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Authors: Alex Dryden
Tags: #Fiction, #Thrillers, #Espionage
To 'J' and to the
Russians who want their freedom
'…On a huge hill, Cragg'd, and steep, Truth stands, and he that will Reach her, about must, and about must go'
John Donne
I DON'T KNOW WHO I'M writing this for but perhaps…
I MET FINN for the first time back in January…
NANA'S DACHA AT BARVIKHA had always been my home, in…
IT WAS NOT ONLY my father who was delighted when…
THE KRASNOZNAMENNIY INSTITUTE, or KI, was where women trained for…
DEAR RUSSIANS, very little time remains to a momentous date…
FINN GETS UP, walks over to the window, stretches, and…
WHEN FINN AND I had met a year before, at…
IN THE SPRING of the year 2000 Vladimir Putin moved…
IN THE AFTERGLOW of our mild hysteria, I walked alone…
WHEN I'D GOT OVER the shock of Patrushev actually appearing…
AS I SIT now in the vault at Tegernsee, I recall how…
AFTER FINN'S SHOWDOWN with the embassy's head of station, he…
THE HEAT OF BODIES in the gasthaus at Tegernsee was…
FINN PREFERS TO WALK. Even when he was in Moscow…
FINN WAS BACK in London on the first Eurostar train…
FINN RETURNS HOME that afternoon after his lunch with Adrian.
FOR TWO WEEKS Finn criss-crosses southern Europe, from the Russian…
ON A FINE CLEAR SUMMER MORNING, when the outlines of…
I WAKE ERRLY the next morning unable to sleep, the worst…
AT THE BEGINNING of September 2001, just after we lost…
AFTER MORE THAN A YEAR without contact between Finn and…
TWO THINGS HAPPENED in the following seven days that changed…
I MOVED INTO FINN'S LIFE and his apartment in Camden…
THERE ARE SEVEN OF US sitting in the Café des…
THE PHOTOGRAPHS LIE SPREAD OUT on a burgundy-coloured coverlet that…
ON THE MORNING of the meeting, it is innocently bright,…
THE DRESDEN FILE gives us five names,' Finn says. He…
THE PLANE LANDED at Scheremetyevo airport north of Moscow at…
I DON'T KNOW what time it was–sometime after midnight certainly–when…
FINN CAUGHT THE TRAIN to Frankfurt, with or with the…
I RETURNED TO LONDON two days after Vladimir had taken…
AT THE END OF APRIL, Finn told me we were…
FINN AND I FLEW to Bucharest one early morning in…
I WOKE UP IN BED. I didn't know where I…
FINN'S EXCITEMENT is so naive. The almost boyish enthusiasm he…
IT IS JUST MORE THAN a week later. Finn and…
FINN SAID HE WOULD CALL twice a day until he…
THERE IS ONLY one photograph of Mikhail in the public…
writing this for but perhaps it's for you. If that makes it sound like a confession, you may wonder what I'm expecting in return. A small part of me, I admit, seeks forgiveness, or at least understanding. But that part of me is less important than the forgiveness I wish to give myself, and which I find elusive.
I am writing to draw a line under the past, with its rot creeping into the present. I know now that if I had done this a long time ago, the present would never have been postponed and things would be different today.
In one of his more fatalistic moments, Finn said to me: 'Anna, you know our story can never be written.'
'Why not?' I asked him.
'Nobody would believe it,' he said.
But I'm here now, sitting in a medieval vault in a house in Tegernsee on the southern borders of Germany, reading Finn's story-our story- and I'm aware that all I have between me and the hostile forces that swim up at me from the pages is the Contender
handgun and the twelve rifle shells on the table by my hand. And now that I've found these notebooks of his, or books of record, as he calls them, buried in this vault along with all the other material of our secret profession, I see his fatalism was short-lived. As I sift through the piles of notebooks, oddments, scraps and sheets of paper, documents and microfiches-their edges stained with cellar dampness-with only the heat of an oil burner to keep me warm, I can see that he has practically written our story himself.
The notebooks certainly contain the facts and, without these facts, my feelings would be drifting in a vacuum, unmoored to the reality that at any moment I may need to use this gun and all my years of training to kill my way out of here. Feelings need to be clothed in reality and the facts–this story–supply the clothes. For days now, I have been reading and rereading Finn's prose, notes and observations—over and over. I'm reading them sitting in this dark stone vault and my eyes are running from the fumes of the oil burner and I strain in the dim light to follow the thread of a story that began long before I met Finn.
According to one note Finn made, our story begins in 1998, when Boris Yeltsin's Russia reached its nadir. A later scrawl in Finn's undisciplined handwriting names 1989 as the beginning, the year the Berlin Wall came down at last. But another, perhaps more thoughtful, observation says that it all started in 1961, when we Russians erected the Wall in the first place.
Whatever the true beginning, however, everything Finn and I have experienced will continue to unfold into a dark and uncertain future, with or without us.
As I read all his disparate and complementary records, the thing that strikes me most deeply about what Finn experienced in his long quest to have the truth accepted in his own country is his sheer obstinacy, the relentless autopilot of individual human endeavour when success seems impossible.
What also strikes me is that Finn's past has dictated his life, much as my past has dictated mine.
'You cannot escape your past, Anna,' he once told me. 'But you don't have to live in it. You don't have to build the present in its image.'
If only Finn had been true to his own belief.
Finn could have had a quiet life. That is the point, I realise, as I sit here shivering in the damp cold. He told me that he chose to pursue this quest, not just for the truth, but to have the truth accepted by his masters in London, and their political masters in the British Government. But did he really choose? Or was it his deep-seated need for acceptance that fed his stubbornness and single-mindedness?
For my part, I know I'm looking for someone or something to find responsible for my own actions, but I can't escape my part.
Oh yes, Finn could have had a quiet life, a beautiful life. He had a great talent for doing nothing, which he called happiness, but he chose to go alone down the Tunnel, as he calls it here, and I hope I'm not deluding myself when I say he would not regret that now, whatever's happened to him. For Finn has disappeared and, as I wait for the crash of sledgehammers against the door upstairs, I'm looking for a clue to tell me something, anything that might help me to find him.
There is much, too, about Finn himself in these notebooks which distracts me from my increasingly urgent task. There are details of his internal struggle to understand his motives, a struggle which I never fully understood, and that he never told me, despite the fusion of our love. During all the time I've known Finn, he never wanted to bring his own past like an evil spirit into our house. So he wrote it down in the notebooks and buried it with our secret story in this vault, which has hidden many things and many people in its long history.
And there is much in the notebooks about his feelings towards me.
'There are three distinct spirits in our relationship,' Finn once said to my grandmother at the dacha in Barvikha. This was back in the freezing winter weeks leading up to the millennium, when perhaps he and I were at our closest, and when trouble seemed far away. 'There's Anna, me and the spirit that joins us.' My grandmother, with her peasant background, was comfortable with the world of spirits. She laughed with mirth and hugged him. Like many people whose lives he touched, Nana loved Finn.
Finn had just given me a charm bracelet. It contained two charms: a rabbit for me and a monkey for him. It was his nickname for me, Rabbit.
'The silver circle that links the charms,' he said, 'is the spirit that joins us.'
'I'm not sure I believe in spirits,' I said.
'I find I can't do without them,' he said breezily.
'How sentimental,' I replied.
But it was typical of Finn instinctively to sense the language, the context, of whoever he was talking to. Nana appreciated his inclusion of spirits. Nana was very superstitious and, my mother told me, she was also psychic. With her deep grey eyes that glinted over her sharp, hooked nose, she even looked witch-like. When Finn described his relationship with me in this way, she replied as if foreseeing the future.
'If only that were true,' she said. 'If only you, Anna and the spirit that joins you both really were the only three things in your relationship. God bless you.'
At this moment, in the cellar, I pause over another scrap of paper he wrote about us, on some Luxembourg hotel notepad. I'm mes-merised by what he's written and can almost feel his presence here, through the words.
'When we make love, and we look into each other's eyes, I see
the child in you, Anna, the spirit in you, and in those moments you are me and I am you.'
It is these brief, aching glimpses of our intimacy that distract me from my task and, to keep my mind away from such thoughts and focused on the present danger, I sit and listen for the slightest sound. Then I slot the gun's firing pin into the mechanism and slide a single round of green spot ammunition into the chamber. With this weapon, I can kill a man at over two hundred yards.
It is hard enough to keep love safe. In my own experience it is prone either to snap suddenly, or to dwindle and fade to a monotonous daily exercise that is not so different from filling the dishwasher or taking the dog for a walk. I have passionately kissed a man at the start of a film while out on a date at the cinema and, by the end, found him physically repulsive to touch. I have also watched other affairs dwindle imperceptibly to the banal, the everyday, the meaningless.
