Volatile molecules for COVID-19: A possible pharmacological strategy?

COVID-19 is a novel coronavirus disease with a higher incidence of bilateral pneumonia and pleural effusion. The high pulmonary tropism and contagiousness of the virus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), have stimulated new approaches to combat its widespread diffusion. In developing new pharmacological strategies, the chemical characteristic of volatility can add therapeutic value to the hypothetical drug candidate. Volatile molecules are characterized by a high vapor pressure and are consequently easily exhaled by the lungs after ingestion. This feature could be exploited from a pharmacological point of view, reaching the site of action in an uncommon way but allowing for drug delivery. In this way, a hypothetical molecule for COVID-19 should have a balance between its lung exhalation characteristics and both antiviral and anti-inflammatory pharmacological action. Here, the feasibility, advantages, and disadvantages of a therapy based on oral administration of possible volatile drugs for COVID-19 will be discussed. Both aerosolized antiviral therapy and oral intake of volatile molecules are briefly reviewed, and an evaluation of 1,8-cineole is provided in view of a possible clinical use and also for asymptomatic COVID-19.     

Plasma pharmacokinetics,bioavailability, and tissue distribution in CD2F1 mice of halomon,an antitumor halogenated monoterpene isolated from the red algae Portieria hornemannii

 The purpose of the present study was to define the plasma pharmacokinetics, bioavailability, and tissue distribution in mice of halomon, a halogenated monoterpene from Portieria hornemanii that is active in vitro against brain-, renal-, and colon-cancer cell lines. Halomon formulated in cremophor : ethanol : 0.154 M NaCl (1 : 1 : 6, by vol.) was injected i.v. at 20, 60, 90, or 135 mg/kg into female CD₂F₁ mice. Doses of 135 mg/kg were also given i.p., s.c., and by enteral gavage to female CD₂F₁ mice and i.v. to male CD₂F₁ mice. Plasma halomon concentrations were measured with a gas-chromatography system using electron-capture detection. Halomon concentrations were also determined in the brains, hearts, lungs, livers, kidneys, spleens, skeletal muscles, fat, red blood cells, and, if present, testes of mice given 135 mg/kg i.v. Halomon plasma pharmacokinetics were well fit by a two-compartment, open linear model and were linear between 20 and 135 mg/kg. Population estimates of parameters describing halomon plasma pharmacokinetics in female CD₂F₁ mice were developed with a standard two-stage technique and also by simultaneous modeling of data from 20-, 60-, 90-, and 135-mg/kg i.v. studies in female mice. Halomon bioavailability was 45%, 47%, and 4% after i.p., s.c., and enteral dosing, respectively. Urinary excretion of the parent compound was minimal. Halomon was distributed widely to all tissues studied but was concentrated and persisted in fat. Halomon concentrations measured in the brain were comparable with concomitant concentrations detected in plasma and most other tissues. These data and models are helpful in the simulation and evaluation of conditions produced by preclinical screening and toxicology studies. Received: 18 February 1996/Accepted: 13 June 1996     

Perillyl alcohol mediated radiosensitization via augmentation of the Fas pathway in prostate cancer cells

BACKGROUND: The management of hormone-insensitive locally advanced prostate cancer is difficult and complex and there is an urgent need for the development of effective chemotherapeutic agents intended for combination with currently available treatment modalities. METHODS: The present paper demonstrates the effectiveness of the monoterpene perillyl alcohol (POH) as potent radiosensitizer on DU145 and PC3 cell lines by performing clonogenic survival assays, cycle analysis, and assays to detect viability, apoptosis, and Fas receptor/ligand by flow cytometry. RESULTS: POH pretreatment resulted in a dose dependent sensitization to kill cell by radiation. Furthermore, POH treatment induced a transient G(2)/M arrest, enhanced the expression of the membrane bound form of the Fas ligand and sensitized the cells to Fas mediated apoptosis. CONCLUSIONS: The unique manner of radiosensitization in addition to its low toxicity profile makes POH a promising new agent for preclinical evaluation as a potential radiosensitizer in the treatment of prostate cancer.     

