 1 / 2 =ln2 / -k (2)

[0035] Where C0 and C are the initial content of Cur and the content under different UV irradiation times, respectively.

[0036] Example 1 Fluorescence Spectrum of (STE-Cur)-(LF-LMP) Quaternary Complex

[0037] Prepare LF, LMP, and STE stock solutions in 10 mM acetate buffer (pH 5.0) and store at 4°C overnight to allow for complete dissolution. Prepare the STE-Cur stock solution by mixing 4.5 mL of the STE stock solution with 0.5 mL of the Cur stock solution and stirring at 25°C for 2 hours. Prepare the LF-LMP stock solution by mixing 2.5 mL of the LF stock solution with 2.5 mL of the LMP stock solution and stirring at 25°C for 2 hours. Slowly pour the STE-Cur stock solution into the LF-LMP stock solution and stir thoroughly for 2 hours to form the (STE-Cur)-(LF-LMP) complex. The final concentrations of LF, LMP, and STE are 0.5%, 1.5%, and 1.5% (w / v), respectively, and the ethanol concentration is 5% (v / v).

[0038] The fluorescence spectrum of Cur in the obtained (STE-Cur)-(LF-LMP) quaternary complex is shown in Figure 1As shown in the figure, it can be seen that the maximum fluorescence emission wavelength of Cur in the (STE-Cur)-(LF-LMP) quaternary complex is located at 467nm, which is significantly blue-shifted (about 37nm) compared to Cur in 5% ethanol and red-shifted (about 8nm) compared to pure ethanol. Obviously, this is closely related to the change of Cur's solvent environment and its interaction with LF, STE and LMP. Combined with the UV-visible scanning spectroscopy results ( Figure 2 ) shows a significant increase in Cur solubility, indicating that the presence of LF and LMP further enhances Cur solubility. Furthermore, compared to Cur alone (5% EtOH), the maximum absorption peak of Cur in the complex undergoes a significant blue shift, indicating a more hydrophobic microenvironment for Cur within the complex. This suggests that the LF-LMP complex facilitates Cur's entry into the deep, hydrophobic cavities of the STE and enhances system stability.

[0039] Comparative Example 1: The (STE-Cur)-(LF-LMP) quaternary complex was replaced by the STE-Cur binary complex. The other conditions were the same as those in the control.

[0040] Example 1

[0041] Prepare a 1.58% (w / v) STE stock solution in 10 mM acetate buffer (pH 5.0) at 25°C. Dissolve Cur in anhydrous ethanol to prepare a 10.8 mmol / L Cur stock solution. Store at 4°C in the dark until ready for use. Add 0.5 mL of the Cur stock solution dropwise to 9.5 mL of the STE stock solution at 25°C and stir for 2 hours to form a STE-Cur complex. The final STE concentration in the STE-Cur complex is 1.5% (w / v) and the ethanol concentration is 5% (v / v).

[0042] The fluorescence spectrum of Cur in the obtained STE-Cur binary complex is as follows: Figure 1 As shown in the figure, the maximum emission wavelength of the STE-Cur binary complex is located at 506 nm, which does not change much compared with that of single Cur (5% EtOH), but is significantly red-shifted compared with the (STE-Cur)-(LF-LMP) quaternary complex, indicating that the structure of STE alone is simple and the interfacial interaction between STE and Cur is weak, resulting in an unstable micelle structure. Figure 2 The results of UV-visible scanning spectroscopy further showed that the maximum absorption peak of Cur in the (STE-Cur)-(LF-LMP) quaternary complex was blue-shifted compared with that in the STE-Cur binary complex, indicating that the microenvironment of Cur in the (STE-Cur)-(LF-LMP) quaternary complex was more hydrophobic, while the hydrophobic environment provided by the STE-Cur micelle system was weak, and the micelle structure of Cur stabilized by STE alone was unstable.

