Comparison of bile salt/phosphatidylcholine mixed micelles in solubilization to sterols and stability

Figures & data

Figure 1 Molecular structures of synthesized sterols.

Notes: (A) Triol; (B) YC-1; (C) YC-10; (D) YC-5.
Abbreviations: Triol, Androst-3β,5α,6β-triol; YC-1, Cholest-24-ene-3β,5α,6β,19-tetrol; YC-10, Androst-2β,3α,5α-trihydroxy-6-one; YC-5, Cholest-3β,5α,6β-triol

Table 1 Solubility of Triol in different BS/PC-MM systems with different weight ratios of BS to PC

BS/PC (w/w)	Solubility of Triol (mg/mL)
	SGC/EPC	SDC/EPC	SBA-Na/EPC	SGC/SPC	SDC/SPC	SBA-Na/SPC
1:1	7.4±0.6	9.0±0.5	11.5±0.8	7.0±0.3	7.4±0.1	10.1±0.3
1.5:1	9.3±0.5	10.9±0.8	18.6±0.7	9.0±0.1	9.9±0.1	15.2±0.1
2:1	10.8±0.3	11.3±1.0	21.5±1.5	11.0±0.9	10.0±0.5	22.9±0.8
3:1	14.0±1.1	19.3±1.5	24.7±1.2	13.2±1.4	23.2±0.2	24.8±0.9
4:1	15.4±1.3	26.4±0.9	32.4±1.8	15.7±0.5	24.6±0.7	32.5±1.3
6:1	24.0±0.9	28.0±1.9	32.8±1.1	22.7±0.7	25.3±0.5	35.5±0.3

Notes: Concentration of PC was fixed at 50 mg/mL; data presented as mean ± SD (n=3).

Abbreviations: BS, bile salt; BS/PC-MM, bile salt/phosphatidylcholine mixed micelles; EPC, egg phosphatidylcholine; PC, phosphatidylcholine; SBA-Na, swine bile acid-sodium salt; SD, standard deviation; SDC, sodium deoxycholate; SGC, sodium glycocholate; SPC, soya phosphatidylcholine; Triol, Androst-3β,5α,6β-triol.

Table 2 Solubility of Triol in different BS/PC-MM systems with different weight ratios of BS to PC at a fixed total concentration

BS/PC (w/w)	Solubility of Triol (mg/mL)
	SGC/EPC	SGC/SPC	SDC/EPC	SDC/SPC	SBA-Na/EPC	SBA-Na/SPC
1:1	11.9±0.3	14.7±0.1	13.9±0.2	12.9±0.4	16.3±0.8	14.5±0.6
1.5:1	10.5±0.2	14.2±0.5	14.9±0.5	14.7±0.3	16.5±0.2	17.4±0.5
2:1	14.8±0.3	15.3±0.3	17.0±0.4	17.3±0.2	19.4±0.6	19.6±0.9
3:1	13.6±0.4	13.2±0.4	17.3±0.1	17.5±0.6	16.8±0.1	15.9±0.2
4:1	10.7±0.1	12.9±0.3	13.2±0.2	13.8±0.2	14.9±0.3	14.4±0.3
6:1	10.4±0.1	12.5±0.1	9.3±0.2	13.2±0.4	14.5±0.3	13.1±0.3

Notes: Total concentration of BS and PC was fixed at 100 mg/mL; data presented as mean ± SD (n=3).

Abbreviations: BS, bile salt; BS/PC-MM, bile salt/phosphatidylcholine mixed micelles; EPC, egg phosphatidylcholine; PC, phosphatidylcholine; SGC, sodium glycocholate; SPC, soya phosphatidylcholine; SDC, sodium deoxycholate; SBA-Na, swine bile acid-sodium salt; SD, standard deviation; Triol, Androst-3β,5α,6β-triol.

Figure 2 Solubility of Triol in different BS/PC-MM systems with their optimized ratios and varied total concentrations of BS and PC (n=3).

Notes: (A) SGC/EPC-MM, (B) SGC/SPC-MM, (C) SBA-Na/EPC-MM, (D) SBA-Na/SPC-MM, (E) SDC/EPC-MM and (F) SDC/SPC-MM.
Abbreviations: BS, bile salt; BS/PC-MM, bile salt/phosphatidylcholine mixed micelles; EPC, egg phosphatidylcholine; MM, mixed micelles; PC, phosphatidylcholine; SGC, sodium glycocholate; SPC, soya phosphatidylcholine; SBA-Na, swine bile acid-sodium salt; SDC, sodium deoxycholate; Triol, Androst-3β,5α,6β-triol.

Table 3 Characterization of different Triol-loaded BS/PC-MM systems at their optimized ratios and total concentrations

Triol-loaded BS/PC-MM systems	Particle size (nm)	PDI	Zeta potential (mV)
SGC/EPC	11.0±0.7	0.171	−20.5±0.4
SGC/SPC	3.9±0.4	0.228	−20.0±0.3
SDC/EPC	7.6±1.1	0.262	−14.0±1.3
SDC/SPC	8.3±0.3	0.240	−14.9±0.6
SBA-Na/EPC	11.4±0.3	0.211	−16.3±1.0
SBA-Na/SPC	7.4±0.4	0.231	−15.8±1.1

Note: Data presented as mean ± SD (n=3).