But neither of these fault lines is true in the case of Finn and myself. We have, or is it had? other troubles. Whatever the truth of our love, we allowed too much to come between us. And the past sneaked into any available crack and fissure in our relationship as frost enters the cement courses of a house.
We had the malevolence of others, with their huge forces arrayed against us, but it was we, ultimately, who allowed their hostility to disrupt the peace we might have found.
We had microphones, watchers, tails, people in every corner. We had two of the world's most experienced intelligence services tracking our every movement. We had shining slivers of time to snatch for last minute walk-ins in hotel rooms across the city. At reception desks, Finn would call me his wife.
'I'm not your wife,' I told him at the beginning. 'I'm a colonel in the SVR. Doing a job, that's all.'
'OK. Colonel,' he might say. 'Allow me to undo the buttons of your shirt.'
'Please, then,' he'd say.
Do I mean that we had the malevolence of others, or do we still have it? Where is Finn? What has happened to him? Is Finn now the past or is he still in the present?
for the first time back in January 1999. Our meeting had been at the Baltschug Hotel, across the Moskva River from the Kremlin. The hotel was to remain a place of good memories for us throughout Finn's residency in Moscow. As he put it, rather unnecessarily, 'It was the venue for our first official fuck, Rabbit, endorsed by Her Majesty and the KGB.'
Finn was sitting at the head of the table in a private meeting room at the Baltschug, in his role of Second Secretary of Trade and Investment. He was holding court with a dozen or so business and security acolytes and their boss Pavel Drachevsky, the billionaire aluminium tycoon. Drachevsky had invited some of his managers: his leaden ex-KGB security boss Chimkov; Alexander, the poet, translator and KGB snake who had worked at the United Nations in Geneva for several years; and my friend Natasha, who tried to act in Russia's confused public relations world as Drachevsky's PR assistant.
Whether Finn knew the lunch was a set-up or not, he didn't show it. When I arrived, deliberately late, he looked completely
relaxed. He was sitting in a high-backed chair at the head of the long table. He wore a slightly creased blue suit and a tie, which could have been tied by a schoolboy, with a large thick knot and one very short end. He had brown hair, brushed straight back over his head and greying slightly at the temples, and his face was lined from laughter. His eyes, which changed through various shades of green and brown, depending on the light, as I discovered later, contained a kind of merry amusement that stopped just short of scorn. Finn always seemed to be enjoying himself.
I noticed that he addressed more of his conversation to Natasha, who is very pretty, than to Drachevsky, who was the reason Finn was there and who can't claim good looks as his forte.
Whether or not Finn knew who I was, I couldn't tell, but as I entered the room dressed in black leather–it embarrasses me now to recall this–I immediately had his attention. Of course I did. That was the idea. Finn's reputation for womanising in Moscow was always considered to be the hook to catch him with, despite our consistent failures up till now, and in the crude logic of the KGB it was considered that Finn would be more of a pushover if I, the youngest female KGB colonel, wore black leather.
I took my seat next to Natasha so that Finn would now look at both of us, and the conversation continued in a relaxed and businesslike way.
And then, at the appointed moment, Natasha looked up the table towards Finn. It was all rehearsed. In her lazy seducer's voice which she and I would practise to our increasing amusement in her apartment over a bottle of wine, she came out with the line we'd told her to say.
'Tell me, Finn, are you a spy?'
This was the signal for the whole table to stop talking and look directly at Finn. I was supposed to read his face.
There was dead silence. It was a foolish, old-fashioned Cold War moment.
But all I saw in Finn's eyes was his continued relaxed amusement. And then, startlingly, he put up his hands like a pickpocket caught in the act.
'How did you guess?' he said.
It was brilliantly done.
It was certainly not what anyone had expected. It threw the business officials at the party into barely concealed consternation. One or two of Drachevsky's managers looked around, breaking the injunction to stay staring at Finn, as if they were personal witnesses to a great diplomatic scandal. 'The Second Secretary of Trade and Investment at the British embassy in Moscow admits to being a spy.' These managers were almost there, at the interview with ORT News, telling the story of the headline with knitted brows and the melodramatic seriousness beloved by TV interviewees.
But I saw immediately that to look at Finn was not to look at a man who appeared to have confessed to anything at all.
At least silence was maintained in the confusion and the rest of us stayed staring at Finn, hoping, I suppose, to unsettle him. Finn, however, treated us like an audience he had spent hours trying to win. This was his moment of triumph, not defeat, he seemed to say.
'I'm double O zero,' he said, and smiled that open, guileless smile that still makes my stomach tighten when I think of it.
There were a few unrestrained grins around the table; Natasha actually laughed out loud. The brain-dead Chimkov and the snake Alexander kept their blank, trained expressions in place. Drachevsky just looked mildly curious. And as for me, I could not help smiling but I resisted aiming it at Finn and looked down at my plate.
The tension, which we thought we would create, but which Finn had effortlessly usurped, was broken. I suspected then that I had been promoted beyond my ability. I was charmed by him.
Finn pressed home his advantage, supremely confident, every inch the willing entertainer. 'I do know some spies,' he said, as if to
compensate for the drudgery of his trade job, and again he rocked those managers who, a moment before, had thought they were at a crucial moment in contemporary history.
'What, here? In Moscow!' one of them said, outraged.
'No,' Finn said, apologetic now that he might have led them to expect too much. His face said how embarrassed he was that he didn't know any spies actually in Moscow. 'No, they're retired now. I only know them socially, I'm afraid.'
Later in the day, Finn called me and asked me to dinner, just as my controllers at the Forest had expected. I accepted, of course. It was my job, after all, but it was easier than I'd feared.
We went to a sushi restaurant which had recently opened up near the church where Pushkin got married; Finn said it was the most expensive dinner he'd ever bought anyone. He'd booked our first room at the Baltschug that night because, as he made clear, he had a discount. I remember I was annoyed by his deliberately aimed presumption. But already he knew I had my orders-we both had our orders-and he decided to make the most of it.
I remember that evening clearly and things Finn said still play over in my mind. He was incredibly indiscreet. He made no attempt to pretend we were anything other than what we were. Looking me in the eye, he said, 'There wouldn't be any need for spies if there weren't any spies, Anna.' And later, in bed, he said, 'You're completely wasted as a colonel, you know. You're the best honey trap there ever was.'
He knew just how to get a rise from me at that first meeting.
One of the reasons why we'd always failed to trap Finn with the spectacular list of professionals we'd lined up for him was because he was so amused by the idea of honey traps. He'd talk to them for hours in order to waste our time, just as some men want only to talk to prostitutes. In this as in other things, Finn was quite childish
and the girls we threw at him were a source of hilarity and disdain for him.
Afterwards, when we'd made love, he said, 'You do realise I'm going to have to use you. I can't kick the youngest female colonel in the KGB out of bed. I'm going to have to sleep with you a lot, if that's OK.'
'Keep up,' I said. We're not the KGB any more. 'We call it the FSB these days.'
But Finn always called us the KGB to the end. 'It's the same thing, isn't it,' he said. 'It's just an old dog with a new name. There's no difference, except that now you've all been to business school as well as Honey Trap High.'
'Are you trying to ruin the evening?' I said.
He turned to me and it was the first time I saw him not play-acting. 'No,' he said. 'No, I don't want to ruin anything.'
We made love again, and afterwards it felt so strange, like we were two lovers who'd known each other for years.
'We've met before, I know it,' Finn said.
'And in what incarnation would that be?' I replied.
He held my hands above my head. 'No incarnation. You were pure dust and so was I.'
I undid his fingers. 'Why don't you do something to jog my memory?' I murmured. But he'd fallen asleep.
Anyway, that's how I first met Finn.


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subset: commoncrawl


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text: Plastitalia has been awarded by a brand new certification from Bureau Veritas (Italy) UNI 11024: Quality requirements for welding of polyethylene pipes for gas, water and other pressure fluids pipelines. This standard sets rules for organizations involved in the welding of plastics pipeline and/or components (e.g.: fabricated bends, etc.). The observation of the UNI 11024 requirements is a guarantee for the market that the organization has a documented system management for the whole welding processes. For a company like Plastitalia this process starts with the customer's request and go on with all stages, eg.: purchasing of the right material and its control, training of internal welders and people, checks of tools and equipment and testing of the item manufactured on the base of an internal sampling plan. All of this is made to provide the market with products that, even made as an handcraft, are under control at the same level of an industrial product.


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H.R.3463 - Ukrainian American Veterans Charter Act106th Congress (1999-2000)
BillHide Overview
|Rep. Bonior, David E. [D-MI-10] (Introduced 11/18/1999)
|House - Judiciary
|Latest Action:
|House - 12/01/1999 Referred to the Subcommittee on Immigration and Claims. (All Actions)
This bill has the status Introduced
Here are the steps for Status of Legislation:
- Introduced
Subject — Policy Area:
- Government Operations and Politics
- View subjects
Summary: H.R.3463 — 106th Congress (1999-2000)All Information (Except Text)
There is one summary for H.R.3463. Bill summaries are authored by CRS.