Phase II trial of perillyl alcohol in patients with metastatic colorectal cancer

Purpose. This is a phase II study of perillyl alcohol in the treatment of patients with metastatic colorectal carcinoma. The primary endpoint is time to progression. Secondary objectives are to evaluate objective response rate and toxicity. Patients and Methods. Eligible patients had metastatic adenocarcinoma of the colon or rectum. Patients received perillyl alcohol orally at a dose of 1200 mg/m². Dose escalation to 1,600mg/m² was allowed. Results. Twenty-seven patients were enrolled. The median time to progression was 1.8 months (range 1 to 3 mo). Four patients received less than one cycle and were not evaluable for response. Of the remaining 23, all had progressive disease. There were no complete or partial responses. Toxicity was relatively mild, with fatigue, nausea and anemia predominating. Three patients withdrew from therapy for toxicity (grade 3 belching, bloating; grade 2 nausea, fatigue, vomiting, anorexia and increase perspiration; grade 1 anorexia). Discussion. Despite preclinical evidence of anticancer activity, oral perillyl alcohol administered at this dose and formulation does not appear to have clinical antitumor activity when used for patients with advanced colorectal carcinoma.     

The biosynthesis of thymol,carvacrol, and thymohydroquinone in Lamiaceae proceeds via cytochrome P450s and a short-chain dehydrogenase

Thymol and carvacrol are phenolic monoterpenes found in thyme, oregano, and several other species of the Lamiaceae. Long valued for their smell and taste, these substances also have antibacterial and anti-spasmolytic properties. They are also suggested to be precursors of thymohydroquinone and thymoquinone, monoterpenes with anti-inflammatory, antioxidant, and antitumor activities. Thymol and carvacrol biosynthesis has been proposed to proceed by the cyclization of geranyl diphosphate to γ-terpinene, followed by a series of oxidations via p-cymene. Here, we show that γ-terpinene is oxidized by cytochrome P450 monooxygenases (P450s) of the CYP71D subfamily to produce unstable cyclohexadienol intermediates, which are then dehydrogenated by a short-chain dehydrogenase/reductase (SDR) to the corresponding ketones. The subsequent formation of the aromatic compounds occurs via keto–enol tautomerisms. Combining these enzymes with γ-terpinene in in vitro assays or in vivo in Nicotiana benthamiana yielded thymol and carvacrol as products. In the absence of the SDRs, only p-cymene was formed by rearrangement of the cyclohexadienol intermediates. The nature of these unstable intermediates was inferred from reactions with the γ-terpinene isomer limonene and by analogy to reactions catalyzed by related enzymes. We also identified and characterized two P450s of the CYP76S and CYP736A subfamilies that catalyze the hydroxylation of thymol and carvacrol to thymohydroquinone when heterologously expressed in yeast and N. benthamiana. Our findings alter previous views of thymol and carvacrol formation, identify the enzymes involved in the biosynthesis of these phenolic monoterpenes and thymohydroquinone in the Lamiaceae, and provide targets for metabolic engineering of high-value terpenes in plants.

