Preparation and in vitro and in vivo evaluation of quercetin-loaded mixed micelles for oral delivery

Abstract

Quercetin (QT) is a plant polyphenol with various pharmacological properties. However, the low water solubility limits its therapeutic efficacy. In the present study, QT-loaded sodium taurocholate-Pluronic P123 (QT-loaded ST/P123) mixed micelles were developed and characterized, and the effect of the formulation on improving the water solubility of QT was investigated. QT-loaded ST/P123 mixed micelles were prepared by thin film hydration-direct dissolution and optimized by uniform design. The optimal formulation possessed high drug loading (12.6%) and entrapment efficiency (95.9%) in small (16.20 nm) spherically-shaped micelles. A low critical micelle concentration indicated that the micelles were stable, and they showed a sustained release pattern, as determined in vitro in simulated gastric fluid and intestinal fluid. Pharmacokinetic evaluation showed the C_max and AUC_0–24 were 1.8-fold and 1.6-fold higher than the QT suspension. The present results indicate that QT-loaded ST/P123 micelles are potential candidates to improve the solubility and oral bioavailability of QT.

Schematic illustration of QT-loaded mixed micelles. ST, P123 and QT were dissolved in EtOH, then evaporated to obtain thin film, followed by hydration to form a uniform system.

Keywords:

• Mixed micelles
• quercetin
• sodium taurocholate
• Pluronic P123

Oral administration remains the preferred drug delivery route because it is convenient and improves patient compliance. In the gastrointestinal (GI) tract, however, gastric acid, bile, and various digestive enzymes cause drug degradation, leading to decreased absorption efficacy. Therefore, new strategies are needed to facilitate drug transport from the GI tract to the blood circulation and improve oral bioavailability [1–3].

Quercetin (QT, 3,3′,4′,5,7-pentahydroxy flavone) is a potent polyphenolic flavonoid antioxidant with various pharmacological properties, including anti-inflammatory [4,5], antioxidative [6,7], antiviral [8,9], and anticancer effects [10]. However, the clinical use of QT is limited by its poor water-solubility, GI instability, minimal oral bioavailability (<17% in rats and <2% in humans) [11], and extensive first pass metabolism before reaching the systemic circulation [12]. To overcome these shortcomings, recent studies have relied on nanotechnology [13] to develop various QT-loaded nanocarriers such as nanoparticles [14–18], polymeric micelles [19–21], conjugates [22], inclusion complexes [23], and nanosuspensions [24]. Mixed micelles with nanosized distribution have several favorable properties, such as increased micelle stability, solubility, and bioactivity. A simple preparation process and low toxicity in the absence of organic solvents make mixed micelles potential candidates for QT delivery, especially for oral administration [25,26]. For instance, lecithin-based mixed polymeric micelles improved the solubility of QT loaded in nanosystems to 5.81 mg/mL compared with 0.17–7.7 μg/mL for free QT in water [12].

Pluronic P123 (P123), composed of PEO₂₀-PPO₆₈-PEO₂₀, is an amphiphilic synthetic polymer containing hydrophilic poly(ethylene oxide) (PEO) block as an outer shell and hydrophobic poly(propylene oxide) (PPO) block as an inner core [27,28]. The PPO core can serve as a “pool” for the encapsulation of hydrophobic drugs in the lipophilic microenvironment, whereas the hydrophilic PEO corona maintains the dispersion stability of micelles [29]. Bile salt is considered as an important biosurfactant with strong solubilization ability and good biocompatibility [30], and mixed micelles containing bile salt increase the solubility and improve the oral bioavailability of hydrophobic drugs [31–33].

In the present work, QT loaded sodium taurocholate/Pluronic P123 (QT-loaded ST/P123) mixed micelles were initially prepared to improve the solubility of QT in water, increase oral absorption, and protect the drug during passage through the GI tract. The mixed micelles were prepared by the thin-film hydration-direct dissolution method and the preparation was optimized by uniform design. The physicochemical properties were characterized and the drug release pattern was evaluated in simulated gastric fluid and intestinal fluid. The pharmacokinetics of QT-loaded ST/P123 micelles were investigated in rats.

