Identification of Plant Material: Corallocarpus Epigaeus L

Identification of Plant Material: Corallocarpus Epigaeus L 6. MATERIALS AND METHODS 6.1 Collection The plant material was collected from the Tirupati, Andhra pradesh, India in the month of February 2014. The plant taxonomical authenticated by Prof; Dr. K. Madhava chetty, Department of Botany, SV University, Tirupati. The voucher specimen of Corallocarpus epigaeus L has been preserved in our laboratory for further collection and reference. Chemicals Various reagents like Mayer’s reagent, Wagner’s reagent, Hager’s reagent, Dragendroff’s reagent, ÃŽ ± napthol solution, Fehlings solution A B, Barfoeds reagent, Millons reagent, Ninhydrins solution, cuso4, ethanol 95%, potassium hydroxide, concentrated HNO3, pyridine, sodium nitroprusside, sodium picrate, concentrated HNO3, pyridine, sodium nitroprusside, sodium picrate, concentrated H2SO4, Glacial acetic acid, Ferric chloride, Ammonium hydroxide solution, Potassium dichromate solution, Thionyl chloride solution, Phenolpthalein, Chloroform, etc. were received from standard suppliers to Dept of Pharmacology, SIPS-Proddatur. 6.1.1 Preparation of Whole Plant ethanolic extract of Corallocarpus epigaeus L The fresh leaves of Corallocarpus epigaeus L. The sieved powder was stored in airtight container and kept at room temperature for further study. The dried powdered material (250gm) was extracted with 95% ethanol using soxhlet apparatus for about 72hours. Figure no 12: SOXHLET APPARATUS DISTILLATION APPARATUS After extraction with solvent, the marc was dried in hot air oven below 50o c and was concentrated by distilling off the solvent and evaporating to dryness. The dried extract was subjected to preliminary phytochemical screening for detection of various phytoconstituents. 6.1.2 Qualitative Phytochemical Analysis74 The ethanolic extract Corallocarpus epigaeus L was subjected to various analytical tests in order to identify various phytoconstituents. Test for Alkaloids Mayer’s test To 1 ml of the extract, a drop or two drop of Mayer’s reagent was added by the side of test tube. Appearance of a white or creamy precipitate indicates presence of alkaloids. Wagner’s Test To 1 ml of the extract, few drops of Wagner’s reagent was added. Development of reddish brown colour indicates the presence of alkaloids. Hager’s Test To the 1 ml of the extract, few drops of Hager’s reagent was added. A prominent yellow colour indicates the test as positive. Dragendroff’s Test To the 1 ml of the extract, few drop of Dragendroff’s reagent was added. A prominent yellow colour indicates the test as positive. Test for Carbohydrates Benedict’s Test To 5 ml of Benedict’s reagent, 1 ml of the extract solution was added and boiled for 2 minute and cooled. Formation of red precipitate shows the presence of carbohydrates. Molisch’s Test To 2 ml of extract, two drops of alcoholic solution of ÃŽ ±-naphthol was added and shaken well. Later 1 ml of concentrated sulphuric acid was added slowly along the side of the test tubes and allowed to stand. A violet ring indicates the presence of carbohydrates. Fehling’s Test To 1 ml of the extract, add equal quantity of Fehling solution A and B were added. Appearance of red precipitate indicates the presence of sugars. Barfoed’s Test To 2 ml of the extract, 2 ml of Barfoed reagent was added and mixed well. It was heated for 1-2 minute in boiling water bath and cooled. Formation of red precipitate Indicates the presence of sugars. Test for Protein and Amino Acids Million’s Test To 2 ml of the extract, few drops of Million reagent was added. A white precipitate Indicates the presence of proteins. Ninhydrin Test To the 2 ml of the extract, two drops of Ninhydrin solution was added. A characteristic purple color indicates the presence of amino acids, proteins and peptides. Biurett Test To 1 ml of the extract, one or two drop of 1% copper sulphate solution was added and tothis 1 ml of ethanol (95%) was added, followed by excess of potassium hydroxide pellets. The pink layer in ethanolic layer indicates the presence of proteins. Xanthoprotein Test To 1 ml of the extract, add 1 ml of concentrated Nitric acid was added resulting in the formation of a white precipitate which is then boiled and cooled. Then 20% sodium hydroxide in ammonia was added. Orange colour indicates the presence of aromatic amino acids. Test for Glycosides Legal’s Test 2 ml of extract was dissolved in the solution of pyridine. Then sodium nitroprusside was added, to make it alkaline. The change in the colour from yellow to orange was not observed, which indicates the presence of glycosides. Baljet’s Test To 1 ml of the extract, 1 ml of sodium picrate solution was added. The colour from yellow to orange reveals the presence of glycosides. Borntrager’s Test To 1 ml of extract, few ml of sulphuric acid was added, boiled, filtered and extracted with chloroform. The chloroform layer was than treated with few ml of ammonia. The formation of red colour indicates the presence shows the presence of anthraquinone glycosides. Keller Killani Test The extract was dissolved in acetic acid containing traces of ferric chloride and was transferred to a test tube containing sulphuric acid. At the junction, the formation of reddish brown colour, which gradually turns to blue, confirms the presence of glycosides. Test for Flavonoids Shinoda Test To 1 ml of extract, magnesium turnings was added and 1-2 drops of concentrated hydrochloric acid was added drop wise. Formation of pink to crimson colour indicates the presence of flavonoids. Alkaline reagent Test The aqueous solution of the extract was treated with 10% ammonium hydroxide solution. Yellow fluorescence indicates the presence of flavonoids. Tests for Tannins and Phenolic compounds Ferric chloride Test To 1 ml of the extract, add few drops of neutral 5% ferric chloride solution. Formation of dark greenish colour shows the presence of phenolic compounds. To the extract add potassium dichromate solution, formation of a precipitate shows the presence of tannins and phenolic compounds. Test for Triterpenoids Two or three granules of tin metal were added to thionyl chloride solution present in a test tube. Later 1 ml of extract solution was added. The formation of pink colour indicates the presence of triterpenoids. Test for Saponins The 1 ml of the extract was diluted with distilled water and the