Identification and Allelopathy of Green Garlic (Allium sativum L.) Volatiles on Scavenging of Cucumber (Cucumis sativus L.) Reactive Oxygen Species.

Garlic and formulations containing allicin are used widely as fungicides in modern agriculture. However, limited reports are available on the allelopathic mechanism of green garlic volatile organic compounds (VOCs) and its component allelochemicals. The aim of this study was to investigate VOCs of green garlic and their effect on scavenging of reactive oxygen species (ROS) in cucumber. In this study, green garlic VOCs were collected by HS-SPME, then analyzed by GS-MS. Their biological activity were verified by bioassays. The results showed that diallyl disulfide (DADS) is the main allelochemical of green garlic VOCs and the DADS content released from green garlic is approximately 0.08 mg/g. On this basis, the allelopathic effects of green garlic VOCs in vivo and 1 mmol/L DADS on scavenging of ROS in cucumber seedlings were further studied. Green garlic VOCs and DADS both reduce superoxide anion and increase the accumulation of hydrogen peroxide of cucumber seedlings. They can also regulate active antioxidant enzymes (SOD, CAT, POD), antioxidant substances (MDA, GSH and ASA) and genes (CscAPX, CsGPX, CsMDAR, CsSOD, CsCAT, CsPOD) responding to oxidative stress in cucumber seedlings.

Diallyl Disulfide Induces Apoptosis and Autophagy in Human Osteosarcoma MG-63 Cells through the PI3K/Akt/mTOR Pathway.

Diallyl disulfide (DADs), a natural organic compound, is extracted from garlic and scallion and has anti-tumor effects against various tumors. This study investigated the anti-tumor activity of DADs in human osteosarcoma cells and the mechanisms. MG-63 cells were exposed to DADs (0, 20, 40, 60, 80, and 100 µM) for different lengths of time (24, 48, and 72 h). The CCK8 assay results showed that DADs inhibited osteosarcoma cell viability in a dose-and time-dependent manner. FITC-Annexin V/propidium iodide staining and flow cytometry demonstrated that the apoptotic ratio increased and the cell cycle was arrested at the G2/M phase as the DADs concentration was increased. A Western blot analysis was employed to detect the levels of caspase-3, Bax, Bcl-2, LC3-II/LC3-I, and p62 as well as suppression of the mTOR pathway. High expression of LC3-II protein revealed that DADs induced formation of autophagosome. Furthermore, DADs-induced apoptosis was weakened after adding 3-methyladenine, demonstrating that the DADs treatment resulted in autophagy-mediated death of MG-63 cells. In addition, DADs depressed p-mTOR kinase activity, and the inhibited PI3K/Akt/mTOR pathway increased DADs-induced apoptosis and autophagy. In conclusion, our results reveal that DADs induced G2/M arrest, apoptosis, and autophagic death of human osteosarcoma cells by inhibiting the PI3K/Akt/mTOR signaling pathway.

Allyl methyl sulfide, a garlic active component mitigates hyperglycemia by restoration of circulatory antioxidant status and attenuating glycoprotein components in streptozotocin-induced experimental rats.

Diabetes is a major noncommunicable life-threatening chronic and pervasive condition that is consuming the world health in a petrifying rate. The circulatory system is one of the major sources of hyperglycemia-induced ROS generation. Historically, garlic has been revered as part of a healthful diet. Organosulfur compounds have been attributed to the medicinal properties and health benefits of garlic. The present study focuses on the ameliorative role of allyl methyl sulfide (AMS) in combating diabetic complications in diabetic rats. Male Wistar rats were randomly divided into four groups. Experimental diabetes was induced by a single intraperitoneal injection (i.p), of streptozotocin (STZ) (40 mg/kg b.w). STZ treated diabetic rats showed significant augment in plasma glucose level, lipidperoxidative (LPO) markers, glycoprotein components (hexose, hexosamine, sialic acid, and fucose), and significant decline in plasma insulin level, nonenzymatic antioxidants and activities of antioxidant enzymes in the circulatory system and tissues. Further, periodic acid-Schiff (PAS) staining of hepatic and renal tissues revealed positive stain accumulation and Western blot investigation of glucose transporter 2 (GLUT 2) in pancreas of STZ-induced hyperglycemic rats. Dietary intervention with AMS (100 mg/kg b.w) for 30 days demonstrated significant protective effects on all the biochemical parameters studied. Besides, biochemical findings were corroborated by histological exertion and Western blot study. The findings of current investigations recommended that AMS can ameliorate the consequences of diabetes due to their antioxidant efficacy and can be used as a potential therapeutic approach. Further studies are warranted to explore the clinical application of AMS.