The phenolic monoterpenes of the Lamiaceae are widely used constituents of pharmaceuticals, cosmetics, and food products (1). Extracts of plants containing thymol or carvacrol are employed in medicine for their antibacterial, anti-spasmodic, antioxidant, and anti-cancer properties. Because of their pungent, warm, and aromatic odors, they also serve as additives to cosmetics and are used in aromatherapy. Thymol and carvacrol are best known as the aroma compounds of oregano and thyme, in which they provide the herbal, pizza-like tastes that are traditionally used in Mediterranean cuisine and food preservation (2). The occurrence of phenolic monoterpenes is restricted to a few genera in the Lamiaceae (Thymus, Origanum, Satureja, and Thymbra), Apiaceae (Trachyspermum), and Verbenaceae (Lippia). Of these, the essential oils of Thymus are the most important commercial source of phenolic monoterpenes (3). Thymus vulgaris L. and Origanum species also produce the structurally related monoterpenes thymohydroquinone and thymoquinone, which were first described in the essential oil of Nigella sativa L. black seed (4). Thymohydroquinone was shown to exhibit anti-cancer activity (5), and thymoquinone displays anti-inflammatory, hepatoprotective, antioxidant, cytotoxic, and anti-cancer activities (6).To date, only a few biosynthetic pathways to pharmaceutically valuable, oxidized terpenes have been completely elucidated, such as those leading to artemisinin, paclitaxel, and the phenolic, labdane-type diterpenes of sage and rosemary (7–9). For monoterpenes, a complex biosynthetic pathway has only been described for menthol and its derivatives in Mentha (10). However, the biosynthetic pathways for phenolic monoterpenes like thymol or carvacrol remain uncharacterized. Most monoterpenes are biosynthesized by fusion of the ubiquitous C₅ intermediates, isopentenyl diphosphate, and its isomer dimethylallyl diphosphate, resulting in the formation of a C₁₀ compound, geranyl diphosphate (GDP). This acyclic intermediate is the substrate for the large enzyme family of monoterpene synthases that form cyclic or acyclic products with an enormous variety of carbon skeletons (11, 12). In previous studies in thyme and oregano, the cyclic monoterpene olefin γ-terpinene was proposed as a precursor of thymol and carvacrol (Fig. 1A). Studies with ³H-labeled γ-terpinene showed that this compound was converted into thymol and carvacrol after incubation with young thyme leaves (13). Furthermore, the essential oils of plant species rich in either thymol or carvacrol have always been reported to contain substantial amounts of γ-terpinene (3). Terpene synthases forming γ-terpinene have been identified and characterized from various Lamiaceae species (14–19). In oregano, the expression of the γ-terpinene synthase OvTPS2 was found to correlate with thymol and carvacrol content in leaves (14). Beyond γ-terpinene, however, no further precursors of phenolic monoterpenes have been identified. The aromatic hydrocarbon p-cymene was suggested to be an intermediate in thymol and carvacrol formation from γ-terpinene (20), but its participation in the phenolic monoterpene pathway and the nature of the enzymes involved in formation of the aromatic ring still remain unknown. Moreover, there is little information about the conversion of the thymol and carvacrol to thymohydroquinone.Open in a separate windowFig. 1.T. vulgaris is composed of different monoterpene chemotypes. (A) Proposed biosynthetic pathway to the phenolic monoterpenes thymol and carvacrol as well as thymohydroquinone and thymoquinone. γ-terpinene and p-cymene were suggested as intermediates in the formation of thymol and carvacrol (17). (B) Essential oil composition of the T. vulgaris chemotypes dominated by carvacrol (C type), thymol (T type), and geraniol (G type). Terpenes were extracted with hexane and analyzed by GC-MS. The following terpenes were identified: 1, α-thujene; 2, α-pinene; 3, myrcene; 4, α-terpinene; 5, p-cymene; 6, γ-terpinene; 7, cis-sabinene hydrate; 8, linalool; 9, nerol; 10, neral; 11, thymoquinone; 12, geraniol; 13, geranial; 14, thymol; 15, carvacrol; 16, geranyl acetate; 17, (E)-β-caryophyllene; 18, thymohydroquinone; and 19, germacrene D. Nonyl acetate (10 µl/mL) was added as internal standard (IS) for quantification.In this study, we investigated the biosynthetic pathway leading to the formation of thymol, carvacrol, and thymohydroquinone in thyme and oregano. We isolated and characterized six cytochrome P450 monooxygenases (P450s) of the CYP71D subfamily from thyme and oregano accessions producing high levels of thymol and carvacrol. When these CYP genes were heterologously expressed and combined with a short-chain dehydrogenase from thyme in vitro or coexpressed in vivo in Nicotiana benthamiana, thymol or carvacrol were formed. Based on the characteristics of the expressed enzymes and their reaction with other substrates, we constructed the biosynthetic pathway leading to thymol and inferred the nature of unstable intermediates. Furthermore, we identified and characterized two P450s of the CYP76S and CYP736A subfamilies that hydroxylate thymol and carvacrol to thymohydroquinone when expressed in vivo in yeast and in N. benthamiana.     

挥发油含氧单萜成分的透皮吸收促进作用与皮肤细胞毒性比较研究

目的比较中药挥发油含氧单萜成分的透皮吸收促进作用与皮肤细胞毒性。方法以中药挥发油中常见的9种含氧单萜(1,8-桉叶素、4-萜品醇、樟脑、薄荷脑、α-松油醇、香芹酮、芳樟醇、香叶醇和柠檬醛)为研究对象,比较其对布洛芬的透皮吸收促进效果和对人角质形成细胞HaCaT的细胞毒性。结果 9种含氧单萜成分均能显著促进布洛芬的透皮吸收,且皮肤细胞毒性远低于经典的化学合成促渗剂氮酮,其中1,8-桉叶素的效果最好。环状含氧单萜(1,8-桉叶素、4-萜品醇、樟脑、薄荷脑、α-松油醇、香芹酮)的透皮吸收促进效果优于链状含氧单萜(芳樟醇、香叶醇、柠檬醛),皮肤细胞毒性没有显著差异。结论环状含氧单萜更适合作为透皮吸收促进剂进行开发。     

Chemical Constituents from Stems of Cistanches deserticola

Objective To study the chemical constituents in the stems of Cistanches deserticola. Methods The compounds were purified by various column chromatographic methods and their structures were elucidated by spectroscopic(1D, 2D NMR, IR, [?]D, and MS) and chemical analyses. Results Seven compounds were isolated and identified as kankanoside D1(1), ajugol(2), cistanin(3), uridine(4), adenosine(5), 6-deoxycatalpol(6), and kankanoside D(7). Conclusion Compound 1 is a new compound.     