Materials and methods

Materials and animals

QT, Pluronic P123 (P123), and sodium taurocholate (ST) were purchased from Sigma-Aldrich (USA). Tween 80 was provided by Tianjin Kermel Chemical Ltd (China). All other chemicals used were of analytical grade.

Male Wistar rats (weight: 200 ± 20 g) were supplied by the Laboratory Animal Center of Shandong University (China). The rats were kept in cages for 2 weeks before experiments and fed with food and water ad libitum. Before the experiments, the rats were fasted for 12 h with free access to water. The procedures complied with the requirements of the Institute’s Animal Care and Use Committee of Shandong University.

Preparation of QT-loaded ST/ P123 mixed micelles

QT-loaded ST/P123 mixed micelles were prepared by the thin-film hydration-direct dissolution method [34,35]. Briefly, QT, ST, and P123 (the total amount of ST and P123 was 200 mg) were dissolved in ethanol and the solvent was removed under reduced pressure to acquire a thin polymeric film in a 50-ml round-bottom flask. The thin film was dried under vacuum overnight, and the resulting dry polymeric film was hydrated with 4 mL of deionized water, and then vortexed for 5 min. Finally, the mixture was centrifuged at 12,000 rpm for 5 min to remove unloaded drugs, resulting in a clear supernatant as the micelle solution.

Optimization of the QT-loaded ST/P123 mixed micelle formulation

The uniform design method was used to determine the optimal formulation of QT-loaded ST/P123 micelles. The complete design consisted of 9 (3²) experimental points. Before using the design, a great number of single-factor experiments were preliminarily conducted using drug loading coefficient (DL) and encapsulation efficiency (EE) as evaluation indexes.

Characterization of QT-loaded ST/P123 mixed micelles

Particle size distribution and zeta potential

The size distribution and zeta potential of QT-loaded ST/P123 micelles were measured using a dynamic light scattering instrument (DLS, DELSA^TM NANO particle size, and zeta potential analyzer, Beckman Coulter Inc., USA) at 25 °C. Measurements were processed in triplicate for each sample.

Surface morphology

The surface morphology of micelles was observed by transmission electron microscopy (TEM, JEM-100SX, JEOL, Japan) after negative staining with a phosphotungstic acid solution (1.5%, w/v).

The thermal characteristics of the QT, blank micelles, physical mixture, and QT-loaded ST/P123 mixed micelles were determined using a TGA/DSC1 instrument (Mettler Toledo, Switzerland) under a 10 mL/min stream of nitrogen purge. All samples were heated from 30 °C to 300 °C at the rate of 10 °C/min in an open aluminum pan, and TA4 Analysis software was used for analysis.

Differential scanning calorimetry (DSC)

Thermograms of QT, blank micelles, a physical mixture of QT and blank micelles, and QT-loaded micelles were recorded on a differential scanning calorimeter (Mettler Toledo). Weighed samples of 10 mg were placed in aluminum pans and heated at a rate of 10 °C from 25 °C to 400 °C in a nitrogen atmosphere. Aluminum oxide was used as the internal standard.

Drug loading and entrapment efficiency

To determine the DL content and EE of micelles, QT-loaded ST/P123 mixed micelles were dissolved in methanol to break up the nanostructures. The concentration of QT was then detected by an ultraviolet-visible spectrophotometer (UV-2102, Shanghai Instrument Ltd., China) at 376 nm. The DL and EE were calculated using the following equations:[36]:

