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JKM > Volume 45(2); 2024 > Article
Yu, Jin, Kim, Noah, Kim, and Lim: A comparative study on enhanced anti-inflammatory activity of Cannabis root extract prepared by combined processing process



Cannabis root is a medicinal plant that has been used in traditional medicine around the world. However, in previous studies using simple extracts, the biological activity was not relatively prominent compared to other herbal medicines. The aim of the present study is to confirm the enhancement of anti-inflammatory activity and bioactive compound of Cannabis root extract prepared by combined processing process.


A series of processes including repeated steam and dry, fermentation, hydrothermal extraction and ethanol extraction was applied to Cannabis root. The antioxidant content of cannabis root extracts obtained through a combined processing process was investigated by analyzing the total phenolic, tannin, and flavonoid contents. Anti-inflammatory effects were tested in LPS-treated RAW264.7 cells. The anti-inflammatory mechanism was examined by western blot. Finally, the component profile of Cannabis root extract was analyzed using High Performance Liquid Chromatography (HPLC) and Thin layer chromatography (TLC).


The cannabis root extract prepared by complex processing process had higher antioxidant and anti-inflammatory effects than simple extract. Total phenolic and tannin contents were significantly increased, and DPPH free radical inhibition activity was strengthened by combined processing process. Increased NO production and iNOS expression in LPS-treated RAW264.7 cells were decreased in a concentration-dependent manner upon extract treatment by complex processing process. Additionally, the Stigmasterol content of Cannabis root extract was increased through a complex processing process.


Further research is needed on the mechanisms and substances that exhibit the anti-inflammatory effects of Cannabis roots extract prepared by complex processing process.

Fig. 1
Total phenolic content, tannin content, flavonoid content of Cannabis root extract prepared by combined processing process.
Fig. 2
DPPH radical scavenging activity of Cannabis root extract by combined processing process. (A) DPPH radical scavenging effects for various concentration (B) IC50 values of DPPH assay. The values are significantly different at *P < 0.05, ***P < 0.001.
Fig. 3
Effect of Cannabis root extract by combined processing process on RAW264.7 cell viability. RAW264.7 cells were incubated with different concentration of Cannabis root extract (0–500 μg/ml) for 24 h. Cell viability was measured by EZ-Cytox assay as described in materials and methods. Results are the means ± SDs of three independent experiments.
Fig. 4
Comparison of anti-inflammatory effects of Cannabis root extract prepared by combined processing process. RAW264.7 cells were co-treated with LPS (1 μg/ml) and Cannabis root extracts for 24 h. (A) Inhibitory effect of Cannabis root extract by combined processing process on NO production. (B) The levels of iNOS protein expression in LPS-induced RAW264.7 cells were immunoblotted and analyzed. Results are the means ± SDs of three independent experiments. #P<0.05, ##P < 0.01 vs. LPS-untreated controls and *P < 0.05, **P < 0.01 vs. LPS-treated control.
Fig. 5
Comparison of compound peaks of Cannabis root extracts (A) HPLC analysis of all Cannabis root extracts prepared by different processing process. (B) Bar chart of peak area results for comparison.
Fig. 6
TLC(Thin-layer chromatography) of the different extracts of Cannabis root showing the content of Stigmasterol. Spot ① indicates Stigmasterol, and Spot ② indicates water extract of Cannabis root, and Spot ③ indicates CSR-Nt, and Spot ④ indicates CSR-4SD, and Spot ⑤ indicates CSR-F-Lr, and Spot ⑥ indicates CSR-4SD-F-Lr, respectively.
Table 1
Schematic diagram of the combined processing process and yield of each processed Cannabis root extract.


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