[0043] Example 2 XRD, DSC, TGA and DTG curves of the (STE-Cur)-(LF-LMP) quaternary complex

[0044] The (STE-Cur)-(LF-LMP) quaternary complex in Example 1 was characterized and analyzed by XRD, TGA and DSC. The XRD pattern of the obtained (STE-Cur)-(LF-LMP) quaternary complex ( Figure 3 A) It can be seen that the characteristic diffraction peaks of Cur in the (STE-Cur)-(LF-LMP) quaternary complex all disappear, indicating that the crystal structure of Cur is destroyed and it is encapsulated in the quaternary complex in an amorphous state. At the same time, the XRD pattern of the (STE-Cur)-(LF-LMP) quaternary complex is similar to that of the LF-LMP binary complex, which further indicates that the LF-LMP binary complex is in the outermost layer of the (STE-Cur)-(LF-LMP) quaternary complex. DSC curve ( Figure 3 B) TGA curve ( Figure 3 C) and DTG curve ( Figure 3 D) obtained similar results, which proves that the structure of the (STE-Cur)-(LF-LMP) quaternary complex is a core-shell structure with the STE-Cur binary complex as the core and the LF-LMP binary complex as the shell, which can further enhance the stability of Cur.

[0045] Comparative Example 2: The (STE-Cur)-(LF-LMP) quaternary complex was replaced by the STE-Cur binary complex. The other conditions were the same as those in the control.

[0046] Example 2

[0047] The STE-Cur binary complex was characterized by XRD, TGA and DSC. Figure 3 A) and DSC curve ( Figure 3 B) It can be shown that Cur is encapsulated in the quaternary complex in an amorphous state. The DTG curve of Cur ( Figure 3D) shows a single main peak, indicating that Cur decomposes via a single mechanism involving overall skeleton fragmentation. Similarly, the STE-Cur binary complex also largely maintains a single mechanism for decomposition, demonstrating that the spatial conformation of Cur loaded on the STE micelles is relatively simple, and due to the weak interfacial interaction between STE and Cur, the resulting micelle structure is unstable. Furthermore, the weight loss rate of the STE-Cur binary complex is significantly greater than that of Cur. This is because Cur forms a complex with STE through hydrogen bonding and hydrophobic interactions. This binding significantly disrupts the original crystal structure of Cur, causing its molecules to loosen and its thermal stability to decrease, leading to thermal decomposition at lower temperatures.

[0048] Example 3 Morphology and Structural Analysis of the (STE-Cur)-(LF-LMP) Quaternary Complex

[0049] The (STE-Cur)-(LF-LMP) quaternary complex in Example 1 was freeze-dried and fractured in liquid nitrogen, and the morphology of the quaternary complex was analyzed by scanning electron microscopy (SEM). The (STE-Cur)-(LF-LMP) quaternary complex in Example 1 was diluted 50 times, and a copper mesh was placed in the diluted sample solution for 2 seconds, then transferred to liquid nitrogen for freeze fixation, and freeze-dried for 24 hours. Finally, the structure and particle size of the quaternary complex were analyzed by transmission electron microscopy (TEM).

[0050] The SEM results of the obtained (STE-Cur)-(LF-LMP) quaternary complex are as follows: Figure 4 As shown in A. It can be seen that the (STE-Cur)-(LF-LMP) quaternary complex presents a smooth layered structure, which indicates that the LF-LMP complex is effectively encapsulated in the outer layer of the STE-Cur complex. TEM ( Figure 4 B) The results show that the particle size of the STE-Cur binary complex is approximately 200 nm, and the (STE-Cur)-(LF-LMP) quaternary complex is a spherical structure with a particle size of 1-2 μm. The particle size is significantly smaller than that of LF-LMP (2-5 μm), indicating that a more compact quaternary complex structure is formed at this time, which can effectively improve the stability of Cur.

[0051] Example 4 UV Stability of Cur in the (STE-Cur)-(LF-LMP) Quaternary Complex

[0052] The (STE-Cur)-(LF-LMP) quaternary complex from Example 1 was placed in a sealed, transparent container and irradiated with a 12W UV lamp at 25°C in the dark for 6, 12, 18, and 24 hours, respectively. The retention rate and half-life of Cur were calculated at different irradiation times.