Critical micelle concentration determination

Critical micelle concentration (CMC) of the mixed micelles was determined by fluorescence spectroscopy using pyrene as the fluorescence probe [37]. When the concentration of polymers increased up to the CMC in an aqueous environment, lipophilic pyrene tended to accumulate in the inside core of micelles, resulting in an alteration in the intensity ratio of I374/I384. The fluorescence spectrum was recorded on a fluorescence spectrophotometer at room temperature. In brief, pyrene was dissolved in acetone, before being diluted to a final concentration of 2.0 × 10⁻⁶ M. A total of 100 μL of pyrene solution was extracted into a colorimetric tube and the solvent was evaporated under nitrogen flow. After that, a series of polymer solutions with different concentrations (ranging from 1 × 10⁻³ M to 3 M) was added into the colorimetric tubes and vortexed for 5 min. After storage in the dark for 24 h, the fluorescence spectra of samples were recorded at 350–500 nm. The excitation wavelength was set at 334 nm, and the excitation and emission slit widths were set at 5 and 2.5 nm, respectively.

In vitro release studies

Dialysis was used to investigate the in vitro release pattern of QT from QT- loaded ST/P123 mixed micelles using a dialysis bag (MWCO 3500 Da, Solarbio, China). QT-loaded ST/P123 mixed micelles (200 μL) were introduced into the dialysis membrane bag, and the end-sealed dialysis bag was then immersed in 50 mL release medium at 37 ± 0.5 °C and shaken at a speed of 100 rpm. After 2 h of incubation in simulated gastric fluid (SGF, pH = 1.2), the bag was transferred into simulated intestinal fluid (SIF, pH = 6.8) for the next 46 h. At predetermined time intervals, 2 mL of the release media was withdrawn and replaced with an equal volume of the fresh release medium. The concentration of QT was determined by the UV method with 375 nm as the determination wavelength, and the cumulative release of QT was calculated according to the formula described below. The release pattern of QT from QT propylene glycol solution was assessed under the same conditions as the control group. All samples were performed in triplicate.

where E_r is DTX accumulative release, %; V_e stands for displacement volume of release medium, 2 mL; V₀ represents the volume of release medium, 80 mL; C_i is the release solution concentration at the displaced time, mg/mL; m_drug stands for QT amount of QT-loaded mixed micelles (mg); and n is the number of times of release medium displacement.

Pharmacokinetic studies

Wistar rats were randomly divided into two groups, and intragastrically administered QT-loaded ST/P123 mixed micelles and QT suspension with the same concentration. QT suspension was prepared by dispersing 40 mg QT in 10 mL of 0.4% carboxymethylcellulose sodium solution. The dose of QT in all samples was maintained at 80 mg/kg. At each time point (0.25, 0.5, 1, 2, 4, 6, 8, 10, 12, and 24 h), 300 μL of blood was collected from the sinus jugularis of rats and placed into heparinized tubes. Samples were then centrifuged immediately (4000 rpm for 10 min) and the separated plasma was stored at –20 °C until analysis. Rat plasma (200 μL) was added to acetonitrile-acetic acid (400 μL, 9:1, v/v), then vortexed for 90 s, hydrolyzed in a 50 °C water bath for 15 min, and then centrifuged (10,000 rpm for 10 min). The supernatant (20 μL) was analyzed by HPLC. The pharmacokinetic parameters were calculated using DAS (version 2.0).

HPLC analysis of QT

The samples were analyzed using an HPLC system (1200 series; Agilent Technologies, USA) and an HPLC column (C18, 4.6 × 250 mm, 5 μm). The mobile phase was a mixture of acetonitrile-water at a ratio of 33: 67 (v:v) containing 0.75% acetic acid. The flow rate was 1 mL/min, and the detection wavelength was 376 nm. Aliquots of 20 μL of each sample were injected into the column, and all operations were performed at ambient temperature.

Statistical analysis

Data expressed as the mean ± standard deviation (SD) are representative of at least three different experiments. Statistical data were analyzed by the Student’s t-test at the level of p < 0.05.

