**4. Anti-inflammatory activity of SMt isolated from vegetal material obtained by biotechnological processes**

Some SMt with significant anti-inflammatory activity have been obtain from MP through employment some biotechnological processes. From cell suspension cultures *Sphaeralcea angustifolia*, two compounds with important anti-inflammatory activity (evaluated in murine models) were isolated. The cell suspension was developed in MS medium with total nitrate 2.74 mM, under this condition was obtained scopoletin, sphaeralcic acid (**22**) and tomentin (**23**). From the CH2Cl2:CH3OH extract sphaeralcic acid (**22**) and tomentin (**23**) were isolated; these compounds showed 58 and 66% antiinflammatory activity, in the carrageenin model at 45 mg/kg administered by intraperitoneal (i.p.) route. On the other hand, in the topical anti-inflammatory model (TPA, 12-O-TetradecanoylPhorbol-13-Acetate), tomentin (225 mM/ear) exhibited 57% inhibition in the formation of auricular edema, while sphaeralcic acid (174 mM/ear) revealed 86% inhibition with a dose-dependent effect and one half of the Effective Dose (ED50) = 93 mM. Sphaeralcic acid is the most active compound in both models (topical as well as systemic) [124, 131].

*Modulation of Secondary Metabolites among Mexican Medicinal Plants by Using Elicitors… DOI: http://dx.doi.org/10.5772/intechopen.99888*

**Figure 1.** *Chemical structure of some polyphenols and other SMt with biological activity.*

#### *Modulation of Secondary Metabolites among Mexican Medicinal Plants by Using Elicitors… DOI: http://dx.doi.org/10.5772/intechopen.99888*

In another study, the anti-inflammatory activity of the cell suspension culture from *S. angustifolia* is described. In this case, aseptic-leaf explants and Naphthalene Acetic Acid (NAA, such as auxin) in several concentrations (0, 0.5, 1.0, and 2.0 mg/L) in combination with a constant concentration of Kinetin (KIN) were used. For the cell suspension culture, they utilized 4% initial inoculum in MS medium with 2.74 mM of the total nitrates, 1 mg/L of NAA and 0.1 mg/L of KIN and supplemented with 30 g/L of sucrose. The main SMt identified in this suspension cultures were the same compounds (scopoletin, tomentin, and sphaeralcic acid). Scopoletin was excrete in the culture medium, although it also accumulated in the biomass. For evaluation of the anti-inflammatory activity, the authors prepared the CH2Cl2:MeOH extract of the cell's suspension from *S. angustifolia* and this extract was administered i.p. in male ICR mice (35 g) employing the carrageenin model. This extract showed ED50 = 137.63 mg/ kg; sphaeralcic aid and tomentin at 45 mg/kg inhibited 67 and 62%, respectively on carrageenan assay and sphaeralcic acid at 1 mg/ear was more active in TPA assay, showed ED50 = 93 mM and tomentin showed 48% of inhibition at 1 mg/ear [132, 133]. In addition, the same extract from biomass of cells in suspension of *S. angustifolia* at 100 mg/kg (with 0.10 mg of scopoletin, 0.10 mg of tomentin and 0.19 mg of sphaerelcic acid), as well as tomentin (20 mg/kg) were active as anti-inflammatory agent and reduced the mean body weight lost in Freund adjuvant- and kaolin/carrageenan-induced arthritis, respectively. In this assay, the organic extract and tomentin reduced the levels of pro-inflammatory interleukins such as IL-1β, IL-6 and TNF-α and increased levels of IL-4 and IL-10 (anti-inflammatory cytokines) [133].

In parallel with obtaining cells *in vitro* of *S. angustifolia*, the authors performed a preclinical phase study (in rats). The CH2Cl2 extract of the aerial parts of *S. angustifolia* (wild material) was tested in chronic inflammation model induced with complete Freund's adjuvant (polyarthritis) The administration of the extract at 100 mg/kg/day during 8 days showed sustained and significant inhibition of edema, being of 62.6% [134]. A double-blind clinical phase study with the extract of *S. angustifolia* (wild material) standardized at 1% hydroxycoumarin content was conducted; the experiment was performed on 130 patients diagnosed with osteoarthritis. 55 of them were treated with standardized extract of *S. angustifolia* (gel) and 75 patients were treated with Diclofenac (2%). The therapeutic effectiveness of the gel administered topically for 4 weeks was 89%, while that of the control group (Diclofenac) was 91.3%; it was highlighted in the study that patients who received the treatment (gel of the standardized extract) did not exhibit adverse effects and did show an improvement in their disorder [135].