volume was made up to 20 ml. The suspension was shaken in a graduated cylinder for 15 minutes. Appearance of foam indicates the presence of saponins. Tests for Fixed Oil Spot test A small quantity of the extract was pressed between two filter papers. Appearance of oily stain on the presence of fixed oils. Saponification Test A few drops of 0.5 N alcoholic potassium hydroxide solution was added to a small quantity of extract along with a drop of phenolphthalein. The mixture was heated on water bath for 2 hrs. Formation of the soap or partial neutralization of alkali indicates the presence of fixed oil. Tests for Steroids Libermann Buchard Test 1ml of the extract was dissolved in 2 ml of chloroform in a dry test tube. 10 drops of acetic anhydride and 2 drops of concentrated sulphuric acid was added to it. The solution turns to red, then blue and finally bluish green, indicating the presence of steroids. Salkowski Test The extract was dissolved in chloroform the extract in chloroform and equal volume of concentrated sulphuric acid was added. Formation of bluish red to cherry red colour in chloroform layer and green fluorescence observed acid indicates the presence of steroids. 6.2 INVITRO ANTICANCER ACTIVITY Cell Lines COLO 320 cell lineswere obtained from sugen Life Sciences Pvt. Ltd., Tirupati and cultured in RPMI 1640 medium (Difco, invitrogen corp, Canada). General information74 Organism Homo sapiens, human Tissue Colon Product Format Frozen Morphology Cells are rounded and refractile Culture Properties Loosely adherent, multicell aggregates Biosafety Level 1 Disease DukestypeC,colorectaladenocarcinomas Age 55 years Gender Female Ethnicity Caucasian Storage Condition Liquid nitrogen vapor phase GenesExpressed Serotonin,norepinephrine,epinephrine,Adreno Cortico Tropic Hormone (ACTH), parathyroid hormone. Tumorigenic effectsYes, in nude mice CommentsCells are weakly positive for keratins Culture Method The base medium for this cell line is formulated RPMI-1640 Medium. 6.2.1 Tryphan Blue dye Exclusion Assay Method75 Table 5: List of instruments Table 6: List of Chemicals Experimental Design The designed study consists of three groups viz: Negitive control, Control, Test. In the Negative control group the cell lines were incubated with the medium for a period of 24 hours. This group was designed to rule out the possibility of any growth inhibitory effect of certain compounds of medium. The control group was designed to rule out the effect of any residual or traces of solvent with which the extract was prepared on the growth inhibition of cell lines. Here the solvent employed was ethanol and hence it is added at the concentration of 0.1% (v/v) in distilled water. In test group different concentrations of test extract i.e, 10, 25, 50, 75 and 100 µg/ml are incubated with colo 320 cell lines for a period of 24 hours. This group was used to study the effect on cell line viability. Table7: Experimental Design to Study the Effect of Ethanolic Extract of Corallocarpus epigaeus L on colo 320 Cell Line Viability by Tryphan Blue Assay Procedure 1. An aliquot of cell suspension being tested for viability was centrifuged for 5 min andsupernatant was discarded. The size of the aliquot depends on the approximate number of cells present. The aliquot is taken such that it contained a convenient number of cells to count in a haemocytometer when suspended in 1 ml PBS and then diluted again by mixing with 0.4% Tryphan blue (e.g., 5 Ãâ€"105 cells/ml). 2. The cell pellet was resuspended in 1 ml PBS or serum free complete medium. Serum proteins stain with Tryphan blue and can produce misleading results. Hence determinations must be made in serum-free solution. 3. Mix 1 part of 0.4% Tryphan blue and 1 part cell suspension (dilution of cells) cells were mixed and allowed to incubate at room temperature for approximately 3 minutes, Cells were counted within 3 to 5 min of mixing with Tryphan blue, as longer incubation periods will lead to cell death and reduced viability counts. Mixing was performed in a well of a microtiter plate or a small plastic tube using 10 to20  µl each of cell suspension and Tryphan blue. 4. A drop of the Tryphan blue/cell mixture was applied on to haemocytometer. The haemocytometer on the stage of a binocular microscope and cells were focused clearly. 5. The no. of unstained (viable) and stained (nonviable) cells were counted separately in the haemocytometer. Seeding of Cells COLO 320 cells were cultured to reach the 80-90% confluency using RPMI 1640 medium. After reaching the desired confluency, culture was collected and centrifuged at 3000 rpm for 10 minutes to get cell pellet. The pellet was resuspended in 1ml of fresh culture media. Cell concentration was determined by Tryphan blue assay was performed by mixing 50  µlitres of culture and 50  µlitres of 0.4% tryphan blue dye. Finally cells seeded in 24 well plates at the concentration 10000 cells/ml and incubated at 5% co2 incubator at 370 c for 24 hours. Drug Treatment Cells were maintained in 24 well plates in triplicate for every concentration, and treated with different concentrations of (10, 25, 50, 75, 100 µgm). Corallocarpus epigaeus L, test compound and control groups were treated with medium and ethanol. The treated cells were incubated for 24 hours in 5% co2 incubator at 370c. Invitro Cytotoxic Assay After 24 hours incubation the cells were collected from each well in eppendroffs and centrifuged at 3000rpm for 10 min to get cell pellet, to the pellet 50 µlit of each medium and tryphan blue was added and mixed well to suspend the pellet. Cytotoxicity was screened by performing tryphan blue assay. Percent of growth inhibition was calculated by using the following formula 6.2.2 Micro Culture Tetrazolium Assay76 Plant Material Used: Whole plant ethanolic extract of Corallocarpus epigaeus L. Principle This assay is based on the capacity of mitochondria succinate dehydrogenase enzymes in living cells to reduce the yellow coloured watersolublesubstrate3(4,5dimethylthiazolyl)2,5diphenyltetrazoliumbromideintoaninsolublepurplecolouredformazanproductwhosecolouredismeasuredbymeansofELISAreaderat540nm.Onlyviablecellswithactivemitochondria reducesignificant amountsof MTT, since reduction of MTT can only occur in metabolically active cells. Figure 13: Reduction of MTT to a formazan compound by mitochondrial Enzymes Cell Lines Human colorectal adenocarcinoma- colo 