Results

Optimization of QT-loaded P123 /ST mixed micelles

Single-factor experiment

Single-factor experiments were performed to obtain optimal formulations. The EE and DL were selected as indexes to optimize the formulation of QT-loaded P123/ST mixed micelles. The results of preliminary experiments showed that carriers ratio (P123:ST), hydration temperature, and the added amount of QT affected the DL and EE. Therefore, single-factor experiments were performed to determine the appropriate ranges of these three factors. Table 1 shows the EE and DL values according to P123:ST ratio, hydration temperature, and QT amount. The highest DL and EE values were obtained at a P123:ST ratio of 1:1, a temperature of 40 °C, and with 30 mg of added QT.

Table 1. Single-factor experiments with investigated factors and observed results at different levels.

Single-factors	Levels	EE (%)	DL (%)
P123/ST (mg/mg)	50:150	21.844	3.682
	70:130	23.291	3.916
	100:100	75.949	11.732
	130:70	50.488	8.118
	150:50	51.356	8.246
Hydration temperature	20 °C	70.944	11.044
	30 °C	77.656	11.964
	40 °C	79.74	12.246
	50 °C	65.967	10.35
Added amount of QT	20 mg	98.253	9.112
	25 mg	90.896	10.203
	30 mg	94.684	12.436
	35 mg	75.349	11.65

Optimization of QT-loaded P123/ST micelles based on uniform design

The independent variables affecting the DL and EE in the uniform design are shown in Table 2. The multiple regression method was used to fit the data to regression equations. The quadratic polynomial equation generated by the design was as follows:

Table 2. Results of uniform design experiments.

No.	X1 (P123:ST)	X2 (added amount of QT)	S1 (DL%)	S2 (EE%)	S ()
1	0.667	20	8.3	90.1	27.346
2	0.667	25	10.3	91.6	30.716
3	0.667	25	8.9	78.4	26.415
4	1	30	12.8	98.3	35.472
5	1	20	8.4	91.9	27.784
6	1	20	8.8	96.5	29.141
7	1.5	25	10.2	91.8	30.600
8	1.5	30	12.6	95.9	34.761
9	1.5	30	12.2	92.8	33.648

where S stands for , X1 represents the ratio of P123 to ST, and X2 represents the amount of added QT.

From the formula, all the dependent variables can be described by the quadratic polynomial with a high correlation coefficient (r > 0.95), and response surface plots (Figure 1) were generated according to the regression equations. These results indicated that the added amount of QT played a crucial role in DL and EE. Increasing the QT in formulations increased the amount of QT likely to be encapsulated in micelles, resulting in increased DL and EE. However, the EE decreased when the QT amount exceeded the entrapment capability of the micelles. Considering all the factors, the optimal formulation was set as follows: P123:ST ratio of 120:80 mg, QT of 30 mg, hydration temperature of 40 °C, and hydration volume of 4 mL.

Figure 1. Response surface plot showing the effects of the P123:ST ratio (X1) and the added amount of QT (X2) on drug loading efficiency (A), and the contours of X1 and X2 (B).

Characterization of mixed micelles

TEM is a practical technique to visualize the morphology of micelles. As shown in Figure 2(A), the mixed micelles generated homogeneous nanometric spheres with an obvious core-shell structure. In addition, the micelles showed good formability and uniform dispersion without aggregation. The outer shell of the micelles was formed by the ST and PEO in P123, and the inner core consisted of hydrophobic PPO and QT.

Figure 2. Characterization of mixed micelles. Transmission electron microscope images of QT-loaded mixed micelles (A); size distribution of micelles without QT (B) and QT-loaded mixed micelles (C); the zeta potential distributions of unloaded mixed micelles (D) and QT-loaded mixed micelles (E); external photos of QT-loaded mixed micelles in water (F-i) and QT in water at the same concentration (F-ii).