Activity of Thioallyl Compounds From Garlic Against Giardia duodenalis Trophozoites and in Experimental Giardiasis.

Fresh aqueous extracts (AGEs) and several thioallyl compounds (TACs) from garlic have an important antimicrobial activity that likely involves their interaction with exposed thiol groups at single aminoacids or target proteins. Since these groups are present in Giardia duodenalis trophozoites, in this work we evaluated the anti-giardial activity of AGE and several garlic's TACs. In vitro susceptibility assays showed that AGE affected trophozoite viability initially by a mechanism impairing cell integrity and oxidoreductase activities while diesterase activities were abrogated at higher AGE concentrations. The giardicidal activities of seven TACs were related to the molecular descriptor HOMO (Highest Occupied Molecular Orbital) energy and with their capacity to modify the -SH groups exposed in giardial proteins. Interestingly, the activity of several cysteine proteases in trophozoite lysates was inhibited by representative TACs as well as the cytopathic effect of the virulence factor giardipain-1. Of these, allicin showed the highest anti-giardial activity, the lower HOMO value, the highest thiol-modifying activity and the greatest inhibition of cysteine proteases. Allicin had a cytolytic mechanism in trophozoites with subsequent impairment of diesterase and oxidoreductase activities in a similar way to AGE. In addition, by electron microscopy a marked destruction of plasma membrane and endomembranes was observed in allicin-treated trophozoites while cytoskeletal elements were not affected. In further flow cytometry analyses pro-apoptotic effects of allicin concomitant to partial cell cycle arrest at G2 phase with the absence of oxidative stress were observed. In experimental infections of gerbils, the intragastric administration of AGE or allicin decreased parasite numbers and eliminated trophozoites in experimentally infected animals, respectively. These data suggest a potential use of TACs from garlic against G. duodenalis and in the treatment of giardiasis along with their additional benefits in the host's health.

Garlic Organosulfur Compounds Reduce Inflammation and Oxidative Stress during Dengue Virus Infection.

Dengue virus (DENV) is a mosquito-borne flavivirus that causes significant global human disease and mortality. One approach to develop treatments for DENV infection and the prevention of severe disease is through investigation of natural medicines. Inflammation plays both beneficial and harmful roles during DENV infection. Studies have proposed that the oxidative stress response may be one mechanism responsible for triggering inflammation during DENV infection. Thus, blocking the oxidative stress response could reduce inflammation and the development of severe disease. Garlic has been shown to both reduce inflammation and affect the oxidative stress response. Here, we show that the garlic active compounds diallyl disulfide (DADS), diallyl sulfide (DAS) and alliin reduced inflammation during DENV infection and show that this reduction is due to the effects on the oxidative stress response. These results suggest that garlic could be used as an alternative treatment for DENV infection and for the prevention of severe disease development.

Diallyl Sulfide and Its Role in Chronic Diseases Prevention.

Diallyl sulfide (C6H10S, DAS) is one of the novel natural organosulfur compounds, which is mostly obtained from the genus Allium plants. Numerous studies have revealed several unique properties of DAS in terms of its health-promoting effects. DAS has proved to be anticancer, antimicrobial, anti-angiogenic, and immunomodulatory like unique functions as demonstrated by the multiple investigations. Diallyl sulfide can also impede oxidative stress and chronic inflammation as suggested by the literature. Studies also explored that DAS could thwart the development of chronic diseases like cancer, neuronal, cardiovascular disease through modulating mechanistic pathways involved in pathogenesis. In this book chapter, we have attempted to give the comprehensive view on DAS about the physiochemical and biological properties, and its preventive role in chronic diseases with a mechanistic overview.