320were obtained by sugen Life Sciences Pvt. Ltd., Tirupati from an authenticated supplier. Stock culture of these cell lines were cultured in RPMI -1640 with 10% inactivated newborn bovine serum, Penicillin (100 IU/ml), Streptomycin (100 µg/ml)) under humidified. The cells were dissociated in 0.2% trypsin and 0.02% EDTA in phosphate buffered saline solution. The stock culture was grown in 25cm 2 tissue culture flasks and cytotoxicity experiments were carried out in 96 well microtiter plates. Procedure Cell lines in the exponential growth phase were selected, washed, trypsinized and suspended in complete culture media i.e, RPMI 1640. The microtiter plates and incubated for 24hrs during which a partial monolayer was formed. They were then exposed to various concentrations of the extract (1-100 µg/ml). Control wells received only the maintenance medium. The plates were incubated at 37 °C and 48 hrs and cells were periodically checked for granularity, shrinkage and swelling. After 48 hrs, the sample solution in wells was flicked off and 50 µl of MTT dye was added to each well. The plates were gently shaken and incubated for 4 hrs at 370C in 5%CO2 incubator. The supernatant was removed and 50  µl of DMSO was added. The plates were gently shaken to solubilise the formed formazan. The absorbance was measured at 540nm. The percentage of growth inhibition was calculated using the following formula, Values of absorbance were converted into percentage of residual viability. Usually the Inhibition concentration 50% (IC50) is chosen as the best biological marker of cytotoxicity. The IC50 value represents the concentration of the test extracts that reduced 50% of cell inhibition. Statistical analysis Statistical evaluation of data was done by one-way analysis of variance (ANOVA) followed by Tukey’s multiple comparison test on viability using computer based fitting program (Prism graph pad prism version 6.03) statistical significance was set at p,0.05. IC50 was calculated by linear interpolation method using the formula IC50 = Ãâ€"(D-C)+C Where A = The first point on the curve, expressed in percent inhibition, that is less than 50% B = The first point on the curve, expressed in percent inhibition, that is greater than or equal to 50% C = The concentration of inhibitor that gives A% inhibition D = The concentration of inhibitor that gives B % inhibition

Practical Demonkeeping Chapter 19-20

19 JENNY'S HOUSE Jenny parked the Toyota behind Travis's Chevy and killed the lights. â€Å"Well?† Travis said. Jenny said, â€Å"Would you like to come in?† â€Å"Well.† Travis acted as if he had to think about it. â€Å"Yes, I'd love to.† â€Å"Give me a minute to go in and clear a path, okay?† â€Å"No problem, I need to check on something in my car.† â€Å"Thanks.† Jenny smiled with relief. They got out of the car. Jenny went into the house. Travis leaned against the door of the Chevy and waited for her to get inside. Then he threw open the car door and peeked inside. Catch was sitting on the passenger side, his face stuck in a comic book. He looked up at Travis and grinned. â€Å"Oh, you're back.† â€Å"Did you play the radio?† â€Å"No way.† â€Å"Good. It's wired into the battery directly; it'll drain the current.† â€Å"Didn't touch it.† Travis glanced at the suitcase on the backseat. â€Å"Keep an eye on that.† â€Å"You got it.† Travis didn't move. â€Å"Is there something wrong?† â€Å"Well, you're being awfully agreeable.† â€Å"I told you, I'm just glad to see you having a good time.† â€Å"You may have to stay the night in the car. You aren't hungry, are you?† â€Å"Get a grip, Travis. I just ate last night.† Travis nodded. â€Å"I'll check on you later, so stay here.† Travis closed the car door. Catch jumped to his feet and watched over the dashboard while Travis went into the house. Ironically, they were both thinking the same thing: in a little while this will all be over. Catch coughed and a red spiked heel shot out of his mouth and bounced off the windshield, spattering the glass with hellish spit. Robert had parked his truck a block away from his old house and walked up, hoping and dreading that he would catch Jenny with another man. As he approached the house, he saw the old Chevy parked in front of her Toyota. He had run through this scene a hundred times in his mind. Walk out of the dark, catch her with the guy, and shout â€Å"Ah ha!† Then things got sketchy. What was the point? He didn't really want to catch her at anything. He wanted her to come to the door with tears streaming down her cheeks. He wanted her to throw her arms around him and beg him to come home. He wanted to assure her that everything would be fine and forgive her for throwing him out. He had run that scene through his mind a hundred times as well. After they made love for the third time, things got sketchy. The Chevy was not part of his preconceived scenes. It was like a preview, a teaser. It meant that someone was in the house with Jenny. Someone who, unlike Robert, had been invited. New scenes ran through his mind: knocking on the door, having Jenny answer, looking around her shoulder to see another man sitting on the couch, and being sent away. He couldn't stand that. It was too real. Maybe it wasn't a guy at all. Maybe it was one of the women from the coven who had stopped over to comfort Jenny in her time of need. Then the dream came back to him. He was tied to a chair in the desert again, watching Jenny make love with another man. The little monster was shoving saltines in his mouth. Robert realized he had been standing in the middle of the street staring at the house for several minutes, torturing himself. Just be adult about it. Go up and knock on the door. If she is with someone else, just excuse yourself and come back later. He felt an ache rising in his chest at the thought. No, just walk away. Go back to The Breeze's trailer and call her tomorrow. The thought of another night alone with his heartbreak increased the ache in his chest. Robert's indecision had always angered Jenny. Now it was paralyzing him. â€Å"Just pick a direction and go, Robert,† she would say. â€Å"It can't be any worse than sitting here pitying yourself.