Engineered andrographolide nanosystems for smart recovery in hepatotoxic conditions

Figures & data

Figure 1 (A) FTIR scan of AG; (B) electrospray ionization mass spectra of AG; (C) X-ray diffraction spectra of AG and CsAGNP nanosystems; (D) FTIR scan of biopolymer Cs; (E) FTIR scan of CsAGNP nanosystem.

Abbreviations: Cs, chitosan; FTIR, Fourier-transform infrared; AG, andrographolide; CsAGNP, chitosan-modified AG nanoparticle.

Figure 2 (A) Size distribution of CsAGNPs; (B) 3-D atomic force microscopy data of CsAGNPs; (C) In vitro dissolution studies for CsAGNPs (blue) in phosphate buffer, pH 7.4.

Abbreviations: AG, andrographolide; CsAGNPs, chitosan-modified AG nanoparticles.

Table 1 Liver function markers and lipid peroxidation in treatment conditions

Groups	AST (IU/L)	ALT (IU/L)	ALP (IU/L)	Hepatic MDA level (nM/mg liver tissue)
Control	48.33±5.44	71.50±8.36	66.00±8.39	45.67±6.56
CCl4	157.83±19.77	201.83±19.53	187.83±17.16	86.83±10.17
AG	106.67±13.41*	136.67±20.26*	140.00±16.20*	63.33±12.16*
CsAGNPs	84.83±7.25*, **	105.50±10.60*, **	108.83±14.93*, **	57.00±8.07*, **

Notes:

* P<0.001, significant difference compared with CCl₄-treated group

** P<0.05, significant difference compared among AG- and CsAGNP-treated groups. Results are expressed as means ± standard deviation (n=6).

Abbreviations: AST, aspartate aminotransferase; ALT, alanine transaminase; ALP, alkaline phosphatase; MDA, malonyldialdehyde; AG, andrographolide; CsAGNPs, chitosan-modified AG nanoparticles.

Figure 3 Histopathological changes of liver cytoarchitecture after various treatments, magnification 400×, normal liver, CCl₄-treated liver, AG-treated liver, CsAGNP-treated liver.

Abbreviations: AG, andrographolide; CsAGNPs, chitosan-modified AG nanoparticles.

Figure 4 Levels of hepatic enzymatic antioxidant markers before and after CCl₄-induced hepatotoxicity. Results are expressed as means ± standard deviation (n=6).

Notes: *P<0.001, significant difference compared with CCl₄-treated group; **P<0.05 significant difference compared among AG- and CsAGNP-treated groups.

Abbreviations: AG, andrographolide; CsAGNPs, chitosan-modified AG nanoparticles; SOD, superoxide dismutase; CAT, catalase; GPx, glutathione peroxidase; GR, glutathione reductase; GST, glutathione S-transferase.

Table 2 Nonenzymatic hepatic antioxidants in CCl₄-induced hepatotoxicity

Groups	Hepatic GSH (μM/mg liver tissue)	Hepatic GSSG (μM/mg liver tissue)	GSH/GSSG
Control	8.618±1.20	0.755±0.09	11.66±2.86
CCl4	4.567±0.66	1.082±0.15	4.28±0.82
AG	6.17±1.21	0.943±0.12	6.69±1.81
CsAGNPs	7.417±1.33*	0.843±0.13*	9.07±2.72*

Notes:

* P<0.05, significant difference compared with CCl₄-treated group. Results expressed as means ± standard deviation (n=6).

Abbreviations: GSH, glutathione; GSSG, GSH disulfide; AG, andrographolide; CsAGNPs, chitosan-modified AG nanoparticles.

Table 3 Serum levels of inflammatory markers

Groups	Serum TNFα level (pg/mL)	Serum IL-6 level (pg/mL)
Control	77.50±9.20	52.8±6.09
CCl4	247.82±20.66	109.33±11.15
AG	149.13±16.26*	71.94±8.12*
CsAGNPs	112.18±12.52*,**	63.84±6.13*

Notes:

* P<0.001, significant difference compared with CCl₄-treated group

** P<0.05, significant difference compared among AG- and CsAGNP-treated groups. Results expressed as means ± standard deviation (n=6).

Abbreviations: AG, andrographolide; CsAGNPs, chitosan-modified AG nanoparticles.

Figure 5 Fluorescein isothiocyanate-labeled nanoparticle localization in the liver after 4 hours of administration.

Notes: (A) Transmitted image; (B) fluorescent image.

Figure S1 Ultraviolet maximum absorbance for andrographolide extracted in 5 μg/mL chloroform solution.

Note: The arrows (numbered 1, 2, and 3), indicate specific peak positions for andrographolide in UV-visible scanning.

Figure S2 Engineered andrographolide nanosystems in water: transmission electron microscopy.

Figure S3 Irregular andrographolide crystals in water: transmission electron microscopy.