Abbreviations: BS/PC-MM, bile salt/phosphatidylcholine mixed micelles; EPC, egg phosphatidylcholine; PDI, polydispersity index; SD, standard deviation; SBA-Na, swine bile acid-sodium salt; SDC, sodium deoxycholate; SGC, sodium glycocholate; SPC, soya phosphatidylcholine; Triol, Androst-3β,5α,6β-triol.

Table 4 Stability of varied Triol-loaded BS/PC-MM systems at their optimized ratios and total concentrations at 25°C±2°C

Triol-loaded BS/PC-MM systems	Appearance
	Month 0	Month 1	Month 3
SGC/EPC	−	−	−
SGC/SPC	−	−	−
SDC/EPC	−	−	+
SDC/SPC	−	−	+
SBA-Na/EPC	−	+	++
SBA-Na/SPC	−	+	++

Notes: “−” indicates transparent; “+” indicates precipitation; “++” indicates severe precipitation.

Abbreviations: BS/PC-MM, bile salt/phosphatidylcholine mixed micelles; EPC, egg phosphatidylcholine; SBA-Na, swine bile acid-sodium salt; SDC, sodium deoxycholate; SGC, sodium glycocholate; SPC, soya phosphatidylcholine; Triol, Androst-3β,5α,6β-triol.

Figure 3 The change of CMC of Triol-loaded BS/PC-MM systems under accelerated testing conditions.

Notes: (A) SGC/EPC-MM (■), SGC/SPC-MM (●); (B) SDC/EPC-MM (■), SDC/SPC-MM (●); (C) SBA-Na/EPC-MM (■), SBA-Na/SPC-MM (●); accelerated conditions: 40°C, 75% RH.
Abbreviations: CMC, critical micelle concentration; BS/PC-MM, bile salt/phosphatidylcholine mixed micelles; EPC, egg phosphatidylcholine; RH, relative humidity; SBA-Na, swine bile acid-sodium salt; SDC, sodium deoxycholate; SGC, sodium glycocholate; SPC, soya phosphatidylcholine; Triol, Androst-3β,5α,6β-triol.

Figure 4 Solubility of synthesized sterols in the optimized SGC/EPC-MM system with ratio in weight 2:1 and the total concentration of 100 mg/mL (n=3).

Abbreviations: EPC, egg phosphatidylcholine; MM, mixed micelles; SGC, sodium glycocholate; Triol, Androst-3β,5α,6β-triol; YC-1, Cholest-24-ene-3β,5α,6β,19-tetrol; YC-5, Cholest-3β,5α,6β-triol; YC-10, Androst-2β,3α,5α-trihydroxy-6-one.

Figure S1 Plot of the fluorescence of pyrene I₁/I₃ intensity ratio versus concentration of surfactant. The CMC corresponds to the center of the sigmoid.

Notes: The pyrene I₁/I₃ ratio varies in response to the position of pyrene, and CMC can be measured through the transition of the pyrene I₁/I₃ ratio. Above the CMC, the ratio reaches a roughly constant value because the pyrene is located in the polar environment of water. As the pyrene migrates to a more hydrophobic environment, ie, the micelles, the ratio experiences a rapid decrease and finally reaches a constant value when the pyrene is completely dispersed in the hydrophobic region of micelles. Boltzmann-type sigmoid is given by the equation in the figure, where the variable y corresponds to the pyrene I₁/I₃ ratio value, the independent variable x is the total concentration of the surfactant, A₁ and A₂ are the upper and lower limits of the sigmoid, respectively; x₀ is the center of the sigmoid and Δx is directly related to the independent variable occur rence. The CMC was determined from the mid-point of the sigmoid.
Abbreviations: CMC, critical micelle concentration; I, fluorescence intensity of pyrene; I₁/I₃, intensity of the third to the first peak of pyrene.

Figure S2 Representative plots of the fluorescence of pyrene I₁/I₃ intensity ratio versus concentration of BS/PC-MM systems under accelerated conditions.

Notes: (A) SGC/EPC-MM, (B) SDC/EPC-MM, (C) SBA-Na/EPC-MM. The pyrene I₁/I₃ ratio data are fitted by a Boltzmann-type sigmoid. The CMC can be obtained from the midpoint of the graph plotted by I₁/I₃ versus logarithm of mixed micelles concentration (intersection of the dashed line and X-axis).
Abbreviations: BS/PC-MM, bile salt/phosphatidylcholine mixed micelles; CMC, critical micelles concentration; EPC, egg phosphatidylcholine; I, fluorescence intensity of pyrene; I₁/I₃, intensity of the third to the first peak of pyrene; MM, mixed micelles; SBA-Na, swine bile acid-sodium salt; SDC, sodium deoxycholate; SGC, sodium glycocholate; w/v, weight/volume.