GLUTATHIONE-LOADED NANOEMULSION 261 were subjected further to a 90-d testing period. The results showed that NE-19 was the most stable formulation for the fabrication of GSH stability. NE-18 showed phase separation at the end of the testing period. Freshly prepared B-19 and NE-19 were milky white, had pleasant odor, and had no phase separation after accelerated centrifugation test at the start of the testing period and remained stable at the end of the 90-d period (Figure 2). No phase separation was found in samples of B-19 and NE-19 kept at 4 and 25°C after each subse- quent time interval, but at 90 d, phase separation was observed in samples of B-19 and NE-19 kept 40 and 40°C ± 75 RH. There was no turbidity seen. The pH values mea- sured at various storage conditions are shown in Table II. The average droplet size and zeta potential, mobility, and electrical conductivity of freshly prepared B-19 recorded were 184.25 nm, -30.85 mV, -2.911 μm cm/Vs, 0.254, and 0.0236 mS/cm, respec- tively. The average droplet size and zeta potential, mobility, polydispersity, and electrical conductivity of NE-19 recorded are shown in Table III. The droplet size distribution and intensity distribution of NE-19 are shown in Figure 3. The Power’s law math model provided the analysis of the behavior of data sets. The vis- cosities of freshly prepared B-19 and NE-19 were found to be 3,363.492 and 4,425.712 cP, respectively. Flow index of B-19 and NE-19 were 0.28 and 0.21, respectively. Confi dence of fi t of the base and NE-19 were found to be 99.6 and 99.8, respectively. Rheograms for B-19 and NE-19 have been shown in Figure 4. DISCUSSION Nanoemulsions are appealing delivery systems because of their high stability, low amount of surfactants, low viscosity, and good development. They are signifi cant because of their small droplet size and close contact with the S. corneum, so the active molecules easily reach the site of action (1). In topical formulations, GSH is at risk of degradation as it oxidizes quickly in water. In GSH-loaded (o/w) nanoemulsion, GSH can be prevented from degradation as it remains inside oil globules dispersed in the aqueous phase. That is why a GSH-loaded o/w nano- emulsion was fabricated to increase the stability of GSH. The modifi ed method was adopted to fabricate a GSH-loaded nanoemulsion to achieve the smallest globule size because the smaller the droplet size, the more stable the nanoemulsion Figure 2. (A) B -19 and (B) NE-19.

JOURNAL OF COSMETIC SCIENCE 262 will be (1,15). Construction of pseudo-ternary phase diagrams is the best way to study all kinds of formulations that can be derived from the mixing of surfactants, water, and oil because the diagrams can cover all probabilities of mixing ratios and possible areas of fi nding nanoemulsion. All kinds of formulations derived from mixing of oil, water, and surfactants can be studied with the help of a ternary phase diagram for nanoemulsion formulations of liquid paraffi n oil, water, and Smix (tween 80 + span 80) with different HLB values 9, 10, 11, and 12. Smix was preferred because a single surfactant was not enough to form the layer at the interface while developing a nanoemulsion. Thirty-three nano- emulsion formulations were derived from pseudo-ternary phase diagrams and subjected to a stability study of 90 days to select the most stable nanoemulsion system. Formula- tion NE-19 was found to be the most stable formulation having a HLB value of 10. The formulation NE-19 was stable because there was an optimum concentration available to make the interfacial fi lm around the oil globules dispersed in the aqueous phase. This ratio of Smix (T80 = 4.26%: S80 = 3.74%) increased the interfacial tension of the interfacial fi lm and developed a stable nanoemulsion (18). Centrifugation involves the application of centripetal force to separate two immiscible liquids and is performed to check whether a nanoemulsion is stable or not. The ability of a formulation to resist change in its physical and chemical characteristics is called its stability (16). NE-19 and B-19 were centrifuged at 5,000 rpm for 10 min. However, no phase separation occurred in the NE-19 and B-19 after centrifugation. The formulations were tested for kinetic stability by centrifugation, and only the formulations that exhibited Table II pH Values for B-19 and NE-19 Kept at 4, 25, 40, and 40°C + 75% RH Storage time 4°C 25°C 40°C 40°C + 75% RH B-19 NE-19 B-19 NE-19 B-19 NE-19 B-19 NE-19 Fresh 5.92 5.79 5.92 5.79 5.92 5.79 5.92 5.79 24 h 5.72 5.53 5.61 5.59 5.64 5.53 5.43 5.49 48 h 5.43 5.39 5.28 5.48 5.35 5.11 4.98 5.38 72 h 5.31 5.28 5.12 5.33 5.21 4.95 4.92 5.24 7 d 5.14 5.16 5.04 5.21 5.09 4.87 4.87 5.09 14 d 4.98 5.03 4.91 4.98 4.93 4.84 4.81 4.92 21 d 4.82 4.92 4.68 4.72 4.82 4.72 4.77 4.81 28 d 4.69 4.78 4.49 4.67 4.71 4.68 4.64 4.73 45 d 4.54 4.76 4.41 4.63 4.63 4.65 4.51 4.69 60 d 4.49 4.61 4.39 4.60 4.54 4.61 4.39 4.67 90 d 4.41 4.53 4.35 4.47 4.48 4.43 4.31 4.37 Table III Average Droplet Size and Zeta Potential, Polydispersity, Mobility, and Electrical Conductivity of NE-19 after Different Time Intervals at 25°C Sample name Droplet size (nm) Zeta potential (mV) Polydispersity index Mobility (μm cm/Vs) Electrical conductivity (mS/cm) Fresh NE-19 96.05 -37.1 0.189 -2.726 0.0141 NE-19 after 30 d 155.0 -34.2 0.208 -2.911 0.0236 NE-19 after 60 d 181.3 -34.8 0.286 -2.684 0.0269 NE-19 after 90 d 190.09 -34.0 0.277 -2.459 0.0289