text: Host Tom Snyder's guests are Pauline Kael, film critic for New Yorker Magazine discusses her work. She claims that she doesn't think of what she does as a profession because she loves it so much. Kael claims the movie companies would be making better movies but that theres so much money in poor or mediocre ones that they don't bother. Jon Alpert, video journalist discusses his work in Cambodia, Iran and Afghanistan. He got to speak to the student militants when he went to Iran and contends Iranians are more interested in eating every day than in the United States. Next he'll go to Cuba. Clip shown from his new documentary, "Third Avenue: Only The Strong Survive", about male prostitutes.


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Testing of Hypotheses
The investigation of a bi-variate relationship is a vital step in explaining and testing the research hypothesis. A relationship of the two variables means that the distributions of values of the two variables are associated. In other words, the variation explained by one variable is patterned in such a manner that its variance is not randomly distributed in connection with other variables. In the context of a bi-variate relationship, the problem arises whether a relationship is real or has arisen by chance.
A bi-variate table is one in which two variables have been cross-classified. It consists of rows and columns. The categories of one variable are labeled for the rows and the categories of the second variable are labeled for the columns (Nachmias and Nachmias, 1992). Generally, significance of the relationship is examined by establishing a null hypothesis, in which it is assumed that there is no difference or no relationship between tow variables.
Table 7 presents the association between respondents' opinion about training and development with employees' conflict. Chi-square (5.95) shows a non-significant association between respondents' opinion about training & development with employees' conflict.


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(Reuters)--Technology and financing company Comdisco Inc., which is in the midst of a bankruptcy reorganization, on Friday reported a fiscal fourth-quarter loss from continuing operations as revenues fell sharply.
Comdisco posts $119M loss in 4Q
The Rosemont-based company also said it is not inclined to accept any of the bids it has received for its flagship information technology leasing business and may scrap plans to sell the unit.
Comdisco, which filed for Chapter 11 bankruptcy in July, said it lost $119 million, or 79 cents a share, in the three months ended Sept. 30, compared with a year-earlier profit of $62 million, or 39 cents a share. The results exclude the technology disaster recovery business that the company sold to SunGard on Nov. 15 for $825 million.
Comdisco said its net loss, which includes that business, narrowed to $142 million, or 94 cents per share, from $168 million, or $1.05 per share, a year earlier. Revenues fell 44 percent to $509 million from $901 million.
Comdisco attributed the quarterly loss to credit losses, lower contribution from its computer equipment remarketing efforts, and lower equity-related gains from its ventures business.
Comdisco had been taking bids for its leasing unit, but said it was "presently not inclined" to accept any offers received during the auction process, which concluded in November. A group of employees led by former Chairman Jack Slevin in October said they would bid for the leasing business.
Comdisco said in a press release it is issuing a "final opportunity" for bidders to submit sealed offers by Jan. 7.
However, "greater value may be achieved through reorganization rather than sale of the Leasing business," the company added.
Comdisco said the U.S. bankruptcy court in the northern district of Illinois had postponed to Jan. 24 the sale hearing for certain of its leasing businesses, previously scheduled for Dec. 20.
The company's Chapter 11 filing in July sparked a sell-off of some of its operating units. At the time, the company said it had agreed to sell its disaster recovery business to Hewlett-Packard for $610 million.
That offer touched off something of a bidding war for the unit, which attracted more attention after the Sept. 11 attack on the World Trade Center left many companies in need of disaster services for their computer systems.
Comdisco eventually accepted the $825 million cash offer from SunGard.


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AS THEY WERE: Muzzy Pep, pictured in 2003, are Nick Munnings, Scott Blackley, Errol Moyle and Luke Bennett.THEY were the band that could not be bothered breaking-up. It's now about seven years sinceMuzzy Pep playedtheir last gig and they say they cansee little pointin breaking up now.
So instead Muzzy Pep is getting back together –not that they actually disbanded, obviously.
"By the end of 2003 we had decided to take it slow, our most successful decision to date," bass player Nic Munnings explained.
"Save for a handful of shows to celebrate the Global Financial Crisis of 2008, we have taken it so slowly that most people assumed that the band had broken up."
Learning the old material involved way "too much effort", Munnings said.So to kick things off again they have written a whole lot of new songs, andthere is a new recording to boot.
The Muzzy Pep line-up also includes Scott Blackley on vocals and guitar, Errol Moyle also on vocals and guitar and Luke Bennett on drums.
Munningssaid the comebackwas a matter of "unfinished business". Despite releasing two EPs and two full-length recordings the bandfelt it had never captured its sound on a recording.
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It was recorded in Sydney's Linear Recording Studios with Nick Franklin at the helm. The band saidit nails their sound.
"We recorded all the basic tracks in seven hours, one shot, we knocked everything over," Munnings said. "We went back and added a few sparkly bits …but there is no sneaky edits, what you hear on the record is what went down. It sounds like we sound," he said.
Muzzy Pep will launch Cyclic on June 10 at The Stag and Hunter Hotel, Mayfield.


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Phytochemical analysis of Andrographis paniculata and Orthosiphon stamineus leaf extracts for their antibacterial and antioxidant potentialMalahubban, M.1,4, Alimon, A.R.1,2*, Sazili, A.Q.2, Fakurazi, S.3 and Zakry, F.A.51Laboratory
of Animal Production, Institute of Tropical Agriculture, of Animal Science, Faculty of Agriculture, 3Department of Human Anatomy, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia 4Department of Animal Science and Fishery, 5Department of Crop Science, Faculty of Agriculture and Food Sciences, Universiti Putra Malaysia Bintulu Campus Sarawak, 97008 Bintulu, Sarawak, Malaysia *Corresponding author email: email@example.com / firstname.lastname@example.com Received 27 March 2013; received in revised form 13 May 2013; accepted 28 May 20132Department
Abstract. Leaves of Andrographis paniculata and Orthosiphon stamineus were extracted with water, ethanol, methanol and chloroform to assess their potential as antibacterial and antioxidant agents. High performance liquid chromatography analysis showed that the methanolic extracts of A. paniculata and O. stamineus leaves gave the highest amounts of andrographolide and rosmarinic acid, respectively. These leaf extracts exhibited antimicrobial and antioxidant activities and, at the highest concentration tested (200 mg/mL), showed greater inhibitory effects against the Gram positive bacteria Bacillus cereus and Staphylococcus aureus than 10 % acetic acid. Andrographis paniculata and O. stamineus methanolic and ethanolic leaf extracts also showed the strongest antioxidant activity as compared with the other extracts tested. The bioactive compounds present in these leaf extracts have the potential to be developed into natural antibacterial and antioxidant agents that may have applications in animal and human health.