† But it's the only thing I'm good at, he thought. A truck rounded the corner and started slowly to roll up the street. Robert was galvanized into action. He ran to the Chevy and ducked behind it. I'm hiding in front of my own house. This is silly, he thought. Still, it was as if anyone who passed would know how small and weak he was. He didn't want to be seen. The truck slowed almost to a stop as it passed the house, then the driver gunned the engine and sped off. Robert stayed in a crouch behind the Chevy for several minutes before he moved. He had to know. â€Å"Just pick a direction and go.† He decided to peek in the windows. There were two windows in the living room, about six feet off the ground. Both were old-style, weighted-sash types. Jenny had planted geraniums in the window boxes outside. If the window boxes were strong enough, he could hoist himself up and peek through the gap in the drawn curtains. Spying on your own wife was sleazy. It was dirty. It was perverse. He thought about it for a moment, then made his way across the yard to the windows. Sleazy, dirty, and perverse would be improvements over how he felt now. He grabbed the edge of the window box and tested his weight against it. It held. He pulled himself up, hooked his chin on the window box, and peered through the gap in the curtains. They were on the couch, facing away from him: Jenny and some man. For a moment he thought Jenny was naked, then he saw the thin straps of her black dress. She never wore that dress anymore. It gave out the wrong kind of message, she used to say, meaning it was too sexy. He stared at them in fascination, caught by the reality of his fear like a deer caught in car headlights. The man turned to say something to Jenny, and Robert caught his profile. It was the guy from the nightmare, the guy he had seen in the Slug that afternoon. He couldn't look any longer. He lowered himself to the ground. A knot of sad questions beat at him. Who was this guy? What was so great about this guy? What does he have that I don't? Worst of all, how long has this been going on? Robert stumbled away from the house toward the street. They were sitting in his house, on his couch – the couch he and Jenny had saved up to buy. How could she do that? Didn't everything in the house remind her of their marriage? How could she sit on his couch with some other man? Would they screw in his bed? The ache rose up in his chest at the thought, almost doubling him over. He thought about trashing the guy's car. It was pretty trashed already, though. Flatten the tires? Break the windshield? Piss in the gas tank? No, then he would have to admit to spying. But he had to do something. Maybe he could find something in the car that would tell him who this home wrecker was. He peered through the Chevy's windows. Nothing much to see: a few fast-food wrappers, a comic book on the front seat, and a Haliburton suitcase on the backseat. Robert recognized it immediately. He used to carry his four-by-five camera in the same model suitcase. He had sold the camera and given the suitcase to The Breeze for rent. Was this guy a photographer? One way to find out. He hesitated, his hand on the car door handle. What if the guy came out while Robert was rummaging through the car? What would he do? Fuck it. The guy was rummaging through his life, wasn't he? Robert tried the door. It was unlocked. He threw it open and reached in. 20 EFFROM He was a soldier. Like all soldiers, in his spare moments he was thinking of home and the girl who waited for him there. He sat on a hill looking out over the rolling English countryside. It was dark, but his eyes had adjusted during his long guard duty. He smoked a cigarette and watched the patterns the full moon made on the hills when the low cloud cover parted. He was a boy, just seventeen. He was in love with a brown-haired, blue-eyed girl named Amanda. She had down-soft hair on her thighs that tickled his palms when he pushed her skirt up around her hips. He could see the autumn sun on her thighs, even though he was staring over the spring-green hills of England. The clouds opened and let the moon light up the whole countryside. The girl pulled his pants down around his knees. The trenches were only four days away. He took a deep drag on the cigarette and stubbed it out in the grass. He let the smoke out with a sigh. The girl kissed him hard and wet and pulled him down on her. A shadow appeared on the distant hill, black and sharply defined. He watched the shadow undulate across the hills. It can't be, he thought. They never fly under a full moon. But the cloud cover? He looked in the sky for the airship but could see nothing. It was silent except for the crickets singing sex songs. The countryside was still but for the shadow. He lost the vision of the girl. Everything was the huge, cigar-shaped shadow moving toward him, silent as death. He knew he should run, sound the alarm, warn his friends, but he just sat, watching. The shadow eclipsed the moonlight and he shivered, the airship was directly over him. He could just hear the engines as it passed. Then he was bathed in moonlight, the shadow behind him. He had survived. The airship had held its bellyful of death. Then he heard the explosions begin behind him. He turned and watched the flashes and fires in the distance, listened to the screams, as his friends at the base woke to find themselves on fire. He moaned and curled into a ball, flinching each time a bomb exploded. Then he woke up. There was no justice; Effrom was sure of it. Not an iota, not one scintilla, not a molecule of justice in the world. If there was justice, would he be plagued by nightmares from the war? If there was any justice would he be losing sleep over something that had happened over seventy years ago? No, justice was a myth, and it had died like all myths, strangled by the overwhelming reality of experience. Effrom was too uncomfortable to mourn the passing of justice. The wife had put the flannel sheets on the bed to keep him cozy and warm in her absence. (They still slept together after all those years; it never occurred to them to do any different.) Now the sheets were heavy and cold with sweat. Effrom's pajamas clung to him like a rain-blown shroud. After missing his nap, he had gone to bed early to try to recapture his dreams of spandex-clad young women, but his subconscious had conspired with his stomach to send him a nightmare instead. Sitting on the edge of the bed, he could feel his stomach bubbling away like a cannibal's caldron, trying to digest him from the inside out. To say that Effrom was not a particularly good cook was