Another plant utilized in Mexican ethnomedicine is *Lopezia racemosa* Cav. Callus cultures in MS medium were obtain with variable amounts of NAA, 2,4-Dichlorophenoxiacetic acid (2,4-D) and 6-BenzylAminoPurine (BAP). The authors carried out 10 treatments with the previously mentioned PGR. In this case, they employed three types of explants (hypocotyl, stem nodule, and leaf) and several treatments. The combination of 1.0 mg/L of 2,4-D plus 0.5 mg/L of BAP was the best. From these callus material two novel compounds: 6-O-palmitoyl-3-O-β-Dglucopyranosylcampesterol (174.0 μg/g of biomass) and 6-O-palmitoyl-3-O-β-Dglucopyranosyl-β-sitosterol were isolated. When quantifying these compounds, the authors observed that the wild plant contains less quantity than the callus. The topical anti-inflammatory activity of the biomass obtained from the callus was evaluate in the TPA model on CD-1 male mice at 1 mg/ear. Three extracts (hexanic, CH2Cl2, and methanol), was tested and showed 48.74, 57.14, and 16. 81% of inhibition, respectively. The CH2Cl2 extract was the most active, with a half-maximal Inhibitory

Concentration (IC50) = 0.93 mg/ear. On the other hand, the pure compound (6-Opalmitoyl-3-O-β-D-glucopyranosyl-campesterol) was tested in the same model at 1 mg/ear showing a 57.14% inhibition, with IC50 = 0.45 mg/ear [100].

The lipophilic extract containing beta-carotene (LMBC) from plant cell cultures of *Cleome spinosa* was evaluate in two *in vivo* models to determine the antiinflammatory and antinociceptive activities in Swiss Webster (SW) mice of both sexes. The callus culture was obtained of the MS medium supplemented with 1 mg/L of 4- amino-3,5,6-trichloropicolinic acid (picloram) and sub-cultured to culture medium with the same composition at 4-week intervals. The anti-inflammatory activity in carrageenan model at 10 mg/kg by i.p. via was evaluated. LMBC was inactive with respect to extract from whole plant, which showed more than 50% inhibition of edema at the same dose. On the other hand, the LMBC (at 50 mg/kg) showed around 68% decrease in writhes, these data were very similar to that shown in wild plant, and the effect was better than dipyrone (at 100 mg/kg) used as positive control. The authors concluded that the results of LMBC are particularly important; since this active SMt of medicinal interest can be continuously obtain from callus cultures [136].

*Buddleja cordata* is other medicinal species utilized to treat diseases related with inflammation. This cell suspension was obtained in MS medium supplemented with NAA (9.05 μM) and Kin (2.32 μM). The anti-inflammatory activity of the extracts from wild plant and of the cell suspension cultures were describe. In both extracts, the verbascoside content was quantified by HPLC methods. The extract of the cells suspension has 87.48 mg verbascoside/g Dry Matter (DM), while that the same extract from wild plant only contained 47.34 mg of verbascoside/g DM. In addition, acute toxicity in Balb/C mice of the both extracts were also determined, with half of a Lethal Dose (LD50) of >2 g/kg. On the other hand, the topical anti-inflammatory effect of the wild plant extract and of the cell suspension was assay. The ED50 values was 3.93 and 1.26 mg/ear, respectively, cell suspension extract was the most active due to its greater content of verbascoside. Evaluation of both extracts in the carrageenan model (systemic inflammation), showed ED50 = 251.26 and 204.62 mg/kg for wild plant and cells suspension extracts, respectively; in this case, the latter extract was more active. In the chronic inflammation model (the arthritis model induced with complete Freud's adjuvant), both extracts showed moderate anti-inflammatory activity (<35%) and favored weight increase in animals with arthritis. The authors concluded that the cell suspension culture of *Buddleja cordata* obtained through the biotechnological process contained a better anti-inflammatory activity; therefore, it represents a source for obtaining this type of secondary metabolite-of-pharmacological-interest [48].

*Cnidoscolus chayamansa* is medicinal plant whit anti-inflammatory, antiprotozoal, hepatoprotective, hypoglycemic and antimycobacterial activities [137–139]. Recently, a biotechnology processes was described to obtain callus using BAP (5 mg/L) and 2,4- D (2.5 or 5 mg/L), this callus was used as a biotechnological alternative for *in vitro* propagation of this plant [140]. After that, this callus was use to establish a cell suspension culture. From the cell suspension, organic extract was prepared and its antioxidant, antibacterial and anti-inflammatory activities were determined, as well as the main SMt was quantified by HPLC analysis. In cell suspension, lupeol acetate (38.1 mg/g DW) was obtained as a main constituent and scopoletin (3.6 mg/g DW) was also quantified; in wild material, both compounds were isolated in low quantity. The organic extract was active against *Staphylococcus aureus, S. coagulase* and *Listeria monocytogenes,* and a moderate antioxidant and anti-inflammatory activities (in TPA and carrageenan models) showed [28, 141].

*Modulation of Secondary Metabolites among Mexican Medicinal Plants by Using Elicitors… DOI: http://dx.doi.org/10.5772/intechopen.99888*