In vitro metabolism by mouse and human liver preparations of halomon, an antitumor halogenated monoterpene

Objectives: To characterize the enzymes responsible for and metabolites produced from the metabolism of halomon, a halogenated monoterpene that is isolated from the red algae Portieria hornemanii and has in vitro activity in the NCI screen against brain, renal, and colon cancer cell lines. Materials and methods: Mouse and human liver fractions, prepared by homogenization and differential centrifugation, were incubated with halomon, extracted with toluene, and analyzed by gas chromatography. Results: In the presence of NADPH, mouse-liver 9,000-g supernatant (S9) fractions metabolized halomon, but boiled S9 fractions did not. NADH could not substitute for NADPH. Further separation of murine hepatic S9 fractions produced a microsomal fraction that contained all of the halomon-metabolizing activity; cytosol had none. Carbon monoxide reduced murine hepatic microsomal metabolism of halomon, whereas an anaerobic, N₂ environment greatly accelerated the disappearance of halomon. Human hepatic microsomes metabolized halomon and required NADPH to do so. Carbon monoxide completely inhibited human hepatic microsomal metabolism of halomon. Unlike murine hepatic microsomal metabolism of halomon, anaerobic conditions did not enhance the metabolism of halomon by human hepatic microsomes. Neither 100 μM diethyldithiocarbamate, 1 μM quinidine, 100 μM ciprofloxacin, 3 μM ketoconazole, nor 100 μM sulfinpyrazone inhibited the metabolism of halomon by human hepatic microsomes. Both murine and human hepatic microsomes produced a metabolite of halomon. The mass spectrum of this metabolite indicated the loss of one chlorine atom and one bromine atom. Conclusions: Halomon is metabolized by mouse and human hepatic cytochrome P-450 enzymes, the identities of which remain unknown. Hepatic metabolism of halomon is very consistent with the concentrations of halomon measured in mouse tissues and urine after i.v. administration of the drug. Received: 23 February 1997 / Accepted: 9 June 1997     

Genotoxicity of lavender oil,linalyl acetate,and linalool on human lymphocytes in vitro

The potential genotoxicity of lavender essential oil and its major components, linalool, and linalyl acetate, was evaluated in vitro by the micronucleus test on peripheral human lymphocytes. In the range of non‐toxic concentrations (0.5–100 μg/ml), linalyl acetate increased the frequency of micronuclei significantly and in concentration‐dependent manner; lavender oil did so only at the highest concentration tested, whereas linalool was devoid of genotoxicity. None of the tested substances led to an increase in nucleoplasmic bridges or nuclear buds frequency. These findings suggest that the mutagenic activity of lavender oil can be related to the presence of linalyl acetate, which seems to have a profile of an aneugenic agent. Environ. Mol. Mutagen. 52:69–71, 2011. © 2010 Wiley‐Liss, Inc.     

Chemopreventive effect of myrtenal on bacterial enzyme activity and the development of 1,2-dimethyl hydrazine-induced aberrant crypt foci in Wistar Rats

Colon cancer remains as a serious health problem around the world despite advances in diagnosis and treatment. Dietary fibers are considered to reduce the risk of colon cancer as they are converted to short chain fatty acids by the presence of anaerobic bacteria in the intestine, but imbalanced diet and high fat consumption may promote tumor formation at different sites, including the large bowel via increased bacterial enzymes activity. The present study was conducted to characterize the inhibitory action of myrtenal on bacterial enzymes and aberrant crypt foci (ACF). Experimental colon carcinogenesis induced by 1,2-dimethylhydrazine is histologically, morphologically, and anatomically similar to human colonic epithelial neoplasm. Discrete microscopic mucosal lesions such as ACF and malignant tumors function as important biomarkers in the diagnosis of colon cancer. Methylene blue staining was carried out to visualize the impact of 1,2-dimethylhydrazine and myrtenal. Myrtenal-treated animals showed decreased levels of bacterial enzymes such as β-glucuronidase, β-glucosidase, and mucinase. Characteristic changes in the colon were noticed by inhibiting ACF formation in the colon. In conclusion, treatment with myrtenal provided altered pathophysiological condition in colon cancer-bearing animals with evidence of decreased crypt multiplicity and tumor progression.