an understatement akin to saying that genocide is not a particularly effective public relations strategy. He had decided that Tater Tots would provide as good a meal as anything, without challenging his culinary abilities. He read the cooking instructions carefully, then did some simple mathematics to expedite the preparation: twenty minutes at 375 degrees would mean only eleven minutes at 575 degrees. The results of his calculation resembled charcoal briquettes with frozen centers, but because he was in a hurry to get to bed, he drowned the suffering Tots in catsup and ate them anyway. Little did he know that their spirits would return carrying nightmare images of the zeppelin attack. He had never been so frightened, even in the trenches, with bullets flying overhead and mustard gas on the wind. That shadow moving silently across the hills had been the worst. But now, sitting on the edge of the bed, he felt the same paralyzing fear. Though the dream was fading, instead of the relief of finding himself safe, at home, in bed, he felt he had awakened into something worse than the nightmare. Someone was moving in the house. Someone was thrashing around like a two-year-old in a pan-rattling contest. Whoever it was, was coming through the living room. The house had a wooden floor and Effrom knew its every squeak and creak. The creaks were moving up the hall. The intruder opened the bathroom door, two doors from Effrom's bedroom. Effrom remembered the old pistol in his sock drawer. Was there time? Effrom shook off his fear and hobbled to the dresser. His legs were stiff and wobbly and he nearly fell into the front of the dresser. The floor was creaking outside the guest bedroom. He heard the guest room door open. Hurry! He opened the dresser drawer and dug around under his socks until he found the pistol. It was a British revolver he had brought home from the war – a Webley, chambered for.45 automatic cartridges. He broke the pistol open like a shotgun and looked into the cylinders. Empty. Holding the gun open, he dug under his socks for the bullets. Three cartridges were held in a plate of steel shaped like a half-moon so the pistol's six cylinders could be loaded in two quick motions. The British had developed the system so they could use the same rimless cartridges in their revolvers that the Americans used in their Colt automatics. Effrom located one of the half-moon clips and dropped it into the pistol. Then he started searching for the sound. The doorknob of his room started to turn. No time. He flipped the gun upward and it slammed shut, only half loaded. The door slowly started to swing open. Effrom aimed the Webley at the center of the door and pulled the trigger. The gun clicked, the hammer fell on an empty chamber. He pulled the trigger again and the gun fired. Inside the small bedroom the gun's report sounded like the end of the world. A large, ragged hole appeared in the door. From the hall came the high-pitched scream of a woman. Effrom dropped the gun. For a moment he stood there, gunfire and the scream echoing in his head. Then he thought of his wife. â€Å"Oh my God! Amanda!† He ran forward. â€Å"Oh my God, Amanda. Oh my†¦Ã¢â‚¬  He threw the door open, leapt back, and grabbed his chest. The monster was down on its hands and knees. His arms and head filled the doorway. He was laughing. â€Å"Fooled you, fooled you,† the monster chanted. Effrom backed into the bed and fell. His mouth moved like wind-up chatter dentures, but he made no sound. â€Å"Nice shot, old fella',† the monster said. Effrom could see the squashed remains of the.45 bullet just above the monster's upper lip, stuck like an obscene beauty mark. The monster flipped the bullet off with a single claw. The heavy slug thudded on the carpet. Effrom has having trouble breathing. His chest was growing tighter with each breath. He slid off the bed to the floor. â€Å"Don't die, old man. I have questions for you. You can't imagine how pissed I'll be if you die now.† Effrom's mind was a white blur. His chest was on fire. He sensed someone talking to him, but he couldn't understand the words. He tried to speak, but no words would come. Finally he found a breath. â€Å"I'm sorry, Amanda. I'm sorry,† he gasped. The monster crawled into the room and laid a hand on Effrom's chest. Effrom could feel the hand, hard and scaly, through his pajamas. He gave up. â€Å"No!† the monster shouted. â€Å"You will not die!† Effrom was no longer in the room. He was sitting on a hill in England, watching the shadow of death floating toward him across the fields. This time the zeppelin was coming for him, not the base. He sat on the hill and waited to die. I'm sorry, Amanda. â€Å"No, not tonight.† Who said that? He was alone on the hill. Suddenly he became aware of a searing pain in his chest. The shadow of the airship began to fade, then the whole English countryside dissolved. He could hear himself breathing. He was back in the bedroom. A warm glow filled his chest. He looked up and saw the monster looming over him. The pain in his chest subsided. He grabbed one of the monster's claws and tried to pry it from his chest, but it remained fast, not biting into the flesh, just laid upon it. The monster spoke to him: â€Å"You were doing so good with the gun and everything. I was thinking, ‘This old fuck really has some gumption.' Then you go and start drooling and wheezing and ruining a perfectly good first impression. Where's your self-respect?† Effrom felt the warmth on his chest spreading to his limbs. His mind wanted to switch off, dive under the covers of unconsciousness and hide until daylight, but something kept bringing him back. â€Å"Now, that's better, isn't it?† The monster removed his hand and backed to the corner of the bedroom, where he sat cross-legged looking like the Buddha of the lizards. His pointy ears scraped against the ceiling when he turned his head. Effrom looked at the door. The monster was perhaps eight feet away from it. If he could get through it, maybe†¦ How fast could a beast that size move in the confines of the house? â€Å"Your jammies are all wet,† the monster said. â€Å"You should change or you'll catch your death.