Comparative functional characterization of eugenol synthase from four different Ocimum species: Implications on eugenol accumulation.

Isoprenoids and phenylpropanoids are the major secondary metabolite constituents in Ocimum genus. Though enzymes from phenylpropanoid pathway have been characterized from few plants, limited information exists on how they modulate levels of secondary metabolites. Here, we performed phenylpropanoid profiling in different tissues from five Ocimum species, which revealed significant variations in secondary metabolites including eugenol, eugenol methyl ether, estragole and methyl cinnamate levels. Expression analysis of eugenol synthase (EGS) gene showed higher transcript levels especially in young leaves and inflorescence; and were positively correlated with eugenol contents. Additionally, transcript levels of coniferyl alcohol acyl transferase, a key enzyme diverting pool of substrate to phenylpropanoids, were in accordance with their abundance in respective species. In particular, eugenol methyl transferase expression positively correlated with higher levels of eugenol methyl ether in Ocimum tenuiflorum. Further, EGSs were functionally characterized from four Ocimum species varying in their eugenol contents. Kinetic and expression analyses indicated, higher enzyme turnover and transcripts levels, in species accumulating more eugenol. Moreover, biochemical and bioinformatics studies demonstrated that coniferyl acetate was the preferred substrate over coumaryl acetate when used, individually or together, in the enzyme assay. Overall, this study revealed the preliminary evidence for varied accumulation of eugenol and its abundance over chavicol in these Ocimum species. Current findings could potentially provide novel insights for metabolic modulations in medicinal and aromatic plants.

The redox-active drug metronidazole and thiol-depleting garlic compounds act synergistically in the protist parasite Spironucleus vortens.

Spironucleus vortens is a protozoan parasite associated with significant mortalities in the freshwater angelfish, Pterophyllum scalare. Control of this parasite is especially problematic due to restrictions on the use of the drug of choice, metronidazole (MTZ), on fish farms. Use of garlic (Allium sativum) is undergoing a renaissance following experimental validations of its antimicrobial efficiency. Ajoene ((E,Z)-4,5,9-trithiadodeca-1,6,11-triene 9-oxide), is a stable transformation product of allicin, the primary biologically active component of garlic. In the current study, an ajoene oil crude extract had a minimum inhibitory concentration (MIC) of 40µg/ml against S. vortens. GC-MS and NMR spectroscopy revealed this ajoene extract contained a mixture of the (E) and (Z)-ajoene isomers along with diallyl disulphide (DADS) and diallyl trisulphide (DATS). The only component of the ajoene crude oil found to substantially inhibit S. vortens growth by optical density monitoring (Bioscreen C Reader) was (Z)-ajoene (MIC 16µg/ml). Ajoene oil acted in synergy with MTZ in vitro, reducing the individual MIC of this drug (4µg/ml) by 16-fold, and that of ajoene oil by 200-fold with a fractional inhibitory concentration (FIC) index of 0.263. This synergistic interaction was confirmed in vivo. S. vortens-infected Pterophyllum scalare angelfish dosed orally with 0.5% (v/w) MTZ combined with 0.05% (v/w) ajoene displayed a significant reduction in faecal trophozoite count, whilst those fed on 0.5% MTZ flakes (half the recommended oral dose) alone did not. This study demonstrates for the first time the synergistic interaction between the synthetic drug MTZ and natural ajoene oil both in vitro and in vivo. Future work should evaluate the potential synergy of ajoene and MTZ against MTZ-resistant bacteria and protists.

Composition of the Essential Oil of Allium neapolitanum Cirillo Growing Wild in Sicily and its Activity on Microorganisms Affecting Historical Art Crafts.