[0053] The retention rate of Cur in the (STE-Cur)-(LF-LMP) quaternary complex under UV irradiation is as follows Figure 5 As shown. After 24 hours of ultraviolet irradiation, the retention rate of Cur in the (STE-Cur)-(LF-LMP) quaternary complex was 75.3%, corresponding to a half-life of 58.3±16.2h, while the retention rate of Cur in the Cur (5% EtOH) solution alone was only 10.3%, corresponding to a half-life of 7.5±1.4h. This shows that the (STE-Cur)-(LF-LMP) quaternary complex has an excellent protective effect on Cur. This is because in the (STE-Cur)-(LF-LMP) quaternary complex structure, Cur is located in the hydrophobic cavity of STE, and STE provides it with the first layer of protection. The addition of the LF-LMP complex provides a second layer of protection for Cur. The tight core-shell structure of the (STE-Cur)-(LF-LMP) quaternary complex effectively improves the photostability of Cur.

[0054] Control Example 3 UV stability of Cur in STE-Cur binary complex

[0055] The STE-Cur binary complex with the same concentration as in Example 4 was placed in a sealed, transparent container and irradiated with a 12W UV lamp at 25°C in the dark for 6 h, 12 h, 18 h, and 24 h, respectively. The retention rate of Cur at different irradiation times was calculated.

[0056] The retention rate of Cur in the STE-Cur binary complex under UV irradiation is as follows Figure 5 As shown in Figure 3 . After 24 hours of UV irradiation, the retention rate of Cur in the (STE-Cur)-(LF-LMP) quaternary complex was 68.9%, corresponding to a half-life of 34.8±3.5 hours. This indicates that the stability of Cur in the STE-Cur binary complex is significantly lower than that of the (STE-Cur)-(LF-LMP) quaternary complex. This is because the spatial conformation of Cur loaded on the STE micelles is relatively simple, resulting in an unstable micelle structure and poor photostability.

[0057] UV stability of Cur in the ternary complex of control example 4LF-LMP-Cur

[0058] Dissolve Cur in anhydrous ethanol to prepare a 10.8 mmol / L Cur stock solution and store at 4°C in the dark until needed. Mix equal volumes of the prepared LF and LMP stock solutions and magnetically stir for 2 hours to form an LF-LMP binary complex. Mix 9.5 mL of the LF-LMP binary complex with 0.5 mL of the Cur stock solution and stir at 25°C for 2 hours to form a LF-LMP-Cur ternary complex.

[0059] The retention rate of Cur in the LF-LMP-Cur ternary complex is as follows Figure 5 As shown in the figure. After 24 hours of UV irradiation, the retention rate of Cur in the LF-LMP-Cur ternary complex was 37.4%, corresponding to a half-life of 16.4±9.3 hours, which is significantly lower than the stability of Cur in the (STE-Cur)-(LF-LMP) quaternary complex. This is because the ketone and bisphenol groups in the Cur structure are extremely sensitive to light and easily cause Cur photodegradation. The poor amphiphilicity of LF-LMP prevents efficient encapsulation of Cur, thus failing to effectively improve the photostability of Cur.

[0060] Example 5 Thermal Stability of Cur in the (STE-Cur)-(LF-LMP) Quaternary Complex

[0061] The (STE-Cur)-(LF-LMP) quaternary complex from Example 1 was placed in an 80°C water bath for 20, 40, 60, 90, and 120 minutes. The samples were then immediately removed and placed in an ice bath. The retention of Cur at different heating times was measured, and its half-life was calculated.

[0062] The retention rate of Cur in the (STE-Cur)-(LF-LMP) quaternary complex under different heating times is as follows Figure 6 As shown in the figure. After heat treatment at 80°C for 120 minutes, the retention rate of Cur in the (STE-Cur)-(LF-LMP) quaternary complex was 82.25%, corresponding to a half-life of 387.5±58.5 minutes. In contrast, the retention rate of Cur in a Cur solution alone (5% EtOH) was close to 0, indicating that Cur had been completely degraded at this point. Clearly, the (STE-Cur)-(LF-LMP) quaternary complex effectively improved the thermal stability of Cur. This is because the LF-LMP binary complex in the (STE-Cur)-(LF-LMP) quaternary complex facilitates Cur's entry into the deep hydrophobic cavity of STE. Cur is distributed in an amorphous state within the quaternary complex, and LF-LMP forms a tighter core-shell structure with STE-Cur through thermal aggregation, thereby slowing the thermal degradation of Cur and improving its thermal stability.

