Hydroxyl Group-Targeted Conjugate and Its Self-Assembled Nanoparticle of Peptide Drug: Effect of Degree of Saturation of Fatty Acids and Modification of Physicochemical Properties

Figures & data

Table 1 Physicochemical Properties of LEU Acetate

Factors	Physicochemical Properties
2D structure
Molecular weight	1269.45 g/mol
Chemical formula	C59H84N16O12. C2H4O2
BCS class	IV
Half life	1 mg intravenous administration: 3hr
Route of administration	IM injection
Dose strength	3.75, 7.5 mg per month
Solubility	≥ 66.66 mg/mL (52.51 mM) in water
log P	−2.7–1.07

Table 2 Physicochemical Properties of Different Degree of Saturation of C18 Fatty Acids Selected for LEU Conjugation

Fatty Acids	Structure	Physicochemical Properties
Stearic acid		Chemical formula: C18H36O2 Features: C18, saturated Molecular weight: 284.48 g/mol Melting Point: 69.3°C log P: 8.23
Oleic acid		Chemical formula: C18H34O2 Features: C18, mono-unsaturated Molecular weight: 282.47 g/mol Melting Point: 13.4°C log P: 7.64
Linoleic acid		Chemical formula: C18H32O2 Features: C18, polyunsaturated Molecular weight: 280.45 g/mol Melting Point: −5°C log P: 7.05

Figure 1 Schematic diagram for the hydroxyl group targeted LEU-oleic acid conjugate (LOC).

Table 3 Composition Gradients of Prep-HPLC

Time (min)	DW (%)	ACN (%)
0	80	20
15	50	50
35	30	70
45	0	100
50	0	100

Figure 2 ¹H-NMR spectra of LEU, fatty acids and LEU-fatty acid conjugate having different saturation levels: (A) LEU acetate, (B) Stearic acid, (C) Oleic acid, (D) Linoleic acid, (E) LSC, (F) LOC, (G) LLC.

Figure 3 MALDI-TOF results of LEU-fatty acid conjugate after purification by Prep-HPLC: (A) LSC, (B) LOC, (C) LLC.

Figure 4 FT-IR spectra of LEU acetate, fatty acids, physical mixtures, and LEU-fatty acid conjugate having different saturation levels (LSC, LOC, LLC).

Figure 5 DSC thermograms of LEU acetate, fatty acids, physical mixtures, and LEU-fatty acid conjugate having different saturation levels: (A) LSC, (B) LOC, (C) LLC.

Table 4 Physicochemical Properties of Self-Assembled L18FNs

Nanoparticle	Particle Size (nm)	Polydispersity Index	Zeta Potential (mV)
LSN	180.11 ± 11.27	0.446 ± 0.108	31.24 ± 4.12
LON	144.47 ± 7.49	0.285 ± 0.028	57.13 ± 2.17
LLN	79.9 ± 0.40	0.144 ± 0.017	76.13 ± 1.32

Note: Mean ± standard deviation (n=3).

Figure 6 FE-SEM images of LEU acetate and self-assembled L18FNs: (A) LEU acetate, (B) LSN, (C) LON, (D) LLN.

Figure 7 FE-TEM images of self-assembled L18FNs: (A) LSN, (B) LON, (C) LLN.

Figure 8 Degradation profiles of LEU (A) and L18FCs (B), and conversion profiles from L18FCs to cumulative LEU concentration (C) in human plasma (n=3).

Figure 9 Permeability profile of LEU (A) and L18FNs (B) as a function of time in pH 7.4 PBS through permeation barrier-membrane (PB-M) using Franz diffusion cell (n=3).

Figure 10 FE-SEM images of the PB-M surface (A), and 120 min after starting LON permeation (B).