Essential oil of the aerial parts of Allium neapolitanum Cirillo collected in Sicily were analyzed by gas-chromatography-flame-ionization detection and gas-chromatography-mass spectrometry. Nineteen compounds were identified in the oil and the main components were found to be (E)-chrysanthenyl acetate (28.1%), (Z)-chrysanthenyl acetate (23.8%), (E)-ß-farnesene (9.6%), dimethyl trisulfide (9.6%), camphor (7.4%), methyl allyl disulfide (6.8%) and 1-methyl-3-allyl trisulfide (5.8%). The essential oil showed good antimicrobial activity against 11 strains of test microorganisms, including several species infesting historical material.

Experimental study on the optimization of extraction process of garlic oil and its antibacterial effects.

BACKGROUND: Garlic oil which is the main active constituent of garlic has a wide range of pharmacological activities, and a broad antibacterial spectrum. It also has a strong anti-cancer activity, and can significantly inhibit a variety of tumors such as liver cancer, gastric cancer and colon cancer. The objective is to study the extraction process of garlic oil and its antibacterial effects. MATERIALS AND METHODS: CO2 Supercritical extraction was used to investigate the optimal processing conditions for garlic oil extraction; filter paper test and suspension dilution test were applied to determine the bacteriostatic action of garlic oil. RESULTS: In the CO2 supercritical extraction experiment, factors influencing the yield of garlic oil were: extraction pressure > extraction temperature > extraction time in descending order. Range analysis showed that the optimal experimental conditions for CO2 supercritical extraction of garlic oil were extraction pressure of 15 Mpa, temperature of 40 °C, and duration of 1 h. Different concentrations of garlic oil could all inhibit the growth of Staphylococcus aureus, Escherichia coli and Bacillus subtilis, suggesting that garlic oil has an antibacterial effect. CONCLUSION: The optimal experimental conditions for CO2 supercritical extraction of garlic oil were: extraction pressure of 15 Mpa, temperature of 40 °C, and duration of 1 h; garlic oil has an antibacterial effect.

Garlic-derived allyl sulfides in cancer therapy.

Garlic (Allium sativam L.) is widely used in traditional herbal remedies and alternative medicine. The potential health benefits of garlic are largely attributed to its metabolic byproducts. Extensive in vivo and in vitro studies has demonstrated that the garlic derivatives possess anti-cancer effects, but the underlying mechanisms are not completely understood. In this mini-review, we aim to summarize the reported biological effects of garlic products as anti-tumor agents, and present the possible molecular mechanisms responsible for the anti-carcinogenesis effects of garlic and its derivatives.

Investigating the responses of Cronobacter sakazakii to garlic-drived organosulfur compounds: a systematic study of pathogenic-bacterium injury by use of high-throughput whole-transcriptome sequencing and confocal micro-raman spectroscopy.

We present the results of a study using high-throughput whole-transcriptome sequencing (RNA-seq) and vibrational spectroscopy to characterize and fingerprint pathogenic-bacterium injury under conditions of unfavorable stress. Two garlic-derived organosulfur compounds were found to be highly effective antimicrobial compounds against Cronobacter sakazakii, a leading pathogen associated with invasive infection of infants and causing meningitis, necrotizing entercolitis, and bacteremia. RNA-seq shows changes in gene expression patterns and transcriptomic response, while confocal micro-Raman spectroscopy characterizes macromolecular changes in the bacterial cell resulting from this chemical stress. RNA-seq analyses showed that the bacterial response to ajoene differed from the response to diallyl sulfide. Specifically, ajoene caused downregulation of motility-related genes, while diallyl sulfide treatment caused an increased expression of cell wall synthesis genes. Confocal micro-Raman spectroscopy revealed that the two compounds appear to have the same phase I antimicrobial mechanism of binding to thiol-containing proteins/enzymes in bacterial cells generating a disulfide stretching band but different phase II antimicrobial mechanisms, showing alterations in the secondary structures of proteins in two different ways. Diallyl sulfide primarily altered the α-helix and ß-sheet, as reflected in changes in amide I, while ajoene altered the structures containing phenylalanine and tyrosine. Bayesian probability analysis validated the ability of principal component analysis to differentiate treated and control C. sakazakii cells. Scanning electron microscopy confirmed cell injury, showing significant morphological variations in cells following treatments by these two compounds. Findings from this study aid in the development of effective intervention strategies to reduce the risk of C. sakazakii contamination in the food production environment and on food contact surfaces, reducing the risks to susceptible consumers.