INTRODUCTION Since ancient times, medicinal plants have been commonly used to treat diseases and to maintain health. However, the medicinal attributes of such plants have not been welldocumented scientifically. Nevertheless, local medicinal plants such as Andrographis paniculata and Orthosiphon stamineus are used traditionally for healing purposes, and have been commercialized recently. Several medicinal plants, including A. paniculata and O. stamineus, have been shown to exhibit antimicrobial and anti-oxidative characteristics due to the presence of various phytochemicals such as flavanoids, phenolic diterpenes, phenolic acids, tannins, carotenoids and tocopherols (Akowuah et al.,
2004; 2005; 2006; Ebrahimabadi et al., 2010; Roy et al., 2010; Edziri et al., 2011; Zargar et al., 2011). In recent years, there have been many reports on the antibacterial and antioxidant potential of several hundreds of native Malaysian medicinal plants, highlighting their potential to be used to treat specific diseases and promote good health (Dung et al., 2008; Ebrahimabadi et al., 2010; Edziri et al., 2011). Medicinal plants have also been found to play an important role in the development of agro-chemicals to improve crop production (Adebowale & Adedire, 2006; Bajpai et al. , 2007; Acda, 2009). Moreover, the frequent use of synthetic and commercial antimicrobial drugs in treatments has led to drug resistance in animal, human and plant pathogens (Bajpai
et al., 2007). The excessive use of synthetic preservative agents and antibiotics such as chloramphenicol in meat products has led to human health concerns. Lipid peroxidation in food lowers the nutritive value of food and causes the flavor and taste of food to deteriorate. Besides that, it raises health issues as it contributes to aging, heart disease, stroke, emphysema, mutagenesis and carcinogenesis (Barlow, 1990; Bera et al., 2006). Medicinal plants showing antimicrobial and antioxidant activities may have the potential to act as safer substitutes because their molecular structures are different from those of microbes and chemical-based pharmaceuticals; therefore, their mode of action is likely to be different (Bera et al., 2006). Moreover, it is necessary to suppress lipid peroxidation to ensure that the food is safe for consumption and its flavour maintained (Bera et al., 2006). Andrographis paniculata Nees (Acanthaceae), known in Malaysia as Hempedu Bumi, is used traditionally as a remedy for fever. The aerial parts of the plant are commonly used to treat cold, hypertension, diabetes, cancer, malaria and snake bite (Perry, 1980). Andrographolides are active components found in the aerial part of this plant. Akowuah et al. (2006) successfully identified and quantified two major bioactive compounds in A. paniculata, namely andrographolide and 14-deoxy-11, 12-didehydroandrographolide. Andrographolide and 14-deoxy-11, 12-didehydroandrographolide have been characterized as bitter and colourless crystalline diterpene lactone, and as colourless needles, respectively. Orthosiphon stamineus Benth (Lamiaceae) is used widely in Southeast Asia for the treatment of eruptive fever, epilepsy, gallstone, hepatitis, rheumatism, hypertension, syphilis and renal calculus. Known locally as Misai Kucing in Malaysia, the plant preparation is commercially available in the form of dried leaves. It is commonly taken as a beverage to improve health and for the treatment for bladder inflammation, gout and diabetes. Orthosiphon stamineus contains several chemically active 468
constituents, including those belonging to the phenolic group, of which twenty have been identified. Among them are sinensetin, eupatorin and rosmarinic acid (Akowuah et al., 2004). Numerous studies on antioxidant properties of A. paniculata and O. stamineus have been conducted using different assay methods (Akowuah et al. , 2004; 2006; Premanath & Devi, 2011). Some reported variations in the findings may be ascribed to different environmental conditions and sample sourcing. In this study, the plants selected were morphologically similar although the chemical composition of the leaves might have been affected by soil fertility, age and genetic variation. The main objective of this study was to determine the antibacterial and antioxidant potential of A. paniculata and O. stamineus leaf extracts, and to quantify andrographolide and rosmarinic acid present in the respective species.
MATERIALS AND METHODS Chemicals HPLC grade methanol, acetonitrile (ACN) and acetone (Merck) were used. Andrographolide and rosmarinic acid (Chroma Dex, USA) were used as reference standards. Ammonical chloroform, chloroform, sulphuric acid (H2SO4), methanol, ethyl ether, hydrochloric acid (HCL), magnesium ribbon, ethanol, ferric chloride, acetic anhydride, acetonitrile, phosphoric acid, 0.5 Follinciocalteu, 2, 2-diphenyl-1-picrylhydrazyl (DPPH), ascorbic acid, nitroblue tetrazolium, nicotinamide adenine dinucleotide, tris-HCl buffer, phenazine methosulphate, nutrient broth (OXOID, U.K.), muller hinton agar (MHA) and tetracycline (OXOID, U.K.). Plant materials Fresh samples of cultivated A. paniculata and O. stamineus were obtained from the Herbal Farm in Universiti Putra Malaysia. Identities of the plant samples were authenticated by the Gene Bank Centre, Faculty of Agriculture, Universiti Putra Malaysia. The leaves of plants were collected
and oven-dried at 60C for 72 h. The dried plants were ground to powder using a Willey mill (Thomas Willey cutting mill model 4), sieved through a 1 mm screen and stored at 4C until further use. Sample preparation Powdered A. paniculata and O. stamineus leaves (50 g) were added to 500 mL of water, methanol, ethanol or chloroform in a water bath (Memmert, Germany) at 40C for 8 h to allow liquid adsorption before transferring to an incubator shaker (Protech SI-50D) at 40C for another 16 h. The extracts were then filtered (Whatman No. 1) then centrifuged at 3500 (KUBOTA 5100, Japan) for 10 min. The supernatants were then filtered under vacuum through filter paper (Whatman No. 1). The filtrates were evaporated using a rotary evaporator (Buchi B-491) at 40C. The concentrated extracts were transferred to glass petri dishes and dried in a drying oven at 40C. Qualitative screening of bioactive compounds Alkaloids test The test for alkaloids was carried out on 5 g ground plant material that had been extracted with 10 mL ammonical chloroform and 5 mL chloroform. After filtration, the supernatant was shaken with drops of 0.5 M sulphuric acid. The appearance of a creamy precipitate indicated the presence of alkaloid. Saponin test The test for saponin was carried out by adding 15 mL methanol to 5 g of the powdered plant extract. After evaporation, the residue was shaken vigorously with ethyl ether and 5 mL 2N HCL. The appearance of a precipitate indicated the presence of saponin. Flavanoids test The alcoholic extract (15 mL) corresponding to 3 g of plant material was treated with a few drops of concentrated HCL and 0.5 g magnesium ribbon. The appearance of a pink-red color indicated the presence of flavanoids.
Tannins test One gram of the ground plant sample was boiled in 20 mL of 70 % ethanol for 2 min. The mixture was filtered and a portion of the filtrate was diluted with sterile distilled water in a ratio of 1:4. Three drops of 10% ferric chloride solution was then added. The appearance of a blue to black precipitate indicated the presence of tannins. Steroids and terpenoids tests Steroids and terpenoids were detected using the Liebermann-Burchard reaction. Plant material (200 mg) was boiled in chloroform before being filtered. The filtrate (2 mL) was added to 2 mL acetic anhydride and 50% concentrated H 2 SO 4 . A blue-green ring indicated the presence of steroids and red colour indicated the presence of terpenoids. High Performance Liquid Chromatography (HPLC) analyses Preparation of sample and standard solution Andrographis paniculata and O. stamineus leaf extracts were analyzed by HPLC equipped with a photodiode array detector. Five mg of powdered extract was dissolved in 1 mL ethanol (for the ethanolic extract), or chloroform (for the chloroform extract), or methanol (for the methanolic extract), or distilled water (for the water extract), and passed through a syringe filter of 0.45 m cutoff. Stock solutions of andrographolide and rosmarinic acid (1 mg/mL) were prepared in methanol for each standard. The two analytical standards were further diluted to a concentration of 120 ppm. A compound standard solution prepared by mixing 250 L of each standard diluted solution was analyzed by HPLC for calibration and retention time reference. The test extracts were identified by comparing the retention times and UV spectra with those of the standards. A series of dilution were made to obtain concentrations ranging from 10-120 ppm. Sample injections were made in triplicate. The amount of andrographolide and rosmarinic acid in the standardized extract was calculated based on the peak area of the
chromatogram of the extract with that of the calibration standard. Linear regression plots were obtained using the Waters Empower 2 software. The results were expressed as mg andrographolide or rosmarinic acid per gram of A. paniculata and O. stamineus powdered extract. Chromatographic conditions Qualitative and quantitative determinations of the major constituents of A. paniculata and O. stamineus leaf extracts were performed on a Waters Delta 600 HPLC System with a Model 600 controller (Waters MA, USA). The system was equipped with a 996 photodiode array detector connected to a computer running Waters Empower 2 software. An AC18 column (Phenomenex, Luna 5 m, 250 x 4.6mm, i.d) guarded by a C-18 security guard cartridge (4 x 3.0 mm, i.d), maintained at room temperature was used as the stationary phase. Andrographolide in samples and in the reference standard was separated by a mobile phase comprising solvent A: 50% water, solvent B: 20% acetonitrile, solvent C: 30% methanol. Standards and samples of rosmarinic acid were separated by a mobile phase comprising solvent A: 0.1% aqueous phosphoric acid and solvent B: acetonitrile. The initial conditions were 85% A and 15% B, with a linear gradient reaching 25% B at t = 12 min. This was maintained for 10 min after which the program returned to the initial solvent composition at t = 25 min, and continued thereafter for another 10 min. The flow rate was 1.0 ml/min and the injection volumes for samples and standards were 10 L. Chromatographic peaks were detected at 230 nm for andrographolide and at 340 nm for rosmarinic acid. Antibacterial potential of Andrographis paniculata and Orthosiphon stamineus leaf extracts prepared with different solvents Test bacteria Two Gram-positive bacteria, viz. Bacillus cereus and Staphylococcus aureus, and two Gram-negative bacteria, viz. Escherichia coli and Salmonella enterica, were obtained
from the Microbial Culture Collection of Laboratory of Bacteriology of Faculty of Veterinary Medicine, Universiti Putra Malaysia, Serdang, Selangor, Malaysia. The bacterial cultures were subcultured and purified from stocks before use. Disc diffusion assay The antibacterial potential of A. paniculata and O. stamineus extracts was assessed from their reaction against four bacterial cultures using the Kirby-Bauer disc diffusion method (Bauer et al., 1966; Zaidan et al., 2005). Colonies picked from fresh pure culture plates were suspended in 5 ml sterile nutrient broth (OXOID, U.K.) and adjusted to a turbidity of 0.5 McFarland standard. Standardized bacterial suspensions were aseptically swabbed on to the surface of sterile Mueller Hinton Agar (MHA) plates. Twenty-five L of each plant extract (50 mg/ mL, 100 mg/mL and 200 mg/mL) were aseptically transferred to 6 mm sterilized paper discs to give a final concentration of 1.25, 2.5, and 5 mg/disc, respectively. The tests were replicated three times. The paper discs treated with herbal extracts were aseptically placed in petri plates containing Mueller-Hinton Agar (MHA) seeded with the respective test bacteria. The treated MHA plates were incubated in an upright position at 37C. The diameters of inhibition zones were measured after incubation for 24 h and the antibacterial properties were evaluated as follows: Resistant : < 7 mm inhibition zone Inhibition : Intermediate inhibition: 7-10 mm Moderate inhibition: 11-15 mm Strong inhibition: 16-20 mm The test bacteria were also assayed for their sensitivity to the antibiotics (OXOID, U.K.) tetracycline (30 g per disc), and 25 L of 10 % acetic acid. Data were subjected for oneway analysis of variance (ANOVA) and when differences were found among treatments, Duncans test was used to compare the mean differences using SPSS software (IBM SPSS version 21), with 0.05 % level of significance.