† Effrom was amazed at the reality shift his mind had made. He was accepting this! A monster was in his house, talking to him, and he was accepting it. No, it couldn't be real. â€Å"You're not real,† he said. â€Å"Neither are you,† the monster retorted. â€Å"Yes I am,† Effrom said, feeling stupid. â€Å"Prove it,† the monster said. Effrom lay on the bed thinking. Much of his fear had been replaced by a macabre sense of wonder. He said: â€Å"I don't have to prove it. I'm right here.† â€Å"Sure,† the monster said, incredulously. Effrom climbed to his feet. Upon rising he realized that the creak in his knees and the stiffness he had carried in his back for forty years were gone. Despite the strangeness of this situation, he felt great. â€Å"What did you do to me?† â€Å"Me? I'm not real. How could I do anything?† Effrom realized he had backed himself into a metaphysical corner, from which the only escape was acceptance. â€Å"All right,† he said, â€Å"you're real. What did you do to me?† â€Å"I kept you from croaking.† Effrom made a connection at last. He had seen a movie about this: aliens who come to Earth with the power to heal. Granted, this wasn't the cute little leather-faced, lightbulb-headed alien from the movie, but it was no monster. It was a perfectly normal person from another planet. â€Å"So,† Effrom said, â€Å"do you want to use the phone or something?† â€Å"Why?† â€Å"To phone home. Don't you want to phone home?† â€Å"Don't play with me, old man. I want to know why Travis was here this afternoon.† â€Å"I don't know anyone named Travis.† â€Å"He was here this afternoon. You spoke with him – I saw it.† â€Å"You mean the insurance man? He wanted to talk to my wife.† The monster moved across the room so quickly that Effrom almost fell back on the bed to avoid him. His hopes of making it through the door dissolved in an instant. The monster loomed over him. Effrom could smell his fetid breath. â€Å"He was here for the magic and I want it now, old man, or I'll hang your entrails from the curtain rods.† â€Å"He wanted to talk to the wife. I don't know nothin' about any magic. Maybe you should have landed in Washington. They run things from there.† The monster picked Effrom up and shook him like a rag doll. â€Å"Where is your wife, old man?† Effrom could almost hear his brain rattling in his head. The monster's hand squeezed the breath out of him. He tried to answer, but all he could produce was a pathetic croak. â€Å"Where?† The monster threw him on the bed. Effrom felt the air burn back into his lungs. â€Å"She's in Monterey, visiting our daughter.† â€Å"When will she be back? Don't lie. I'll know if you are lying.† â€Å"How will you know?† â€Å"Try me. Your guts should go well with this decor.† â€Å"She'll be home in the morning.† â€Å"That's enough,† the monster said. He grabbed Effrom by the shoulder and dragged him through the door. Effrom felt his shoulder pop out of its socket and a grinding pain flashed across his chest and back. His last thought before passing out was, God help me, I've killed the wife.

Natural disasters Essay

BTA’S overseas offices work closely with the British diplomatic and cultural staff, the local travel trade and media to stimulate interest in Britain. In the UK it has partnerships with other organizations such as the British Council. The National Tourist Boards (NTB’s) These were established in 1969 when the Development of Tourism Act was passed. The English Tourism Council (ETC) replaced the English Tourism Board in 1999 These are responsible for promoting domestic tourism in their own countries. There are 4 main tourist boards. They all have similar objectives which include: Advising on matters of tourism   Contribution to the economy by creating wealth and employment   Making a good image for their countries as tourism destinations   To help the sustainable tourism development   And to research the consumer requirements in travel and tourism The Voluntary Sector These are usually non profitable or charity organizations and are done by volunteers. Examples include the Tourism Concern and the National Trust. The national trust was founded in 1895 and their aims are to preserve historic places or natural beauty permanently for the nation. The tourism concerns aim is to campaign for fair and ethically traded tourism. New Technologies Travel and Tourism is dependent on technology in the 21st century in order to keep up with consumer expectations and to be able to operate in an efficient way. Many use the Computer Reservations Systems (CRS) or the Global Distributions Systems (GDS) which are explained earlier in the project. The 4 main GDS are:   Sabre   Amadeus   Galileo   Worldspan External Pressures Travel and Tourism is also vulnerable against the external pressures for which it has no control over such as: Currency Fluctuation. State of the Economy Government Legislation Climate Changes   Natural Disasters   War and Acts of Terrorism Currency Fluctuation The exchange rates are always changing. When the value of the pound increases this encourages the outbound tourism as the strength of the pound against the other currencies means holidaymakers get more for their money Legislation Tax revenue also has a big impact on the travel and tourism industry. When air passenger duty (APD) introduced in 1997 every flight from the UK to Europe has had i 10 added on to them and outside of the EU i 20. EU Legislation has had a big impact as well. The EU directive on packaged holidays means that any company that is an organization must have a bond or other financial protection in place or they could be prosecuted. Climate Change When the UK has a poor summer it leads to a bad effect of tourism in the seaside destinations. With the use of the internet people can see what the weather can be like in advanced and if it is bad then those people are likely to looks elsewhere. Natural Disasters Natural Disasters such as earthquakes, tornadoes and floods all affect tourism. For example the hurricanes that hit Florida would have affected the tourism there in the summer months. War, Acts of Terrorism Travel and Tourism is vulnerable to war and terrorism acts. The Middle East and the events of September 11th in the USA has affected peoples confidence of flying and travelling to these parts of the world especially. Impact on Host Communities There can be usually a positive and negative impact on host communities. There are Economic, Social and Environmental impacts on tourism. Economic Travel and Tourism has a big impact on the UK economy in the income generated, numbers employed, and balance of payments. A lot of industries benefit from direct