The mean size of the QT-loaded mixed micelles was 16.20 ± 2.12 nm, with a polydispersity index (PDI) of 0.123 ± 0.071, and that of the micelles without QT was 17.67 ± 1.97 nm (PDI: 0.213 ± 0.054) (Figure 2(B) and (C)). Analysis of the zeta potential of blank ST/P123 and QT-loaded ST/P123 micelles showed that the surface was negatively charged, with zeta-potential values of −8.65 ± 0.972 mV and −10.0 ± 1.192 mV, respectively, indicating the good stability of micelles (Figure 2(D) and (E)). The solubility of QT in the optical QT-loaded micelle solution determined using the UV method was 7.08 mg/mL, which was higher than that of free QT in water (0.17–7.7 μg/mL). With respect to the surface morphology, QT-loaded mixed micelles aqueous solution was clear and transparent, while the QT dispersing in water at equal mass appeared turbid, non-transparent and insoluble state (Figure 2(F)).

Figure 3 shows the thermograms of QT, blank micelles, the physical mixture of QT and blank micelles, and QT-loaded micelles. QT (a) exhibited a sharp endothermic peak at 315 °C, which could be attributed to its crystalline nature, as reported previously [38]. The blank micelles (c) showed a broad endothermic peak at 90 °C, whereas in the physical mixture with QT (b), a smaller peak at 320 °C was observed for QT, indicating the presence of remaining crystalline drugs. The QT-loaded micelles showed no endothermic peak around the melting point of crystal QT (d), suggesting that the QT in micelles was in a noncrystalline state.

Figure 3. DSC curves of QT (a), a physical mixture of QT and blank micelles (b), blank micelles (c), and QT-loaded micelles (d).

CMC determination

When the concentration of polymers increases above the CMC, P123, and ST can self-assemble into aggregates in an aqueous environment. Therefore, the CMC value is an important parameter and micelles with a low CMC show high stability and anti-dilution properties [39]. As a hydrophobic molecule, pyrene was used as a probe because it preferentially enters the hydrophobic microenvironment of aggregates, and its solubility significantly increases when the concentration of polymers increases above the CMC. The CMC values of ST/P123 mixed aggregates were obtained by plotting the I374:I384 ratio of the emission spectrum profile against the concentration of aggregates, as shown in Figure 4. The inflection point of the ST/P123 mixed micelles was 3.25 × 10⁻³ g/L.

Figure 4. CMC value of ST/P123 mixed micelles according to the I1:I3 ratio of the emission spectra profile.

Drug release profile in simulated GI tract

The in vitro profile of the release of QT from ST/P123 micelles was determined in a simulated GI tract including SGF (pH 1.2) and SIF (pH 6.8). As shown in Figure 5, 29.20% of the QT was released from QT-loaded ST/P123 micelles in SGF within 2 h and then 78.56% in SIF within 24 h. However, 37.96% of the QT was released from propylene glycol in SGF within 2 h and then 99.64% in SIF within 11 h. Therefore, the QT-loaded mixed micelles showed a controlled and sustained release pattern.

Figure 5. Cumulative released amount of QT as a function of time in SGF/SIF. Values are presented as the mean ± SD (n = 3).

In vivo pharmacokinetics

A part of QT was metabolized into glycuronate or sulfated compounds after entering blood through the oral absorption. The hydrolysis process could revert these glycuronate or sulfated compounds back to the QT form, and then all the QT would be analyzed with the method of HPLC [40]. The total concentration of QT that was absorbed into blood from the QT-loaded ST/P123 mixed micelles and QT suspension during a 24 h period was determined. Pharmacokinetic analysis showed that the total QT plasma concentration was significantly higher for the QT-loaded ST/P123 mixed micelles than for the QT suspension (p < 0.05), as determined by plasma concentration-time curves (Figure 6). In accordance with the result of in vitro drug release, QT was detectable in plasma within 24 h, indicating that QT was released from QT-loaded ST/P123 mixed micelles in a sustained manner over a long period of time. The pharmacokinetic parameters are listed in Table 3. For QT-loaded ST/P123 micelles, the peak concentration (C_max) of the drug in mixed micelles (5.13 mg/L) was 1.8-fold higher than that of the QT suspension (2.82 mg/L). The area under the concentration-time curve (AUC_0–24), a key drug therapeutic index, was 80.42 mg/L × h for QT-loaded ST/P123 mixed micelles, which was 1.6-fold higher than that of the QT suspension at 50.38 mg/L × h (Table 3).