Cytotoxic effects of dillapiole on MDA-MB-231 cells involve the induction of apoptosis through the mitochondrial pathway by inducing an oxidative stress while altering the cytoskeleton network.

Breast cancer is the world's leading cause of death among women. This situation imposes an urgent development of more selective and less toxic agents. The use of natural molecular fingerprints as sources for new bioactive chemical entities has proven to be a quite promising and efficient method. Here, we have demonstrated for the first time that dillapiole has broad cytotoxic effects against a variety tumor cells. For instance, we found that it can act as a pro-oxidant compound through the induction of reactive oxygen species (ROS) release in MDA-MB-231 cells. We also demonstrated that dillapiole exhibits anti-proliferative properties, arresting cells at the G0/G1 phase and its antimigration effects can be associated with the disruption of actin filaments, which in turn can prevent tumor cell proliferation. Molecular modeling studies corroborated the biological findings and suggested that dillapiole may present a good pharmacokinetic profile, mainly because its hydrophobic character, which can facilitate its diffusion through tumor cell membranes. All these findings support the fact that dillapiole is a promising anticancer agent.

Dillapiole, isolated from Peperomia pellucida, shows gastroprotector activity against ethanol-induced gastric lesions in Wistar rats.

Peperomia pellucida is a plant used in traditional medicine to treat gastric ulcers. Although this gastroprotective activity was reported, the active compounds have not been identified. Therefore, the aim herein was to identify the most active compound in the gastroprotective activity of P. pellucida using an ethanol-induced gastric ulcer experimental rat model. A gastroprotective effect was observed when the hexane and dichloromethane extracts were tested, with the higher effect being obtained with the dichloromethane extract (82.3 ± 5.6%) at 100 mg/kg. Dillapiole was identified as the most active compound in this extract. Although there have been previous reports on dillapiole, this is the first on its gastroprotective activity. Rats treated with this compound at 3, 10, 30 and 100 mg/kg showed 23.1, 56.1, 73.2 and 85.5% gastroprotection, respectively. The effect elicited by dillapiole at 100 mg/kg was not attenuated by pretreatment with indomethacin (10 mg/kg, s.c.), a prostaglandin synthesis blocker, NG-nitro-l-arginine methyl ester (70 mg/kg, i.p.), a nitric oxide (NO) synthase inhibitor, or N-ethylmaleimide (10 mg/kg, s.c.), a blocker of sulfhydryl groups. This suggests that the gastroprotective mechanism of action of dillapiole does not involve prostaglandins, NO or sulfhydryl groups.

Cyclic sulfoxides garlicnins B2, B3, B4, C2, and C3 from Allium sativum.

Several novel sulfides, called garlicnins B2 (1), B3 (2), B4 (3), C2 (4), and C3 (5), were isolated from acetone extracts of garlic, Allium sativum L. and characterized. These garlicnins are capable of suppressing M2 macrophage activation and they have a novel skeleton of cyclic sulfoxide. The structures of the former 3 and latter of 2 were deduced to be 2-(sulfenic acid)-5-(allyl)-3,4-dimethyltetrahydrothiophene-S-oxides and 2-(allyldithiine)-5-(propenylsulfoxide)-3,4-dimethyltetrahydrothiophene-S-oxides, respectively. The mechanism of the proposed production of these compounds is discussed. The identification of these novel sulfoxides from garlic accumulates a great deal of new chemistry in the Allium sulfide field, and future pharmacological investigations of these compounds will aid the development of natural, healthy foods and anti-cancer agents that may prevent or combat disease.

Effect of Allium sativum and Allium cepa oils on different stages of Boophilus annulatus.