and indirect income generated by visitors to the area. Indirect income is brought in from the multiplier effect and this is when money spent by visitors is taken into the local economy. This can also be applied to numbers employed. The opposite to this is leakage Social The local communities can also benefit from public, private and voluntary sectors. Services such as restaurants, shopping complexes and transport services are all examples. Also areas that are neglected are usually done up in order to impress. However, the impact on tourism on host communities are often more negative than positive. Environmental. There are big concerns about what tourism is having on the environmental side of tourism. This has led to big demanding of sustainable tourism. Type of Impact Positive Impacts Negative Impacts Economic Multiplier Effect Jobs Foreign Currency Earnings Money to Develop the area and infrastructure Leakage Jobs can be part time Seasonal Low Paid Visitor Management costs Rise in House Prices Social/Cultural Supports Culture Lead to crime/vandalism Language Dress Music Conflict Environmental Improve and Maintain tourist places Improved infrastructure Preserve Wildlife Pollution Noise Litter Resource Depletion Overcrowding. Lead to crime/vandalism E2 – Scale of the UK Industry and its Economic Significance For E2 I am going to find out statistics on each of the sectors of the travel and tourism industry. I will try to find out the numbers employed in each of these components and the number of participants in these components as well as the consumer spending. I will also show the source from which I obtained these statistics from Component Numbers Employed Number of Participants Consumer Spending Accommodation and Catering Hotels and other Accommodation – 318,7000 Restaurant and Cafi ‘s 356. 0000 Bars, Pubs, Clubs 364. 1000. Source: Office for National Statistics This shows the consumer spending in accommodation in percent Hotels 41% B&B 8% Flat/Chalet 9% Youth Hostel 1% Camping 2% Source: www. staruk. com Travel Agents Below are some branches and how many there are. Lunn Poly 797 Going Places 738 Thomas Cook 390 Another example is the CO-OPERATIVE WHOLESALE SOCIETY LTD with 289 branches this employs around 35,200 people Source: AbtaResearch. com 7. 2 million UK residents book their holidays person to person in travel agents but this is been out done by the internet which is getting 10. 9 million Source: www. staruk. com Tour Operators 83. 5000 people employed in travel agencies and tour operators. Source: Office for National Statistics 257 million visits overall Source: staruk. com Transport There are 87,000 people employed in this type of work Source VisitBritain. co. uk This shows the percentage of trips. Car 73% Train 12% Bus 4% Coach 1% Caravan   Motor Cycle Bicycle   Plane 5% Boat/Ship Other 2% Source: www. staruk. com   Ã¢â‚¬â€œ less than 1% Visitor Attractions Libraries, museums, culture 77. 0000 Source: Office for National Statistics Alton Towers 2,650,00 Tower of London 2,422,181 Lego Land 1,620,000 Flamingo Land 1,197,000 Source: www. staruk. com. Tourism Development There are 450 people employed and 60% of these are employed in overseas markets Source: Visitbritain. co. uk The government participate in tourism development and promotion because without it they would lose out on money from tourists Source: Visit Britain. co. uk Incoming Tourists 24. 2 million trips made spending around i qq. 7 billion Domestic Tourists 167. 3 million trips made and spending around i 26. 6 billion Outgoing Tourists 24. 18 million trips made spending around i 11. 618 million Business Tourism Another significant part of the travel and tourism industry is the business tourism. The table below shows the amount of trips that were made by business tourism and the spending in this for the UK residents in the year of 2001. Trips (Millions) Spent (i Millions) 22. 8 5,670 Source: United Kingdom Tourism Survey / International Passenger Survey E2 – Scale of the UK Travel and Tourism Industry Travel and Tourism is a multi-billion pound industry in the UK. Visit Britain estimates that domestic and overseas tourists spent a total of around i 26 million in 1998. 60% of this spending was covered by the Accommodation and Catering sector. The scale of the industry can be established by looking at: Travel and Tourism revenue and its contribution to the national economy Employment Statistics   Inbound, Outbound and Domestic tourist numbers within the UK Travel and Tourism Revenue Inbound and Domestic tourism contributes to the national economy   A wide range of industries benefit from direct and indirect income, this of which is generated by tourist   The Travel and Tourism industry contributes to the economy by tourists paying VAT and taxes o products   Local Travel and Tourism industries contribute to local government finances through payment of business rates Contribution to the Balance of Payments. The Balance of Payments refers to the difference in the nations economy between the income generated from exports and the cost of imports   It is harder to measure in tourism as tourists are paying for services rather than products and therefore it is described as being invisible   Inbound Tourism is seen as an export because overseas tourists are bringing money into the UK economy. They are buying British products and services   Outbound Tourism is seen as an import because it is taking money out of the UK economy There is a deficit in the balance of payments when the cost of imports is greater than the income of exports. Outbound UK tourists spending more money abroad than the Inbound (overseas) visitors do in the UK   There is a surplus in the balance of payments when the income of exports is greater than the cost of imports. Inbound tourists spending more money in the UK than the money taken out by outbound tourism   More money is being spent by the British abroad than overseas visitors to the UK and therefore there is a deficit in the balance of payments   The British Government is trying to resolve this by encouraging more overseas visitors to the UK. E2 – Scale of the UK Industry and its Economic Significance. This is a multi billion pound industry and has a major influence on the economy in the terms of consumer spending and employment. Examples of organizations that produce statistics are:   Government statistics   British tourist authority National and regional tourist boards   Industry bodies such as ABTA, Mintel and Keynote To assess the scale of the UK industry and its economic significance we have to analyse the UK travel and tourism revenue and its contribution to the national economy and also look into employment statistics and research into inbound, outbound and domestic tourism in the UK. Visit England states that in the year of 1998 the UK industry was worth i 61,201 million making the UK the fourth largest earner of foreign exchange. This is what the money was spent on Trips (Millions) Spending (i m) UK Residents 122. 