Suspensions of antibiotics in self-emulsifying oils as a novel approach to formulate eye drops with substances which undergo hydrolysis in aqueous environment

Figures & data

Figure 1. Degradation pathway of cefuroxime sodium (Wang & Notari, 1994).

Figure 2. Mechanism of VAN degradation in aqueous environment (Antipas et al., 1994).

Table 1. The composition of investigated SEO formulations.

Formulation	Antibiotic		Surfactant		Oil			Additives
	CEF	VAN	Tween 20	Span 80	Miglyol	Olive oil	Castor oil	Sodium citrate
CEF_TwM	5.0		5.0		90.0
CEF_TwO	5.0		5.0			90.0
CEF_TwR	5.0		5.0				90.0
CEF_SpM	5.0			5.0	90.0
CEF_SpO	5.0			5.0		90.0
CEF_SpR	5.0			5.0			90.0
VAN_TwM		5.0	5.0		90.0
VAN_TwO		5.0	5.0			90.0
VAN_TwR		5.0	5.0				90.0
VAN_SpM		5.0		5.0	90.0
VAN_SpO		5.0		5.0		90.0
VAN_SpR		5.0		5.0			90.0
CEF_TwMc	5.0		5.0		88.0			2.0
VAN_TwMc		5.0	5.0		88.0			2.0

Table 2. HPLC analysis parameters for CEF and VAN.

Parameter	CEF	VAN
Mobile phase	Acetate buffer pH 3.4:acetonitrile (95:5 v/v)	Phosphate buffer pH 3.0:acetonitrile (92:8 v/v)
Detection	Spectrophotometer: 273 nm	Spectrophotometer: 210 nm
Injection	20 μl	20 μl
Flow rate	1.5 ml/min	1.0 ml/min
Retention time	Approximately 10 min	Approximately 8 min
Column + guard column	LiChrospher^® 100, C18 (5 µm) 125 × 4 mm

Table 3. Composition of Mueller Hinton 2 LAB-AGAR™ (BioMaxima, Lublin, Poland).

Ingredient	Amount (g/l)
Beef extract powder	2.0
Starch	1.5
Casein acid hydrolysate	17.5
Agar	17.0
Water	Up to 1000 ml

Table 4. Formulations and microorganisms tested in the microbiological study.

Antibiotic 3.75% (w/w)	Formulation (solution or emulsion)	Storage of SEO	Microorganism
CEF	Solution	T0	Staphylococcus aureus ATCC 6538P
	CEF_TwM	T0 and 12 months at 25 °C
VAN	Solution	T0	Bacillus subtilis ATCC 6633
	VAN_TwM	T0 and 12 months at 25 °C

Figure 3. Microscopic images of SEO suspensions with CEF (in SEO consisted of span and Miglyol) and VAN (in SEO consisted of tween and castor oil).

Figure 4. The appearance of the obtained emulsions after dilution of SEO suspensions with water (1:10 w/w). (A) CEF emulsions with Miglyol, (B) VAN emulsions with olive oil, and (C) CEF emulsions with castor oil.

Figure 5. Microscopic images of the emulsions obtained after dilution CEF_TwM (CEF in SEO consisted of tween and Miglyol) and VAN_SpR (VAN in SEO composed of span and castor oil) suspensions with water (1:10 w/w).

Table 5. The physicochemical properties of the emulsions obtained after dilution with water (1:10) of VAN and CEF suspensions in SEO.

SEO	Oil	Surfactant	Zeta potential (mV)	pH	d50 (µm)
CEF
CEF_TwR	Castor oil	Tween 20	−43.4 ± 1.2	6.4	6.5 ± 0.2
CEF_SpR		Span 80	−66.3 ± 1.5	6.4	13.5 ± 1.0
CEF_TwM	Miglyol	Tween 20	−41.7 ± 0.7	6.4	6.2 ± 0.3
CEF_SpM		Span 80	−68.4 ± 1.0	6.4	7.9 ± 0.6
CEF_TwO	Olive oil	Tween 20	−51.0 ± 0.8	6.8	7.8 ± 0.5
CEF_SpO		Span 80	−55.6 ± 1.1	6.5	7.4 ± 0.6
VAN
VAN_TwR	Castor oil	Tween 20	−36.8 ± 0.5	5.4	4.1 ± 0.2
VAN_SpR		Span 80	−47.0 ± 0.7	5.7	14.1 ± 1.0
VAN_TwM	Miglyol	Tween 20	−31.9 ± 0.6	5.7	5.7 ± 0.3
VAN_SpM		Span 80	−40.7 ± 0.7	5.8	4.1 ± 0.4
VAN_TwO	Olive oil	Tween 20	−44.1 ± 0.9	5.6	4.0 ± 0.5
VAN_SpO		Span 80	−49.5 ± 1.1	5.6	4.6 ± 0.5

Figure 6. Stability of CEF and VAN in 5% aqueous solutions stored for 4 weeks at a temperature of 25 °C (VAN_25, CEF_25) and 40 °C (VAN_40, CEF_40).