The study is aimed to investigate the acaricidal effect of Allium sativum (garlic) and Allium cepa (onion) oils on different stages of Boophilus annulatus hard tick. Engorged B. annulatus females were collected from naturally infected cattle. A number of engorged ticks were incubated at 28 °C and 85 % relative humidity to lay eggs, which were incubated to obtain larvae that were used in the study. The used garlic and onion oils were prepared by steam distillation and were analyzed by gas chromatography. These oils were dissolved in ethanol, methanol alcohols, and, partially, in water. The oils were tested in different concentrations; 1, 2, 5, 10, and 20 %. These concentrations were applied on adult ticks by adult immersion test; on larvae by larval immersion technique and on eggs. The 20, 10, and 5 % of garlic oil dilutions in ethanol and methanol killed all adult ticks and larvae within 24 h. Similar results were obtained for 10 and 20 % garlic oil dissolved in water. The effect of 10 % aqueous solution of garlic oil on embryonated eggs was clear as its addition to these eggs led to their in ability to hatch, deformity in shape, and change in color. The 10 and 20 % onion oil in ethanol and methanol alcohols killed 76-86 % of the adult ticks within 72 h post-application. While, all larvae died within 24 h postsubjected to these two concentrations. These concentrations (10 and 20 %) of onion oil in water killed 56-80 % of the treated ticks. Moreover, 10 % aqueous solution of onion oil prevented hatching of embyonated eggs. We concluded that garlic and onion oils have acaricidal effect on all stages of B. annulatus at concentrations higher than 5 %. Only garlic oil could kill 100 % of adult ticks at concentrations from 5 % in alcohols.

Allium sativum-derived allitridin inhibits Treg amplification in cytomegalovirus infection.

This study investigated the effects of allitridin compound on murine cytomegalovirus (MCMV)-induced regulatory T cell (Treg; CD4(+) CD25(+) Foxp3(+) ) amplification in vivo and in vitro. One hundred twenty MCMV-infected mice were allocated at random into two groups for treatment with allitridin or placebo. Another 120 mock-infected mice were randomly allocated as controls for the allitridin treatment and placebo treatment groups. The mice were euthanized at various time points after infection (out to 120 days) to evaluate the effects of treatment on Treg presence and function, as well as MCMV infective load. Co-culture with mouse embryo fibroblasts (MEF) and MCMV was performed to evaluate allitridin-mediated Treg and anti-CMV effects. The maximum tolerance concentration (MTC) of allitridin was used to treat cells for 3 days. Changes in Foxp3 mRNA and protein levels, percentages of T cell subsets, and Treg-related cytokines (IL-10 and TGF-ß) were measured. Allitridin treatment did not influence Foxp3 expression and Treg proportion in uninfected mice, but did down-regulate each in infected mice during the chronic infection period. Additionally, allitridin treatment reduced the MCMV load in salivary glands. MTC allitridin treatment of co-cultures partially blocked MCMV induction of Foxp3 mRNA and protein expression. In vitro treatment with allitridin also increased significantly the percentages of Tc1, Tc2, and Th1, reduced the secreted levels of IL-10 and TGF-ß1, and significantly suppressed viral loads. In conclusion, allitridin can promote MCMV-induced Treg expansion and Treg-mediated anti-MCMV immunosuppression. Therefore, allitridin may be useful as a therapeutic agent to enhance the specific cellular immune responses against CMV.

Effect of allyl sulfides from garlic essential oil on intracellular ca2+ levels in renal tubular cells.