Figure 6. The plasma concentration-time curves of QT suspension and QT-loaded mixed micelles. Values are presented as the mean ± SD (n = 5).^*p < 0.05 vs. QT solution.

Table 3. Pharmacokinetic parameters after oral administration (values are presented as the mean ± SD, n = 5).

	QT suspension	QT-loaded ST/P123
Dose (mg/kg)	80	80
Tmax (h)	6	8
Cmax (mg/L)	2.82 ± 0.51	5.13 ± 0.53
AUC(0–24) (h.mg/L)	50.38 ± 5.26	80.42 ± 1.34
MRT(0–24) (h)	11.40 ± 0.24	10.56 ± 0.28

Discussion

Drug delivery systems based on polymeric micelles have gained attention because of their potential to increase drug absorption by protecting drugs from the GI tract environment, providing controlled drug release at target sites, increasing the time of residence of a drug in the gut, and improving the bioavailability and delivery of poorly water-soluble drugs [41]. Amphiphilic polymers that assemble into nanocarriers with a hydrophobic core and a hydrophilic shell have been developed to improve the delivery of poorly soluble drugs [42,43]. The encapsulation of hydrophobic drugs in the core protects them from degradation, increasing bioavailability, and the small size of micelles increases circulation time and favors biodistribution [44,45]. Here, QT-loaded ST/P123 mixed micelles were developed, characterized, and optimized, and shown to improve the solubility of QT, increase oral absorption, and protect the drug during passage through the GI tract.

For oral administration, the particle size and surface properties of nanocarriers could affect the absorption rate in the GI tract [19]. QT-loaded mixed micelles showed a slight decrease in the average size comparing with the micelles without QT, which could be attributed to the formation of a compact hydrophobic core due to the addition of QT and increased hydrophobic interaction between drugs and carriers. A small size prolongs the retention time of micelles in intestinal villi and increases the internalization ability via the intestinal epithelium [46,47]. The nanoparticles with sizes of 10–100 nm can avoid being filtered out by the kidneys or absorbed and metabolized by the liver [48]. Therefore, QT-loaded mixed micelles with a particle size of 16.2 nm can avoid rapid metabolism and elimination, ensure stability and long circulation time, and improve targeting abilities by increasing permeability and retention.

The zeta potential has an important effect on the storage stability of colloid dispersion systems, and particle aggregation is likely to occur if the zeta potential of particles is too low to provide sufficient electric repulsion or steric barriers [49]. The zeta potential of QT-loaded ST/P123 micelles was slightly higher than that of blank micelles. This change in zeta potential could be attributed to the dissociation of acidic phenolic OH groups on the molecule of QT [50]. Chitkara et al. prepared QT-loaded poly(lactic-co-glycolic acid) nanoparticles by the emulsion diffusion evaporation method and obtained a zeta potential of −6.06 mV [11]. Xu et al. used a self-assembly method to prepare QT-loaded MPEG-PCL nanomicelles. The zeta potential of the nanomicelles was only −1.56 mV [51]. The higher the zeta potential value, the higher the electrostatic repulsion, which results in a greater stability of particles. Compared with the above results, the QT-loaded ST/P123 micelles with a zeta potential of −10.0 mV could have better stability.

In the present study, ST and P123 mixed micelles were used to increase the water solubility of QT. The capacity of the polymer micelles to increase the aqueous solubility of hydrophobic molecules is related to their hydrophobic core, which provides a suitable microenvironment for a hydrophobic solute [52]. The effects of mixed polymeric micelles prepared using P123 on improving the solubility of QT were reported previously [53]. The presence of ST in micelle preparations increased the solubilities of poorly soluble drugs, which was consistent with previous studies [48].