Antioxidant potential of Andrographis paniculata and Orthosiphon stamineus leaf extracts prepared with different solvents According to Wong et al. (2006), the antioxidant activity of the plant extracts basically depends on the composition of the extracts, hydrophobic or hydrophilic nature of the antioxidants, type of solvent used for extraction process, method of extraction, temperature and conditions of the test systems. Therefore, it is necessary to use more than one method for evaluation of antioxidant activity of plant extracts to encompass various mechanisms of antioxidant actions. The present study employed three tests to determine antioxidant activity of A. paniculata and O. stamineus leaf extracts as follows: Determination of total phenolic content Total phenolic content was estimated using the Folin-Ciocalteu colorimetric method described previously (Lu et al., 2011) with a slight modification. The sample extracts (0.2 mL) at appropriate dilutions were oxidized for 4 min with 1 mL of 0.5 M Folin-Ciocalteu reagent before the reaction was neutralized with 1 mL saturated sodium carbonate (75 g/ L). After 2 h incubation at room temperature, the absorbance of the resulting blue colour was measured at 760 nm on a spectrophotometer (Spectronic 20 GenesysTM). Quantification was performed against a standard curve of gallic acid. Results were expressed as mg equivalent to gallic acid (GAE) per 100 g of dry weight. Determination of free radical-scavenging activity The antioxidant activity of each extract was measured using the stable radical 2, 2diphenyl-1-picrylhydrazyl (DPPH) as a standard reagent. This was determined as described by Hatano et al. (1988) with minor modification. Briefly, stock solutions of the extract were prepared in ethanol. Ascorbic acid was taken as standard. Different concentrations of extract and a positive
control of ascorbic acid in ethanol solution (1 mL) were added to 3 ml of 0.004% ethanolic DPPH free radical solution. An equal amount of ethanol was used as a blank. After incubation at room temperature for 30 min in the dark, the absorbance was measured at 517 nm using a UV spectrophotometer (Spectronic 20 GenesysTM). Scavenging activity (%) was calculated using the following formula: DPPH radical scavenging (%) = [(Ac As) Ac] x 100 Where Ac is the absorbance of the control and As is the absorbance of the test sample after 30 min. Determination of superoxide radical scavenging activity The superoxide scavenging ability of the test samples was assessed according to the method of Ahn et al. (2012) with slight modifications. The reaction mixture comprised nitroblue tetrazolium (0.1 mM) and nicotinamide adenine dinucleotide (0.1 mM) with or without the test sample in a total volume of 1 ml of Tris- HCL buffer (0.02 M, pH 8.3). The reaction was started by adding 10 M phenazine methosulphate to the reaction mixture, and the change in the absorbance was recorded at 560 nm every 30 sec for 2 min. The percent inhibition was calculated against a control without the test sample. The results were compared with that of quercetin, with the percentage inhibition of superoxide anion generation calculated as: Scavenging (%) = [(Ac As) Ac] x 100 Where Ac is the absorbance of the control and As is the absorbance of the tested sample. Data on phenolic content, free radiclescavenging activity and superoxide radical scavenging activity were analyzed for oneway analysis of variance (ANOVA) and Duncans test was used to compare the mean differences using SPSS software (IBM SPSS version 21) at 0.05 % level of significance.
RESULTS AND DISCUSSION Qualitative screening for bioactive compounds Analyses on the methanolic extracts of A. paniculata and O. stamineus leaves indicated the presence of alkaloids, saponins, flavonoids, tannins, terpenoids and steroids (Table 1). This finding was consistent with several reports on the effectiveness of methanol in the extraction of bioactive compounds (Akowuah et al., 2005; 2006; Zargar et al., 2011). The chloroform extracts of both leaves were mainly poor in bioactive compound contents, except for alkaloids and flavonoids in the O. stamineus samples. Ethanolic extracts of O. stamineus leaves also showed the presence of all the bioactive compounds tested for (Table 1). The discrepancies in the levels of bioactive compounds in the two plant extracts could be related to their polarity differences (Akowuah et al., 2005). Analysis of andrographolide in A. paniculata leaf extracts by HPLC Figure 1 shows the chromatograms obtained for HPLC analyses of the water, ethanol, methanol and chloroform extracts of A.
paniculata leaves. The peak of andrographolide (AP) was identified by comparison of the retention time with that of the AP reference sample. AP quantification exhibited good linearity in the range from 10 to 120 ppm on the HPLC calibration curve (y = 479942x-175328; R = 0.998). Andrographis paniculata leaves extracted with methanol gave the highest peak as compared to samples obtained with the other extraction solvents. Extraction with methanol thus yielded the highest amount of AP (73.40 mg/g), followed by ethanol (35.85 mg/g), water (26.62 mg/g) and chloroform (15.63 mg/g) (Table 2). These observations are similar to that reported by Akowuah et al. (2006). Analysis of rosmarinic acid in O. stamineus leaf extracts by HPLC The HPLC chromatogram of the O. stamineus leaf extract is shown in Figure 2. Rosmarinic acid (RA) was identified in the water, ethanol, methanol leaf extracts, but it was not detected after extraction with chloroform. The RA peak was confirmed by comparison with the retention times of the reference standard. Various concentrations of the calibration standard exhibited a good
Table 1. Qualitative analysis of bioactive compounds in Andrographis paniculata and Orthosiphon stamineus leaf extracts prepared in different solvents Bioactive Compounds Sample Water Extract A. paniculata O. stamineus Methanol Extract A. paniculata O. stamineus Chloroform Extract A. paniculata O. stamineus EthanolExtract A. paniculata O. stamineusNote: present (+); absent (-).
+ + + + +
+ + + + +
+ + + + + +
+ + + + +
+ + + + + +
Figure 1. HPLC chromatograms (absorbance at 230 nm) of andrographolide (AP) from Andrographis paniculata leaves extracted with different solvents: (a) reference marker; (b) water extract; (c) ethanol extract; (d) methanol extract; (e) chloroform extract
linear variation with their photometric absorbance (y = 788923x 84260; R= 0.996) from 10 to 120 ppm. As shown in Figure 2 and Table 2, the results indicated that greatest peak was observed in the methanolic extract of O. stamineus as compared with extracts using other solvents. Accordingly, the highest recovery of RA (17.23 mg/g) was with methanol, followed by water (8.45 mg/g) and ethanol (1.10 mg/g).
Table 2. HPLC quantification of andrographolide in A. paniculata leaf extract and rosmarinic acid in O. stamineus leaf extract Extract solvents Water Ethanol Methanol Chloroform Andrographolide Rosmarinic acid
mg/g 26.62 35.85 73.40 15.63 1.06 c 0.20 b 0.72 a 0.72 d 08.45 0.02b 01.10 0.07c 17.32 0.41a Not detected
Values in each column bearing the same letter are not significantly different (P>0.05)
Figure 2. HPLC chromatograms (absorbance at 340 nm) of rosmarinic acid (RA) from Orthosiphon stamineus leaves extracted with different solvents: (a) reference marker; (b) water extract; (c) ethanol extract; (d) methanol extract; (e) chloroform extract (the arrow indicates the RA was undetected)
Antibacterial activity of A. paniculata and O. staminues leaf extracts Extracts of A. paniculata and O. stamineus prepared with water, ethanol, methanol, and chloroform were screened for their antibacterial activity against four species of bacteria. The extracts were tested at concentrations of 1.25, 2.50, and 5.00 mg/disc in the disc diffusion assay, which is an effective and rapid method to determine antimicrobial properties (Rovinsky & Cizadlo, 1998). Acetic acid that is commonly used as a food preservative, and tetracycline that is a synthetic antibiotic, respectively served as
controls. The diameters of the inhibition zones resulting from the different extracts are presented in Figure 3. In general, all extracts of A. paniculata and O. stamineus in different solvents exhibited at least some degree of bacterial growth inhibition. Among the treatments and controls, 30 g/disc of tetracycline (the control) showed the strongest anti-bacterial effect with S. enterica, S. aureus and B. cereus. Among the leaf extracts, the methanolic extract of A. paniculata exhibited the strongest inhibitory effects across the bacteria tested, especially at its highest concentration (5.00 mg/disc).