Diallyl sulfide (1), diallyl disulfide (2), and diallyl trisulfide (3), which are major organosulfur compounds of garlic (Allium sativum), are recognized as a group of potential chemopreventive compounds. In this study, the early signaling effects of 3 were examined on Madin-Darby canine kidney (MDCK) cells loaded with the Ca(2+)-sensitive dye fura-2. It was found that 3 caused an immediate and sustained increase of [Ca(2+)](i) in a concentration-dependent manner (EC(50) = 40 µM). Compound 3 also induced a [Ca(2+)](i) elevation when extracellular Ca(2+) was removed, but the magnitude was reduced by 45%. In Ca(2+)-free medium, the 3-induced [Ca(2+)](i) level was abolished by depleting stored Ca(2+) with 1 µM thapsigargin (an endoplasmic reticulum Ca(2+) pump inhibitor). Elevation of [Ca(2+)](i) caused by 3 in the Ca(2+)-containing medium was not affected by modulation of protein kinase C activity. The 3-induced Ca(2+) influx was inhibited by nifedipine and nicardipine (1 µM). U73122, an inhibitor of phospholipase C, abolished ATP (but not the 3-induced [Ca(2+)](i) level). These findings suggest that 3 induced a significant [Ca(2+)](i) elevation in MDCK renal tubular cells by stimulating both extracellular Ca(2+) influx and thapsigargin-sensitive intracellular Ca(2+) release via as yet unidentified mechanisms. Furthermore, the order of the allyl sulfide-induced [Ca(2+)](i) elevation and cell viability was 1 < 2 < 3. The differential effect of allyl sulfides on Ca(2+) signaling and cell death appears to correlate with the number of sulfur atoms in the structure of these allyl sulfides.

In vitro efficacy of diallyl sulfides against the periodontopathogen Aggregatibacter actinomycetemcomitans.

The in vitro antibacterial effects of diallyl sulfide (DAS) against the Gram-negative periodontopathogen Aggregatibacter actinomycetemcomitans, the key etiologic agent of the severe form of localized aggressive periodontitis and other nonoral infections, were studied. A. actinomycetemcomitans was treated with garlic extract, allicin, or DAS, and the anti-A. actinomycetemcomitans effects of the treatment were evaluated. Garlic extract, allicin, and DAS significantly inhibited the growth of A. actinomycetemcomitans (greater than 3 log; P < 0.01) compared to control cells. Heat inactivation of the garlic extracts significantly reduced the protein concentration; however, the antimicrobial effect was retained. Purified proteins from garlic extract did not exhibit antimicrobial activity. Allicin lost all its antimicrobial effect when it was subjected to heat treatment, whereas DAS demonstrated an antimicrobial effect similar to that of the garlic extract, suggesting that the antimicrobial activity of garlic extract is mainly due to DAS. An A. actinomycetemcomitans biofilm-killing assay performed with DAS showed a significant reduction in biofilm cell numbers, as evidenced by both confocal microscopy and culture. Scanning electron microscopy (SEM) analysis of DAS-treated A. actinomycetemcomitans biofilms showed alterations of colony architecture indicating severe stress. Flow cytometry analysis of OBA9 cells did not demonstrate apoptosis or cell cycle arrest at therapeutic concentrations of DAS (0.01 and 0.1 µg/ml). DAS-treated A. actinomycetemcomitans cells demonstrated complete inhibition of glutathione (GSH) S-transferase (GST) activity. However, OBA9 cells, when exposed to DAS at similar concentrations, showed no significant differences in GST activity, suggesting that DAS-induced GST inhibition might be involved in A. actinomycetemcomitans cell death. These findings demonstrate that DAS exhibits significant antibacterial activity against A. actinomycetemcomitans and that this property might be utilized for exploring its therapeutic potential in treatment of A. actinomycetemcomitans-associated oral and nonoral infections.

Studies on purification of allicin by molecular distillation.

BACKGROUND: With recent improvements in living standards, people have been giving more consideration to the healthcare effects of foods. In this respect, allicin, which is the most important organosulfur compound in garlic and plays a key role in physiological function, has been receiving much attention. RESULTS: Allicin obtained from garlic by supercritical CO(2) extraction was purified by molecular distillation (MD). The effects of operating conditions such as absolute pressure (AP), distillation temperature (DT) and feed flow rate (FFR) on allicin purity and yield were studied. The optimal AP, DT and FFR levels in first-stage MD were found to be 200 Pa, 50 °C and 15 mL min(-1) respectively. After three stages of MD the contents of allicin, diallyl disulfide (DADS) and diallyl trisulfide (DATS) were 68.04, 9.19 and 5.91% (w/w) respectively. CONCLUSION: This study has provided a safe and effective method for the purification of allicin.