3 14,030 Overseas Visitors 25. 7 12,671 Total 148. 0 26,701 This information was taken from the Collins text book on Travel and Tourism An example of a visitor attraction for which I have found statistics on is Flamingo Land. This visitor attraction employs about 375 staff a year and work either full time or part time from March to November. Flamingo Land has been able to increase its employment as well because of the opening of a 26 bed roomed staff accommodation block and this has brought in staff from as far away as Wales and Scotland. Most of the staff who are employed at Flamingo Land though are from closer in and in the local areas of Scarborough, Malton, York, Whitby and Pickering E3 A Full Explanation of the Present Structure of the Industry In order to give a clear explanation of the industry I am going to describe the role of the 6 components within it using a case study for each to explain the components role. I am also going to explain the chain of distribution within the industry. The 6 components are: 1. Accommodation and Catering 2. Tour Operators 3. Travel Agents 4. Transport 5. Visitor Attractions 6. Tourism Promotion and Development 1. Accommodation and Catering Accommodation can be either serviced or self-catering Serviced Accommodation Hotels and serviced accommodation are all different in terms of size, quality, turnover and facilities. National tourist boards use classification systems for hotels, bed and breakfast’s and guest houses. They are awarded star ratings for the range of facilities and services provided Examples of serviced accommodation are:   Hotels   Lodge   Bed Only   Bed and Breakfast   Half Board   Full Board All Inclusive Self Serviced Accommodation (Self-Catering) There is a wide range of self serviced accommodation available. Examples are:   Camping Youth Hostels   Apartments   Villas/Chalets Boat   Cottages   Guest House Especially popular are places with self catering accommodation combined with activities and entertainment. Examples of this are Center Parcs and Butlins Catering Examples of catering are:   Fast Food Outlets – McDonalds, Pizza Hut, Burger King. Restaurants – Frankie and Bennies Cafi ‘s   Pubs – Rose and Crown   Takeaways – Dominos Pizza   In-house Restaurants (Supermarkets) – Asda, Tesco’s   Road-side Catering – Burger bars outside football grounds Restaurant and takeaway markets have continued to grow partly down to the popularity in ethnic restaurants and takeaway shops. Pub meals have remained popular. An example of accommodation and catering is the YHA (Youth Hostel Association) and this operates a network of over 227 youth hostels in the UK. It takes in groups and individuals. This is an example of a voluntary organization. 2. Tour Operators Tour operators arrange the following:   Transport Accommodation   Leisure Activities Holiday packages are usually sold through travel operators and the travel agencies receive a commission for been the one who sells the product for them. Tour operation markets provide products and services for three main categories of tourism which are:   Outbound   Inbound   Domestic Outbound Operators The majority of outbound operators are situated in the UK and they organize packaged holidays. Examples of outbound operators are:   Thomson Holidays   First Choice Holidays   Airto.

Global Warming - Is It Man-Made or Mother Nature or Both

Global Warming - Is It Man-Made or Mother Nature or Both? Global warming is define as the global increase in the average temperature of the earth, that near the air-surface and the oceans which caused by the emission of the gases. These include carbon dioxide, methane and nitrogen that trap the sun’s heat in the earth. It’s happened since the mid-twentieth century and its projected would be continuation. Global warming is an extremely severe problem facing the world today. Its effects and causes have been on the rise and people need to do something about it before the problem gets any worse.Global warming is an increase in the earths temperature, which can be caused by the use of fossil fuels, and industrial or agricultural processes.†¦show more content†¦The heat wave caused forest fires and the forest fires are also affecting the atmosphere due to heat and specially smoke, on the other hand the destruction of forests is initiating a cycle of heat, which will cause global warming and global warming will cause forest fire and so on to hold over toxic clouds in the air. Again the non believers would say this was just a natural occurrence. Believer would say global warming had a part in the weather patterns which has changed and caused this heat wave. The massive record-breaking snow storms in past five winters had a lot of the east Europe buried under large amounts of snow. Doubters of global warming just pointed out what they saw was just plain older Mother Nature doing her thing and the believers of global warming would say this was caused by man. Usually these unusual weather happenings tell us there are some climate changes happening already. For the past several years skeptics and believers alike have debated back and forth about the causes of global warming. The believers say that if the man-made causes are left untouched and steadily get worse these causes may not be able to be reversed. A few examples of these are: the rainforest reductions, depletion of the ozone, and the use of fossil fuels. Doubters claim that the warming of the earth is just a natural process and that the overall effect man has on nature is vastly overrated. TheShow MoreRelatedGlobal Warming Is The Greatest Challenge That Our Planet Essay1393 Words   |  6 PagesIntroduction: Global warming is the greatest challenge that our planet is facing today. 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