Recently, many biochemical and epidemiologic studies have revealed that polyphenols of various foods and herbs are beneficial to human health, and some extracts of polyphenolrich plants, such as green tea and grape seed, have been applied to functional foods or supplements. The potent biological activities of green tea polyphenols in, for example, cancer and cardiovascular disease prevention, 1) have in particular been attracting scientists in medicinal and pharmaceutical fields. Tea polyphenols were also shown to have antibacterial, 2-5) antiviral, 6) and antifungal activities 7) in addition to their inhibitory effects on exotoxins. 4,8) In these studies, only a few types of cultures such as from the American Type Culture Collection (ATCC) were examined according to different protocols.2,5,9,10) And therefore differences in susceptibility against the same species were indicated. 11)These differences were partly ascribable to the limited number of trial strains used in previous studies. Furthermore, in addition to tea polyphenols, other types of polyphenols, such as hydrolyzable and condensed tannins, are widely distributed in the plant kingdom and have the potential to be functional material. In the present study, we examined the antibacterial activities of 10 different plant polyphenols by comparing their minimum inhibitory concentrations (MICs) against a total of 96 strains of four groups of food-borne pathogenic bacteria including gram-positive and gram-negative species. Taking into consideration the future application of the results in the environmental and sanitation fields, we selected polyphenols that can be obtained in large amounts from plant material. MATERIALS AND METHODSChemicals Epigallocatechin (EGC, 1) and epigallocatechin-3-O-gallate (EGCg, 2) were isolated from commercial green tea, 12) punicalagin (3) was isolated from the peel of Punica granatum, 13) tannic acid (4) was purchased from Kanto Chemical Co. (Tokyo, Japan), castalagin (5) was isolated from wood of Castanea crenata, 14) and prodelphinidin oligomers (6) were isolated from the bark of Elaeocarpus sylvestris var. ellipticus; an aqueous acetone extract of the dried bark was separated by Sephadex LH-20 column chromatography with H 2 O containing increasing proportions of MeOH and finally with H 2 O-acetone (1 : 1, v/v). Prodelphinidin oligomers were then eluted out with 80-100% MeOH and aqueous acetone. Thiol degradation of 6 with mercaptoethanol-HCl 15) showed that this prodelphinidin is composed of EGC and EGCg, and the molecular weight of the peracetate was estimated to be 2000 daltons (peak top molecular weight) by gel permeation chromatography with a TSK-gel G4000H6 column (TOSOH) using polystylene as a standard. Geraniin (7) was isolated from the leaves of E. sylvestris var. ellipticus, 16) and loquat procyanidins (8) were isolated from the seeds of Eriobotrya japonica; fresh seeds were extracted with 80% acetone-H 2 O, and the extract was separated on a Diaion HP-20 column using H 2 O containing increasing amounts of MeOH, which ...

Recently, many biochemical and epidemiological studies have revealed that polyphenols of various foods and herbs have benefits to human health, and some extracts of polyphenol-rich plants, such as green tea and grape seed, have been added to foods or supplements. The antibacterial activity of various plant polyphenols and plant extracts have also been evaluated in several pharmaceutical studies. 1,2) Although tea polyphenols and their extracts have been examined in detail, [3][4][5][6] other extracts of polyphenol-rich plants have not been properly evaluated, because the activity of their chemical constituents has not been clearly demonstrated. 7,8) Many reports on the antibacterial activity of pure polyphenols have been published, 4,[9][10][11][12] however, each result was not able to be compared directly because different methods of evaluation were applied and various bacterial species were used. In order to overcome this problem, we compared the activity of a wide variety of polyphenols against many bacterial species to clarify their antibacterial spectrum, using the same standard method of the minimum inhibitory concentration (MIC). As a result, we found a relatively simple structure-activity relationship, and it was applied to the plant extracts containing various types of polyphenols. MATERIALS AND METHODSChemicals Twenty-two pure or partially pure polyphenols and 26 plant extracts were used in this study (Fig. 1). Polyphenols 1-10 were the same as in our previous study. 12)Epigallocatechin (EGC, 1) and epigallocatechin-3-O-gallate (EGCg, 2) were isolated from commercial green tea; punicalagin (3) was isolated from the peel of Punica granatum; tannic acid (4) was purchased from Kanto Chemical Co., Japan; castalagin (5) was isolated from the wood of Castanea crenata; and prodelphinidins (6) were isolated from the bark of Elaeocarpus sylvestris var. ellipticus; geraniin (7) was isolated from the leaves of E. sylvestris var. ellipticus; loquat procyanidins (8) were isolated from the seeds of Eriobotrya japonica; theaflavins (9) was obtained from black tea; and loquat-treated green tea polyphenols (10) were prepared by treatment of commercial green tea with unripe loquat fruit. Gallic acid monohydrate (11) was purchased from Wako Pure Chemical Industries, Japan. Thearubigin (12) was prepared from black tea as follows: aqueous acetone extract was successively partitioned with AcOEt and n-BuOH. The nBuOH layer was concentrated and subjected to Sephadex LH-20 column chromatography. Elution of 50% acetone yielded 12. (ϩ)-Catechin (13) and (Ϫ)-epicatechin gallate (ECg, 14) were isolated from gambir and green tea, respectively. Myricitrin (15) and rutin (16) were obtained from bark of Myrica rubra and flower bud of Sophora japonica, respectively. Theaflavin mixture (17) was separated from AcOEt soluble fractions of aqueous acetone extracts of black tea. Pyrocatechol (18), pyrogallol (19), protocatechuic acid (20), and caffeic acid (21) were purchased from Sigma Aldrich Japan. Resveratrol (22) was isolated from dried...

Legionella organisms are prevalent in manmade water systems and cause legionellosis in humans. A rapid detection method for viable Legionella cells combining ethidium monoazide (EMA) and PCR/real-time PCR was assessed. EMA could specifically intercalate and cleave the genomic DNA of heat-and chlorine-treated dead Legionella cells. The EMA-PCR assay clearly showed an amplified fragment specific for Legionella DNA from viable cells, but it could not do so for DNA from dead cells. The number of EMA-treated dead Legionella cells estimated by real-time PCR exhibited a 10 4 -to 10 5 -fold decrease compared to the number of dead Legionella cells without EMA treatment. Conversely, no significant difference in the numbers of EMA-treated and untreated viable Legionella cells was detected by the real-time PCR assay. The combined assay was also confirmed to be useful for specific detection of culturable Legionella cells from water samples obtained from spas. Therefore, the combined use of EMA and PCR/real-time PCR detects viable Legionella cells rapidly and specifically and may be useful in environmental surveillance for Legionella.