The physicochemical status of QT in the present formulation was assessed by DSC. The results of DSC demonstrated that the phase state of QT in mixed micelles had changed and the drug was highly dispersed in the micelles system. The change in the structure of QT in mixed micelles suggested that QT in the micelles was in a noncrystalline state, such as an amorphous or disorder crystalline phase of molecular dispersion [28], which was favorable to improve the solubility of poorly water-soluble drugs.

The delivery of drugs by the oral route requires resistance against degradation in the acidic environment of the GI tract and in the presence of digestive enzymes and bile salts. CMC is the threshold at which polymer molecules start to assemble into polymeric micelles, and a low CMC value denotes a greater stability [54]. In the present study, low CMC values indicated that the mixed micelles composed of ST/P123 maintained their integrity even upon extreme dilution in the GI tract.

QT-loaded mixed micelles showed a controlled and sustained release pattern. The slower release of QT from QT-loaded ST/P123 micelles compared with that of QT propylene glycol in SGF/SIF could be attributed to the encapsulation of QT in the core of micelles. With the uptake of water, micelles are likely to swell, leading to drug diffusion and the erosion of polymers [55]. However, the diffusion of water into micelles could be delayed because of the hydrophobic core structure, which contributed to their sustained release. This property contributed to preventing rapid leakage and precipitation within the GI lumen during drug delivery via oral administration.

The in vivo pharmacokinetic study was performed to investigate the oral bioavailability of QT. As shown in Figure 6, the total QT plasma concentration of the QT-loaded ST/P123 mixed micelles was significantly higher than that of the QT suspension, which could have a positive effect on the therapeutic index. The C_max of the free QT in mixed micelles was higher than that of the QT suspension, indicating that a greater amount of QT was absorbed in the GI tract from QT-loaded micelles. The results might be attributed to the in vivo behaviors difference between QT suspension and QT-loaded mixed micelles. For QT suspension, it was difficult for hydrophobic QT to be absorbed by mucus, leading to the reduced accumulation in the blood circulation and a low C_max. However, for QT-loaded mixed micelles, the micelles increased the drug solubility and stability in the GI tract and the small particle size enabled the whole micelles to penetrate across the pre-epithelial medium mucus and then be uptaken into enterocyte [54]. In addition, free QT could enter the enterocyte with the help of various surfactants after the diffusion of the micelles, and many drugs were absorbed into the enterocyte by passive diffusion. Meanwhile, the component P123 in mixed micelles could inhibit P-gp transporter in the enterocyte to prevent the efflux of QT [56,57]. Therefore, the ST/P123 mixed micelles dramatically increased the total concentration of QT in plasma and significantly improved the oral bioavailability of QT.

In this study, QT-loaded mixed micelles composed of Pluronic P123 and ST copolymers for oral administration were prepared and the formulation was optimized by uniform design. The prepared QT-loaded micelles showed high stability and a controlled and sustained release pattern. The results indicate that QT-loaded ST/P123 mixed micelles could be a potential nanoformulation to enhance the solubility and oral bioavailability of QT.

Author contributions

Conceived and designed the experiments: Guangxi Zhai, Zhen Lu, Cuiping Bu. Performed the experiments: Hui Zhang, Mengrui Liu, Meiqi Lu. Contributed reagents/materials/ analysis tools: Zhen Lu, Cuiping Bu, Weicheng Hu. Wrote the paper: Guangxi Zhai, Zhen Lu, Cuiping Bu, Weicheng Hu. All the authors reviewed the results and approved the final version of the manuscript.

Disclosure statement

No potential conflict of interest was reported by the authors.

Funding

This work was supported by the National Natural Science Foundation of China [grant number 31300463]; the Natural Science Foundation of Jiangsu Province [grant number BK20130414]; Qing Lan Project of Jiangsu Province; and the Science Research Program of Taian [grant number 201440774-30B].

Acknowledgments

The authors gratefully acknowledge the help of Xingzhen Zhang, who provided technical assistance with HPLC.