Figure 7. HPLC chromatograms of 5% CEF solution (CEF_T0) in comparison with the solution stored for seven days at 25 °C (CEF25_T7) and 40 °C (CEF40_T7).

Figure 8. HPLC chromatograms of 5% VAN solution (VAN_T0) in comparison with the solution stored for 30 days at 25 °C (VAN25_T30) and 40 °C (VAN40_T30).

Figure 9. Changes in the content of CEF and VAN in the tested SEO suspensions during one year of storage at 40 °C (the formulation symbols correspond to the compositions presented in Table 1).

Figure 10. Changes in the content of degradation products in CEF and VAN SEO suspensions during storage at 40 °C for 12 months. *No peaks indicating the presence of degradation products were observed.

Figure 11. Examples of HPLC chromatograms of SEO suspensions stored for a year at 40 °C.

Table 6. Characteristics of emulsions prepared from the stored SEO suspensions (12 months at 40 °C).

Name	Oil	Surfactant	Time	Zeta potential (mV)	pH	d50 (µm)
CEF
CEF_TwM	Miglyol	Tween 20	T0	−41.7 ± 0.7	6.4	6.2 ± 0.3
			12 months	−42.3 ± 0.5	6.4	6.0 ± 0.3
CEF_SpM		Span 80	T0	−68.4 ± 1.0	6.4	7.9 ± 0.6
			12 months	−65.4 ± 0.8	6.4	8.1 ± 0.8
VAN
VAN_TwM	Miglyol	Tween 20	T0	−31.9 ± 0.6	5.7	5.7 ± 0.3
			12 months	−34.3 ± 0.3	5.8	5.2 ± 0.2
VAN_SpM		Span 80	T0	−40.7 ± 0.7	5.8	4.1 ± 0.4
			T12	−41.9 ± 0.8	5.7	4.5 ± 0.5

Figure 12. Stability of CEF and VAN in SEO (Miglyol and tween 20) with citrate sodium stored for 6 months at 40 °C.

Table 7. Physicochemical parameters of emulsions prepared from the stored (6 months, 40 °C) SEO (Miglyol and tween 20) suspensions with CEF or VAN containing sodium citrate.

Time (months)	pH	Zeta potential (mV)	Median droplet size (d50) (µm)
CEF_TwMc
0	7.1	−26.6 ± 0.4	5.33 ± 0.4
6	7.1	−26.8 ± 0.2	5.42 ± 0.5
VAN_TwMc
0	6.5	−28.3 ± 0.6	5.92 ± 0.3
6	6.5	−28.6 ± 0.7	6.06 ± 0.2

Figure 13. SEO (Tween 20 and Miglyol) emulsions with CEF or VAN, with the addition of sodium citrate, obtained by diluting the suspension with water at a ratio of 3:1 (w/w).

Figure 14. The inhibition zones of tested formulations: with CEF (on the left) and VAN (on the right). Freshly prepared SEO formulations and antibiotic solutions (T = 0) and SEO formulations after 12 months of storage at 25 °C (T = 12 months) are compared.

Table 8. The inhibition zones (mean ± SD) measured in the microbiological study of the freshly prepared (T = 0) and stored formulations (n = 5).

Formulation – antibiotic concentration 3.75% (w/w)	Inhibition zones (mm)
	CEF	VAN
Aqueous solution	51.6 ± 0.5	41.4 ± 0.5
Emulsion from SEO_TwM (T = 0)	51.8 ± 0.5	41.4 ± 0.4
Emulsion from SEO_TwM (T = 12 months)	51.8 ± 0.2	41.2 ± 0.4

Wang D, Notari RE. (1994). Cefuroxime hydrolysis kinetics and stability predictions in aqueous solution. J Pharm Sci 83:577–81. doi: 10.1002/jps.2600830427.

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Antipas AS, Velde DV, Stella VJ. (1994). Factors affecting the deamidation of vancomycin in aqueous solutions. Int J Pharm 109:261–9. doi: 10.1016/0378-5173(94)90388-3.

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Data availability statement

Upon reasonable request, data can be obtained from the corresponding author.