Figure 3. Susceptibility of pathogenic bacteria to Andrographis paniculata and Orthosiphon stamineus extracts prepared with different solvents. Note: Ctrl: Control; Ac: Acetic acid (10%); Tc: Tetracycline (30 g/disc). Heights of shaded bars represent inhibition diameters
This extract, although less effective than the tetracycline control, achieved an overall bacterial inhibition effect stronger than that shown by 10% acetic acid. Methanolic extracts of A. paniculata and O. stamineus at the highest tested concentration (5.00 mg/disc) in the present study showed the strongest bacterial inhibitory effects as compared with extracts prepared with other solvents. Similar observations have been reported by many researchers for various plant extracts (Negi et al., 2005; Parekh & Chanda, 2007, 2010; Al-Bayati, 2008; Kaushik & Goyal, 2011). The effectiveness of methanolic extracts of A. paniculata and O. stamineus against bacteria in the present study could be due to their polarity. Antibacterial activities of polar extracts could be attributed to the presence of several types of compounds belonging to different classes, such as flavonoids (Guillen & Manzanos, 1998), and the more polar thermo-labile and/or thermo-stable phenolics (Sokmen et al., 1999). The large amount of
andrographolide and rosmarinic acid detected in the methanolic extracts of A. paniculata and O. stamineus, respectively, as presented in the Table 2, may be significant contributors to the antibacterial effects observed. The Gram positive bacteria B. cereus and S. aureus were found to be more susceptible to the highest concentrations (5.00 mg/disc) of methanolic extract of O. stamineus and A. paniculata extracts of water, ethanol, methanol, and chloroform than the Gram negative bacteria (Figure 3). A report by AlBayati (2008) similarly indicated that B. cereus and S. aureus were more sensitive to plant essential oils and extracts as compared with Gram negative bacteria. The tolerance of Gram-negative bacteria to the herbal extract could be due to the presence of an outer membrane in bacteria that acts as a molecular filter to hydrophilic compounds (Benz & Bauer, 1988; Hammer et al., 2001; Al-Bayati, 2008).
Antioxidant activity of A. paniculata and O. staminues leaf extracts prepared with different solvents Total phenolic content In general, total phenolic contents of O. stamineus leaf extracts were higher as compared with similar extracts from A. paniculata leaves (Figure 4). The phenolic content of the O. stamineus methanolic extract was the highest at 1115.61 mg/100g GAE, followed by the ethanolic extract, the water extract and the chloroform extract. The average phenolic contents of the different O. stamineus extracts were significantly different from one another (p<0.05). Among the A. paniculata leaf extracts, methanolic and ethanolic extracts were again the highest in their phenolic contents, at 264.82 mg/100g GAE and 262.57 mg/100g GAE, respectively, followed by water and chloroform extracts. DPPH and superoxide scavenging activities Antioxidant activities of A. paniculata and O. staminues leaves extracted with different solvents were assessed by determining DPPH free radical scavenging and superoxide scavenging capacity. Figure 5 presents the
DPPH scavenging activity of A. paniculata and O. stamineus extracts in different solvents. In general, O. stamineus extracts produced higher DPPH scavenging ability as compared with A. paniculata extracts. Among the O. stamineus extracts, the methanolic extract of O. stamineus showed significantly higher DPPH scavenging activity at 97.5% as compared with the other solvents attempted. Among A. paniculata extracts, those prepared from methanol or ethanol (32.7% and 29.9% DPPH, respectively) were significantly higher in DPPH scavenging activity than extracts prepared with water or chloroform (IC50 value of 0.059 g/L ascorbic acid). The chloroform-extract exhibited significantly lower DPPH scavenging activity as compared with water, ethanol and methanol-extracts from both A. paniculata and O. stamineus. Results of superoxide scavenging activity of A. paniculata and O. stamineus leaves extracts are shown in Figure 6. Similarly, as shown in O. stamineus leaf extracts yielded higher superoxide scavenging activity as compared with A. paniculata extracts. Among the O. stamineus leaves extracts, methanolic and
Figure 4. Total phenol contents of Andrographis paniculata and Orthosiphon stamineus leaf extracts prepared with different solvents. Values in each column bearing the same letter are not significantly different (P>0.05)
Figure 5. DPPH scavenging activity of A. paniculata and O. stamineus extracts prepared with different solvents. Values in each column bearing the same letter are not significantly different (P>0.05)
Figure 6. Superoxide scavenging activity of A. paniculata and O. stamineus extracts prepared with different solvents. Values in each column bearing the same letter are not significantly different (P>0.05)
ethanolic extracts were again significantly higher in superoxide scavenging activity at 93.6% and 91.8% inhibition of superoxide anion generation respectively, as compared to water and chloroform extracts. 477
Methanolic and ethanolic extracts of A. paniculata showed a similar pattern of superoxide scavenging activity the corresponding extracts of O. stamineus. The percentage inhibition of superoxide anion
generation in the methanolic and ethanolic extracts were 39.1% and 36.4%, respectively, being significantly higher than the extracts prepared with chloroform (IC50 value 0.052 g/ml Quercetin). In the present study, methanolic and ethanolic extracts of A. paniculata and O. stamineus produced higher DPPH and superoxide scavenging activities than extracts prepared with other solvents. As in the case of their anti-bacterial properties, this observation could be explained by the presence of andrographolide and rosmarinic acid present in the leaves of A. paniculata and O. stamineus, respectively. The antioxidant effect of phenolic compounds is mainly due to their redox properties, and is the result of various possible mechanisms: free radical scavenging activity, transition metal-chelating activity and/or singlet oxygen-quenching capacity (Yao & Ren, 2011). Phenolic compounds are also known to play an important role in stabilizing lipid peroxidation and to inhibit various types of oxidizing enzymes (Gallo et al., 2010). Due to the above reasons, herbal plants have been regarded as sources of natural bioactive compounds. As mentioned above, the total phenolic compounds in O stamineus leaves were higher when extracted with methanol as compared with other solvents. These results are consistent with the earlier reports that showed a positive correlation between phenolic content and antioxidant activity (Akowuah et al., 2005; 2006; Tawaha et al., 2007; Premanath & Devi, 2011). Akowuah et al. (2005) reported that the methanol and water extracts of O. stamineus exhibited high free radical-scavenging properties. The methanol and ethanol extracts of A. paniculata recorded the highest phenolic content and also had the highest peroxide scavenging activity. This is in agreement with Akowuah et al. (2006) who also found that methanol extracts of A. paniculata exhibited high free radical scavenging activity as compared with water extract. The present study found that A. paniculata and O. stamineus extracted with polar solvents were higher in antioxidant activity rather than nonpolar solvents, in agreement with many 478
previous reports (Akowuah et al., 2005; 2006; Tawaha et al., 2007; Premanath & Devi, 2011). Different solvents of varying polarities were used for the extraction of bio-active compounds, including andrographolide in A. paniculata, and rosmarinic acid in O. stamineus leaves. The highest yields of these two compounds were obtained by methanol extraction. The methanolic extracts of A. paniculata and O. stamineus exhibited antimicrobial and antioxidant properties. The inhibitory effects against the Gram positive bacteria B. cereus and S. aureus shown by methanolic extracts of both plants were superior to that achieved with 10 % acetic acid. Methanolic and ethanolic extracts of A. paniculata and O. stamineus leaves also produced the strongest antioxidant activities as compared with extracts using other solvent. Such leaf extracts are potential candidates as natural antibacterial and antioxidant agents that can be beneficial to animal and human health. Acknowledgements. The authors would like to thank the Ministry of Higher Education Malaysia for financial support and Laboratory of Animal Production, Institute of Tropical Agriculture, Universiti Putra Malaysia for research and overall support.
