## 4 Biological Activity of Cannabis Metabolites
### Biological Activity of Cannabis Alkaloids and Other Nitrogen-Containing Compounds
#### 4.2.1 In Vitro Activity of Cannabis Nitrogen-Containing Compounds  
Hordenine from barley (Hordeum vulgare) is reported to have a repellent activity towards grasshoppers [29]. It also possesses antibacterial activity, with a minimal inhibitory concentration (MIC) of 2.5 mg/mL against the Pseudomonas aeruginosa PAO1 strain [64]. [PERSON] et al. (2018) also tested the quorum-sensing (QS) inhibitory activity of hordenine [64]. Acylated homoserine lactones (AHLs) are gram-negative autoinducers necessary for biofilm formation. Two notable AHLs were confirmed extracellularly by LC-MS/MS from \(P\). Figure 15: Structure of some polyamines from the three different types: diamines, triamines, and tetraamines. aeruginosa PAO1: N-(3 oxcododecanoyl)-L-homoserine lactone (3-oxo-C12-HSL) and N-butanol-L-homoserine lactone (C4-HSL). Thus, [PERSON] et al. (2018) tested the effect of hardening exposure during 24 h in the culture medium, with concentrations of 0.5, 0.75, and 1.0 mg/mL [64]. They observed a reduction of 69%, 74%, and 79% for C4-HSL, with respect to the listed concentrations, when compared to control. They noted a reduction ranging from 24% to 66% for 3-oxo-C12-HSL with the same hardening concentrations, suggesting interference by hardening in AHL production. Additionally, they reported hardening's ability to inhibit biofilm formation in the PAO1 strain, with concentrations lower than 1.0 mg/kg, as well as significantly enhance the efficacy of the antibiotic netilmicin in biofilm formation inhibition [64]. Hordenine is also capable of disrupting and thinning preformed biofilms, especially when combined with netilmicin, and suppressing virulence factors of _P. aeruginosa_, namely protease and elastase activity, pyocyanin and pyoverdine production, and rhamn and alginate activities, even downregulating QS-related genes _lasl_, _lasR_, _rhll_, and _rhll_ by nearly 50% [64]. Similarly, hordenine's quorum-sensing inhibitory activity was tested against the _Serratia marcescens_ NJ01 strain by [PERSON] et al. (2019) [65]. They reported an MIC of 2.5 mg/mL for _S. marcescens_, which is the same as _P. aeruginosa_ PAO1, and noted that concentrations of 25, 50, and 100 mg/mL of hordenine in an LB broth medium for 24 h resulted in a diminution of C4-HSL levels of 40%, 60%, and 80%, respectively [65]. Withholding details, they found results resembling those obtained with _P. aeruginosa_ PAO1, with hardening reducing biofilm autoinducer levels, inhibiting biofilm formation, lessening virulence factors, altering preformed biofilm architecture (resulting in higher antibiotic sensitivity with ciprofloxacin), and downregulating QS-related genes. [PERSON] and [PERSON] (2018) investigated hordenine's \(\beta_{2}\)-adrenergic receptor (\(\beta_{2}\)-AR) agonistic capabilities in HEK293 cells expressing either human or mouse \(\beta_{2}\)-AR, using a luciferase assay [66]. After 6 h of hordenine exposure, the cells were tested for luciferase activity. They found hordenine to be an agonist of mouse \(\beta_{2}\)-AR in a dose-dependent manner, with little to no effect with 10 uM, a twofold increase in luciferase activity compared to control with 30 uM of hordenine, and a significant more than sevenfold increase at 100 uM of hordenine. Regarding HEK293 cells expressing human \(\beta_{2}\)-AR, hordenine at 100 uM was also confirmed as an agonist of the receptor. HCAA Activity In Vitro  
Synthesized N-trans-coumaroyltyramine was tested for antiproliferative activity against Jurkat and U937 human cell lines. Cells were treated with concentrations of 30, 60, and 90 uM of N-trans-coumaroyltyramine for 18 h, and total cell death was measured by the trypan blue exclusion method and a nonradioactive cell proliferation method by [PERSON] (2002) [67]. They reported a notable decrease in the number of living cells with 30 uM of N-trans-coumaroyltyramine (40% for U937, 32% for Jurkat), with increasing efficacy at higher concentrations.