Limonene

Food Chem Toxicol. 2015 Jul;81:160-70.

The antioxidant and antigenotoxic properties of citrus phenolics limonene and naringin.[Pubmed: 25896273]

Phenolic compounds not only contribute to the sensory qualities of fruits and vegetables but also exhibit several health protective properties. Limonene and naringin are the most popular phenolics found in Citrus plants.
METHODS AND RESULTS:
In this study, we investigated the antioxidant capacities of Limonene and naringin by the trolox equivalent antioxidant capacity (TEAC) assay and the cytotoxic effects by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay in Chinese hamster fibroblast (V79) cells. The genotoxic potentials of Limonene and naringin were evaluated by micronucleus (MN) and alkaline COMET assays in human lymphocytes and V79 cells. Limonene and naringin, were found to have antioxidant activities at concentrations of 2-2000 µM and 5-2000 µM respectively. IC50 values of Limonene and naringin were found to be 1265 µM and 9026 µM, respectively. Limonene at the concentrations below 10,000 µM and naringin at the all concentrations studied, have not exerted genotoxic effects in lymphocytes and in V79 cells.
CONCLUSIONS:
Limonene and naringin at all concentrations revealed a reduction in the frequency of MN and DNA damage induced by H2O2.

Phytomedicine . 2019 Feb;53:37-42.

Gastroprotective effect of limonene in rats: Influence on oxidative stress, inflammation and gene expression[Pubmed: 30668410]

Abstract Background: In an increasing search for natural products that may heal the ulcers and avoid its recurrence, Limonene appears as a promising candidate. Hypothesis/purpose: The present study aimed to investigate the protective effect of Limonene in ethanol-induced gastric ulcers, in addition, to investigate the involvement of antioxidant and anti-inflammatory activities, besides the modulation of gene expression. Study design: Male Wistar rats were orally treated with vehicle (8% tween 80), carbenoxolone (100 mg/kg) or Limonene (25, 50 or 100 mg/kg) and then orally received ethanol to induce gastric ulcers formation. Methods: The activity of myeloperoxidase (MPO) was measured. Levels of glutathione (GSH) and activities of glutathione peroxidase (GPx), glutathione reductase (GR) and superoxide dismutase (SOD) were measured. We investigated the anti-inflammatory effect of Limonene measuring the levels of pro-inflammatory cytokines tumor necrosis factor-a (TNF-a), interleukin-6 (IL-6), interleukin-1β (IL-1β) and anti-inflammatory cytokine interleukin-10 (IL-10) by ELISA. Additionally, we investigate through real-time PCR (qPCR) the gene expression of nuclear factor-kappa B (Nf-κb), Gpx, Il-1β, Mpo, and Il-10. Results: Our results showed that Limonene 50 mg/kg was the lowest effective dose, offering 93% of reduction in gastric ulcer area compared with the vehicle. There was an increase in mucus production and higher preservation of gastric mucosa integrity after treatment with Limonene.There was a reduction in the MPO activity, a biomarker of neutrophils infiltration, and an increase in GPx activity, suggesting an antioxidant effect. Limonene displayed anti-inflammatory activity through decreasing the levels of TNF-a, IL-6, and IL-1β and increasing the level of IL-10. Limonene could down-regulate the expression of Nf-κb, Il-1β, and Mpo and up-regulate the expression of Gpx. Conclusion: Our results demonstrate that oral treatment with Limonene exerts gastroprotection through local mucosal defense mechanisms, such as increasing the mucus production, modulation of the oxidative stress and inflammatory response and inhibition of Nf-κb expression. Keywords: Antioxidant; Gastric ulcer; Gene expression; Inflammation; Limonene; qPCR.

Biotechnol Bioeng. 2015 Sep;112(9):1738-50.

Coupling limonene formation and oxyfunctionalization by mixed-culture resting cell fermentation.[Pubmed: 25786991]

Metabolic engineering strategies mark a milestone for the fermentative production of bulk and fine chemicals. Yet, toxic products and volatile reaction intermediates with low solubilities remain challenging. Prominent examples are artificial multistep pathways like the production of perillyl acetate (POHAc) from glucose via Limonene. For POHAc, these limitations can be overcome by mixed-culture fermentations.
METHODS AND RESULTS:
A Limonene biosynthesis pathway and cytochrome P450 153A6 (CYP153A6) as regioselective hydroxylase are used in two distinct recombinant E. coli. POHAc formation from glucose in one recombinant cell was hindered by ineffective coupling of Limonene synthesis and low rates of oxyfunctionalization. The optimization of P450 gene expression led to the formation of 6.20 ± 0.06 mg gcdw (-1) POHAc in a biphasic batch cultivation with glucose as sole carbon and energy source. Increasing the spatial proximity between Limonene synthase and CYP153A6 by a genetic fusion of both enzymes changed the molar Limonene/POHAc ratio from 3.2 to 1.6. Spatial separation of Limonene biosynthesis from its oxyfunctionalization improved POHAc concentration 3.3-fold to 21.7 mg L(-1) as compared to a biphasic fermentation. Mixed-cultures of E. coli BL21 (DE3) containing the Limonene biosynthesis pathway and E. coli MG1655 harboring either CYP153A6, or alternatively a cymene monooxygenase, showed POHAc formation rates of 0.06 or 0.11 U gcdw (-1) , respectively.
CONCLUSIONS:
This concept provides a novel framework for fermentative syntheses involving toxic, volatile, or barely soluble compounds or pathway intermediates.