[0063] Control Example 5 Thermal stability of Cur in the STE-Cur binary complex

[0064] The STE-Cur binary complex, at the same concentration as in Example 5, was placed in an 80°C water bath for 20, 40, 60, 90, and 120 minutes. The samples were then immediately removed and placed in an ice bath. The retention of Cur at different heating times was measured, and its half-life was calculated.

[0065] After heat treatment at 80°C for 120 min, the retention rate of Cur in the STE-Cur binary complex was 43.37%, corresponding to a half-life of 85.6±15.9 min ( Figure 6 The stability of Cur in the STE-Cur binary complex is significantly lower than that of the (STE-Cur)-(LF-LMP) quaternary complex. This is because the spatial conformation of the STE-Cur binary complex is relatively simple and lacks the protection of the polyelectrolyte complex LF-LMP, resulting in poor thermal stability of Cur in STE-Cur.

[0066] Control Example 6 Thermal stability of Cur in the ternary complex of LF-LMP-Cur

[0067] The LF-LMP-Cur ternary complex, at the same concentration as in Example 5, was placed in an 80°C water bath for 20, 40, 60, 90, and 120 minutes. The samples were then immediately removed and placed in an ice bath. The retention of Cur at different heating times was measured, and its half-life was calculated.

[0068] After heat treatment at 80°C for 120 min, the retention rate of Cur in the LF-LMP-Cur ternary complex was 55.91%, corresponding to a half-life of 126.3 ± 28.1 min ( Figure 6 The stability of Cur in the LF-LMP-Cur ternary complex is significantly lower than that in the (STE-Cur)-(LF-LMP) quaternary complex. This is because the LF-LMP complex can encapsulate Cur inside the aggregates, while the LF-LMP-Cur ternary complex has a loose structure. The lack of STE interaction prevents the LF-LMP-Cur ternary system from forming an effective core-shell structure, resulting in the lower thermal stability of Cur in the LF-LMP-Cur ternary complex than that in the (STE-Cur)-(LF-LMP) quaternary complex.

[0069] Example 6 Thermal stability of Cur in high concentration (STE-Cur)-(LF-LMP) quaternary complex

[0070] By varying the concentrations of LF, LMP, and STE in Example 5, high-concentration (STE-Cur)-(LF-LMP) quaternary complexes were prepared and their thermal stability toward Cur was analyzed. Specifically, 8%, 20%, and 11.1% (w / v) stock solutions of LF, LMP, and STE were prepared in 10 mM acetate buffer (pH 5.0) and stored at 4°C overnight. 4.5 mL of the LF, LMP, and STE stock solutions were mixed with 0.5 mL of the Cur stock solution and stirred at 25°C for 2 hours to prepare the STE-Cur stock solution. 2.5 mL of the LF stock solution was mixed with 2.5 mL of the LMP stock solution and stirred at 25°C for 2 hours to prepare the LF-LMP stock solution. The STE-Cur stock solution was slowly poured into the LF-LMP stock solution and stirred thoroughly for 2 hours to form the (STE-Cur)-(LF-LMP) complex. The final concentrations of LF, LMP, and STE were 2%, 5%, and 5% (w / v), respectively, and the concentration of ethanol was 5% (v / v).

[0071] The resulting high-concentration (STE-Cur)-(LF-LMP) quaternary complex was placed in an 80°C water bath for 20, 40, 60, 90, and 120 minutes. The samples were then immediately removed and placed in an ice bath. The retention of Cur at different heating times was measured, and its half-life was calculated.

[0072] The retention rate of Cur in the high concentration (STE-Cur)-(LF-LMP) quaternary complex under different heating times is as follows Figure 7 As shown. After heat treatment at 80°C for 120 minutes, the retention rate of Cur in the high-concentration (STE-Cur)-(LF-LMP) quaternary complex was 88.63%, corresponding to a half-life of 513.5±40.4 minutes, significantly higher than the 59.87% retention rate of Cur in the LF-LMP-Cur ternary complex and the 48.71% retention rate of Cur in the STE-Cur binary complex. This result is similar to that in Example 5, indicating that LF-LMP forms a more compact core-shell structure with STE-Cur through thermal aggregation, thereby slowing the thermal degradation of Cur and improving its thermal stability. Furthermore, this result also demonstrates that as the final concentrations of LF, LMP, and STE in the (STE-Cur)-(LF-LMP) quaternary complex system increase to 2%, 5%, and 5% (w / v), respectively, Cur maintains excellent thermal stability in the high-concentration quaternary complex.