REFERENCES Acda, M.N. (2009). Toxicity, tunneling and feeding behavior of the termite, Coptotermes vastator, in sand treated with oil of physic nut, Jatropha curcas. Journal of Insect Science 9: 1-7. Adebowale, K.O. & Adedire, C.O. (2006). Chemical composition and insecticidal properties of the underutilized Jatropha curcas seed oil. African Journal Biotechnology 5: 901-906. Ahn, Y.J., Ganesan, P. & Kwak, H.S. (2012). Comparison of polyphenol content and antiradical scavenging activity in methanolic extract of nanopowdered and powdered peanut sprouts. The Journal of Korean Society of Applied Biological Chemistry 55: 793-798.
Akowuah, G.A., Ismail, Z., Norhayati, I. & Sadikun, A. (2005). The effects of different extraction solvents of varying polarities on polyphenols of Orthosiphon stamineus and evaluation of the free radical-scavenging activity. Food Chemistry 93: 311-317. Akowuah, G.A., Zhari, I., Norhayati, I. & Mariam, A. (2006). HPLC and HPTLC densitometric determination of andrographolides and antioxidant potential of Andrographis paniculata. Journal of Food Composition Analysis 19: 118-126. Akowuah, G.A., Zhari, I., Norhayati, I., Sadikun, A. & Khamsah, S.M. (2004). Sinensetin, eupatorin, 3-hydroxy-5, 6, 7, 4-tetramethoxyflavone and rosmarinic acid contents and antioxidative effect of Orthosiphon stamineus from Malaysia. Food Chemistry 87: 559-566. Al-Bayati, F.A. (2008). Synergistic antibacterial activity between Thymus vulgaris and Pimpinella anisum essential oils and methanol extracts. Journal of Ethnopharmacology 116: 403406. Bajpai, V.K., Rahman, A. & Kang, S.C. (2007). Chemical composition and anti-fungal properties of the essential oil and crude extracts of Metasequoia glyptostroboides Miki ex Hu. Industrial Crop Production 26: 28-35. Barlow, S.N. (1990). Toxicological aspects of antioxidants used as food additives. In Hudson, B.J.F. (ed.), Food antioxidants, pp. 253-307. Amsterdam, Elsevier. Bauer, R.W., Kirby, M.D.K., Sherris, J.C. & Turck, M. (1966). Antibiotic susceptibility testing by standard single disc diffusion method. American Journal of Clinical Pathology 45: 493-496. Benz, R. & Bauer, K. (1988). Permeation of hydrophilic molecules through the outer membrane of gram negative bacteria. European Journal of Biochemistry 176: 1-19. Bera, D., Lahiri, D. & Nag, A. (2006). Studies on natural antioxidant for stabilization of edible oil and comparison with synthetic antioxidants. Journal of Food Engineering 74: 542-545. 479
Dung, N.T., Kim, J.M. & Kang, S.C. (2008). Chemical composition, antimicrobial and antioxidant activities of the essential oil and the ethanol extract of Cleistocalyx operculatus (Roxb.) Merr and Perry buds. Food Chemistry and Toxicology 46: 3632-3639. Ebrahimabadi, A.H., Ebrahimabadi, E.H., Djafari-Bidgoli, Z., Kashi, F.J., Mazoochi, A. & Batooli, H. (2010). Composition and antioxidant and antimicrobial activity of the essential oil and extracts of Stachys inflate Benth from Iran. Food Chemistry 119: 452-458. Edziri, H.L., Smach, M.A., Ammar, S., Mahjoub, M.A., Mighri, Z., Aouni, M. & Mas-touru, M. (2011). Antioxidant, antibacterial, and antiviral effects of Lactuca sativa extracts. Industrial Crop Production 34: 1182-1185. Gallo, M., Ferracane, R., Graziani, G., Ritieni, A. & Fogliano, V. (2010). Microwave assisted extraction of phenolic compounds from four different spices. Molecules 15: 6365-6374. Guillen, M.D. & Manzanos, M.J. (1998). Composition of the extract in dichloromethane of the aerial parts of a Spanish wild growing plant Thymus vulgaris L. Flavour & Fragrance Journal 13: 259-262. Hammer, K.A., Carson, C.F. & Riley, T.V. (2001). Antimicrobial activity of essential oils and other plant extracts. Journal of Applied Microbiology 86: 985-990. Hatano, T., Kagawa, H., Yasuhara, T. & Okuda, T. (1988). Two new flavonoids and other constituents in licorice root: their relative astringency and radical scavenging effects. Chemical and Pharmaceutical Bulletin 36: 1090-2097. Kaushik, P. & Goyal, P. (2011). Evaluation of various crude extracts of Zingiber officinale rhizome for potential antibacterial acitivity: a study in vitro. Advance Microbiology 1: 7-12. Lu, M., Yuan, B., Zeng, M. & Chen, J. (2011). Antioxidant capacity and major phenolic compounds of spices commonly consumed in China. Food Research International 44: 530-536.
Negi, P.S., Chauhan, A.S., Sadia, G.A., Rohinishree, Y.S. & Ramteke, R.S. (2005). Antioxidant and antibacterial activities of various seabuckthorn (Hippophae rhamnoides L.) seed extracts. Food Chemistry 92: 119-124. Parekh, J. & Chanda, S. (2010). Antibacterial and phytochemical studies on twelve species of Indian medicinal plants. African Journal of Biomedical Research 10: 175-181. Parekh, J. & Chanda, S.V. (2007). In vitro antimicrobial activity and phytochemical analysis of some Indian medicinal plants. Turkish Journal of Biology 31: 53-58. Perry, L.M. (1980). Medicinal plants of East Southeast Asia: Attributed Properties and Uses. MIT Press, Cambridge, MA. Premanath, R. & Devi, N.L. (2011). Antibacterial, antifungal and antioxidant activities of Andrographis paniculata Nees. leaves. International Journal Pharmaceutical Science Research 2: 2091-2099. Rovinsky, S.A. & Cizadlo, G.R. (1998). Salvia divinorum Epling et Jtiva-M. (Labiatae): An ethnopharmacological investigation. The McNair Scholarly Review 3: 142-156. Roy, S., Rao, K., Bhuvaneswari, C., Giri, A. & Mangamoori, L.N. (2010). Phytochemical analysis of Andrographis paniculata extract and its antimicrobial activity. World Journal of Microbiology and Biotechnology 26: 85-91.
Sokmen, A., Jones, B.M. & Erturk, M. (1999). The in vitro antibacterial activity of Turkish plants. Journal of Ethnopharmacology 67: 79-86. Tawaha, K., Alali, F.Q., Gharaibeh, M., Mohammad, M. & El-Elimat, T. (2007). Antioxidant activity and total phenolic content of selected Jordanian plant species. Food Chemistry 104: 1372-1378. Wong, C., Li, H., Cheng, K. & Chen, F. (2006). A systematic survey of antioxidant activity of 30 Chinese medicinal plants using the ferric reducing antioxidant power assay. Food Chemistry 97: 705711. Yao, Y. & Ren, G. (2011). Effect of thermal treatment on phenolic composition and antioxidant activities of two celery cultivars. LWT-Food Science and Technology 44: 181-185. Zaidan, M.R.S., Noor Rain, A., Badrul, A.R., Adlin, A., Norazah, A. & Zakiah, I. (2005). In vitro screening of five local medicinal plants for antibacterial acitivity using disc diffusion method. Tropical Biomedicine 22: 165-170. Zargar, M., Azizah, A.H., Roheeyati, A.M., Fatimah, A.B., Jahanshiri, F. & Pak-Dek, M.S. (2011). Bioactive compounds and antioxidant activity of different extracts from Vitex negundo leaf. Journal of Medicinal Plants Research 5: 2525-2532.


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Hight adjustable foot activated power liftgate w/jam protection, Wireless Smartphone Charger, charge indicator light, Bird's Eye View Monitor, 360 degree overhead view in drive and reverse and perimeter scan, 5 Door Smart Power locks, remote entry and hands free power rear liftgate
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hilary ison
Hilary Ison is a sensitive and skilled vocalist with wide musical interests and a penchant for early and new chamber music. Since her return to Vancouver in 2014 she has been delighted to sing with many of this city's leading ensembles, including the Vancouver Chamber Choir, musica intima, and Cor Flammae. Hilary's has been singing since infancy, and she continues to follow music's adventuresome path.


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Thank G-d for deadlines.
Without them, I'd get nothing done. Seriously. I put the "pro" in "procrastinating."
I delay entering expenses (mostly paper receipts) until a few weeks before April 15 (Ugh, adulting)
I wait until only a couple weeks before to book a flight (Oy, too many choices)
I edit and go over my stories seconds before I hit the stage (I hate rehearsing)
I have put off getting a new desk and chair for YEARS because I couldn't find the best one for my space and my wrists. Only now am I dealing with it because my back hurts so much. (See my, IMHO, hilarious video showing you the current state of my home office) Otherwise, I'd probably wait another five, ten, fifteen… who knows? (Hello, perfectionism)
I wanted and needed to throw things away and clean up my apartment, so I had people over for New Year's Eve to give myself a deadline. (Do whatcha gotta do, Gelf)
This procrastination (or some might say avoidance) is partially why I created this year of 30-day challenges. Among other things, I wanted to write more and this gives me a built-in deadline every single day.