[0073] Example 7 Storage Stability of (STE-Cur)-(LF-LMP) Quaternary Complex

[0074] The (STE-Cur)-(LF-LMP) quaternary complex in Example 1 was stored in the dark at 25° C. for 30 days, and the particle size and PDI value of the sample were measured every 5 days.

[0075] The physical stability of the (STE-Cur)-(LF-LMP) quaternary complex was evaluated by measuring the changes in the average particle size and PDI value. The results of particle size and PDI values ​​were shown in Table 1. Figure 8 After 30 days, the average particle size and PDI value of (STE-Cur)-(LF-LMP) did not change significantly, while the average particle size and PDI value of the STE-Cur binary composite changed significantly, indicating that the quaternary composite (STE-Cur)-(LF-LMP) has good physical stability.

[0076] Although the present invention has been disclosed above in terms of preferred embodiments, it is not intended to limit the present invention. Anyone familiar with this technology can make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention should be based on the definition of the claims.

Claims

1. A (STE-Cur)-(LF-LMP) quaternary complex for improving the stability of curcumin (Cur), characterized in that: The quaternary complex uses LF, LMP, STE and Cur as raw materials, a STE-Cur binary complex as a core and a LF-LMP binary complex as a shell to form a core-shell structure, and self-assembles to form a quaternary complex (STE-Cur)-(LF-LMP).

2. The (STE-Cur)-(LF-LMP) quaternary complex for improving the stability of curcumin according to claim 1, characterized in that: The method for preparing the (STE−Cur)−(LF−LMP) quaternary complex comprises the following steps: (1) Prepare a STE stock solution in 10 mM pH 5.0 acetate buffer at 25°C. Dissolve Cur in anhydrous ethanol to prepare a Cur stock solution with a concentration of 10-200 mM. 0.5 mL of Cur mother solution was added dropwise to 4.5 mL of STE mother solution and stirred at 25 °C for 2-24 h to prepare the STE−Cur complex; (2) Prepare the LF-LMP polyelectrolyte complex by preparing the LF and LMP stock solutions separately in 10 mM pH 5.0 acetate buffer at 25°C. Mix the LF and LMP stock solutions in equal volumes and stir at 25°C for 2-24 h. (3) Slowly add 5 mL of STE−Cur mother solution into 5 mL of LF−LMP mother solution and stir thoroughly at 25 °C for 2–24 h to obtain the (STE−Cur)−(LF−LMP) quaternary complex.

3. A (STE-Cur)-(LF-LMP) quaternary complex for improving the stability of curcumin according to any one of claims 1 or 2, characterized in that: The degree of esterification of the LMP is 30%-45%.

4. A (STE-Cur)-(LF-LMP) quaternary complex for improving the stability of curcumin according to any one of claims 1 or 2, characterized in that: The final concentration of STE in the (STE−Cur)−(LF−LMP) complex system is 0.5%-5% w / v.

5. A (STE-Cur)-(LF-LMP) quaternary complex for improving the stability of curcumin according to any one of claims 1 or 2, characterized in that: The final concentration of ethanol in the (STE−Cur)−(LF−LMP) complex system is 1%-5% v / v, and the final concentration of Cur is 0.5-10 mM.

6. A (STE-Cur)-(LF-LMP) quaternary complex for improving the stability of curcumin according to any one of claims 1 or 2, characterized in that: The final concentration of LF in the (STE−Cur)−(LF−LMP) complex system is 0.1%-2% w / v, and the final concentration of LMP is 0.5%-5% w / v.

7. A (STE-Cur)-(LF-LMP) quaternary complex for improving the stability of curcumin according to any one of claims 1 or 2, characterized in that: The stirring frequency of the stirrer was 50-300 r / min, and the temperature was kept constant at 25°C.

8. Use of the (STE-Cur)-(LF-LMP) quaternary complex prepared according to any one of claims 2 to 7 for improving the stability of curcumin in the preparation of foods, beverages, and medicines.

Images