However, with this focus on self-care this month, I'm becoming keenly aware of all of the areas where I'm putting things off and not taking care of myself. A big source of my agita is technology.
For a few months, the right speaker on my computer has been crackly.
Is it urgent? No. Is it annoying? Yes. Is it something I can live with? Of course, but I don't want to anymore, so a few weeks ago I decided to take action.
First stop. Google. I tried to troubleshoot my problem only to come up with bupkus.
Then, I went to Apple's site and since I haven't contacted them in so long (because I've put it off), I didn't even know you could call someone for tech support (literally a call for help.) I'm glad to know it's an option, but the guy couldn't help me with the speakers. For my other tech issues, I had to make more space and upgrade the software on my devices (no surprises there), so I did, but I still had unresolved issues (who doesn't?)
I made an appointment with the Apple Genius bar.
And that's where I found myself yesterday at the calmest place on Earth - Grand Central. I was greeted by the incredibly friendly Royal. I liked him right away. First, because it's a cool name and secondly because my favorite color is purple, which is a symbol of royalty. (Go with me on this.)
Before we even started working on the problem at hand, I was embarrassed by how smudgy my laptop was. I thought the marks were from my hands burying down into the laptop and thus, there forever, but Royal thought otherwise.
He got right on it cleaning my computer and he had some good shit (Whoosh), too. After some elbow grease, he made those marks disappear. He's a Genius and a magician?! Self-care FTW!
I told him about my Gelf-care challenge and he was intrigued. So much, in fact, that he wanted to read this very blog and possibly join the Facebook group I started in case anyone wants to do these challenges with me.
We tackled the speakers and he was straight with me. He told me it would take 3-5 days and cost $111. I didn't want go give up my computer for that long (How will I write this daily blog?)
He gave me a few other places to call because he's honest, helpful and looking out for #5 (He has a family, so I'm assuming I'm further down on the list.)
I asked him about helping me with being locked out of my iPad. He was happy to help, but he had to restart it, which took an inordinate amount of time.
I didn't want to waste one second, Hell, I was in the presence of a Genius and not just any Genius. A Royal Genius who holds the answers to my burning tech questions that are a "royal" pain in the ass (You're welcome.) With any luck, he can solve my problems (tech-related ones, that is) and help me feel better about some of my life choices (not syncing my phone often, etc. You get the picture.)
I asked him about merging calendars, phone backups, a Java alert that's popped up since I upgraded my software, iMovie file transfers and the list went on. He patiently answered my questions, explained things clearly and took care of business.
He even gave me a hot tip on a place (Five Below) where you can get tech accessories for $5 and under.
While we were waiting for my devices to restart, programs to load, etc., we totally bonded. He told me I was his favorite customer ever because I was so friendly and fun. I was flattered. I told him how much I appreciated his help and how I love connecting with people and above all, we talked about the importance of just being real.
And when we were done, we hugged. That's right. I hugged a Genius. It was so genuine and heartfelt and I started to sing "You give good hugs" a la "You Give Good Love" from Whitney Houston.
Royal had no idea what I was referring to, so I shared my Whitney wisdom with him. After all he'd given me the "royal" treatment, it only seemed fair.
So whether you're putting something off or putting up with something, take one step to take care of it today. Not only will you be helping yourself, you might be helping the world get a little closer. One exchange. One connection. Maybe even one hug at a time.
Are you a procrastinator or are you crushing your to-do list every day? If it's the former, tell me why. If it's the latter, please help. Share your wisdom below.


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Thank you for helping me grow Mini print
Thank you for helping me grow Mini print
A really sweet and pretty thank you print that is perfect for saying thank you to your teacher! Featuring this garland of pretty flowers and leaves and the words, this print is printed onto archival paper with pigmented inks and fits perfectly into an A5 frame .Thank you for helping me grow' painted and printed using a vintage typewriter. This comes packaged in a biodegradable cello wrap with a sticker explaining what it is.
made from:
Printed on to 300gsm thick, white recycled card and with a thick white envelope and hand packaged into a biodegradable cello wrap.
A6 size card and envelope.
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One of the best things about life here at the Villages of Citrus Hills is that we are located close to a number of unique Florida towns like Crystal River. And in addition to all of the fun things there are to do here in our community, there are even more options just 20 minutes up the road in Crystal River.
If you are looking for a fun way to spend a day in a nearby community, here are five popular ideas that are definitely worth checking out in Crystal River.
Swim With Manatees
While most visitors to Crystal River settle for simply seeing our beautiful manatees from the shore, residents of Crystal River know that you haven't really embraced the manatee culture until you get in the water and swim alongside them.
Companies like River Ventures Manatee Tour Center, located in downtown Crystal River, make swimming with manatees a breeze. All you have to do is show up at their office, watch a quick video breaking down the important things to know about getting in the water with these huge mammals, and then you'll be off to a local manatee hotspot with a snorkel and mask in hand.
Kings Bay Scenic Cruise
Nearby Kings Bay offers a wide range of water activities that keep Crystal River residents busy all year round, but one of the easiest ways to enjoy the water is to head out on a Kings Bay Scenic Cruise with Plantation Adventure Center.
This specific tour is a one-hour excursion that will get you up close and personal with some of the area's local wildlife. At the same time, you will have the privilege of taking in some of the best views of Kings Bay without ever having to actually get in the water.
Classic Sunset Cruises
For a more romantic version of the Kings Bay Scenic Cruise, you might decide to opt for the Classic Sunset Cruises offered by Wild Coast Charters. These two-hour tours feature live guitar music that provides the perfect accompaniment to the stunning views of a Kings Bay sunset.
The Classic Sunset Cruises are also popular because they feature a six-person limit on a boat that is capable of holding up to 20 guests, so there is always going to be plenty of room to enjoy this one.
Scalloping Charter
While swimming with manatees and Kings Bay cruises are popular activities throughout the year in Crystal River, one of the most popular activities between the months of July and September is always going to be heading out on a scalloping charter.
There are a number of companies that offer scalloping excursions, but each of them will have you snorkeling in shallow waters looking for scallops that you will be able to take home with you and prepare however you like.
Crystal River Segway Tours
If you are looking for something outside-the-box and you aren't afraid of looking like a goofy tourist, you'll have a hard time finding an excursion that is more fun than the Crystal River Segway Tours. And if you really want to experience Crystal River, this is a great introduction to the town.
The Segway Tours start in the Little Caesar's parking lot with a quick Segway training session. After that, it's off to explore the streets, parks, and waters of Crystal River.
Whether you are more interested in swimming, boating, or Segways, there is something for everyone located just up the road in Crystal River. And with Crystal River being so close to us here at the Villages of Citrus Hills, you are free to explore each of these fun activities whenever you like.


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We currently do not have an online form setup to register but you may print out the form below and mail it in or bring it by our gym located at
84 Pearl Industrial Hoschton, Ga 30548
Summer SESSION times and class offerings are posted on the sign up sheet. Registration opens Monday, April 22nd, and will be on a first come first serve basis. The only way to reserve your spot in any class is to turn in the bottom portion of your Summer_Session_2019_Registration_Sheets with payment to the front desk. Registrations will be accepted throughout the session.


text: Displaying 1 of 1 2013 Format: Atlas, Map, Printed Cartographic Material, Cartographic Material Title: Concise world atlas / DK ; [Dorling Kindersley Cartography]. Edition: 6th ed. Scales differ. Publisher, Date: London ; New York : Dorling Kindersley, c2013. Description: 1 atlas (xxxii, 350 p.) : col. ill., col. maps ; 34 cm. Contents: The world -- North America -- South America -- Africa -- Europe -- Asia -- Australia & Oceania -- Index-gazetteer. Summary: An encyclopedic view of the planet that includes high definition maps for the whole world complemented by photographs, diagrams, and topic maps. Genre: Atlases. Other Author: Dorling Kindersley Cartography (Firm) Other Title: World atlas concise DK world atlas Concise atlas of the world. Reference world atlas. Notes: "Everything you need to know about our planet today"--p. [1] of cover. "Previously published as Concise atlas of the world and also includes content published in Reference world atlas." [rev. American ed.] Relief shown by shading. Depths shown by gradient tints. Includes index. ISBN: 9781465402271 1465402276 System Availability: 1 Current Holds: 0 # Local items: 1 # Local items in: 1 # System items in: 1 Request It Add to My List Expand All | Collapse All Find It Recommended for You Map It Annotations Large Cover Image Librarian's View Series Information Similar Titles Similar Series Summary Reader Reviews Displaying 1 of 1