Qualitative aspect of drug action

Qualitative aspect of drug action Qualitative aspect of drug action Schild plot Schild plot: Schild plot is defined as pharmacological method of receptor classification. By using schild plot dose-effect curve for an agonist is determined in the presence of various concentrations of a competitive antagonist for its receptor in the presence of agonist i.e. equilibrium dissociation constant is calculated. The experiment is carried out for series of dose ratios for a given effect. For example the ratio of the dose of agonist (A) to produce a specific effect (e.g.,half maximal effect) in the presence of the antagonist (B) to the dose required in the absence of the antagonist (A) is calculated. This is determined for several doses of antagonist and then log ((A/A) -1) versus the negative log B is plotted. If the regression of log ((A/A) -1) on -log B is linear with a slope of -1, then this indicates that the antagonism is competitive and by definition the agonist and antagonist act at the same recognition sites. If the slope of the regression is not -1, then by defini tion the antagonist is not competitive or some other condition is in effect. This might include multiple binding sites or pharmacokinetic interactions. Agonist: Agonist is a drug which has both affinity and efficacy. Antagonist: Antagonist is a drug which has affinity and zero efficacy. Affinity:Affinity is a property of a drug; it measures how tight a drug binds to a receptor. To bind to a receptor a functional group of the drug should bind to the complementary receptor. The binding capacity of the drug defines the action of the drug. Efficacy: Efficacy of a drug can be defined as ability of drug which activates the receptor to produce desired effect after binding. Affinity and efficacy are explained in the equation as: K+1 ÃŽ ± A + R AR* Response K-1 ÃŽ ² K+1 B + R BR No Response K-1 Where A is agonist, B is antagonist, K+1 is association rate constant for binding, K-1is dissociation rate constant for binding ÃŽ ±- Association rate constant for activation ÃŽ ²- Dissociation rate constant for activation By using law of mass action affinity is explained as B + R BR Drug free receptor drug-receptor complex At equilibrium KB = [R] [B] KB = Equilibrium dissociation constant [BR] Hill-Langmuir equation: this equation explains drug occupancy [RT] = [R] + [BR] If [RT] = Total number of receptors then by substituting this in law of mass action equation [RB] = [B] [RT] KB + [B] By this equation it is determined that drug occupancy (affinity) depends on drug concentration and equilibrium dissociation constant. Equilibrium dissosciation constant: EQUILIBRIUM DISSOCIATION CONSTANT (Kd) : It is the characteristic property of the drug and the receptors. It is defined as the concentration of the drug required to occupy 50 % of the receptors. The higher the affinity of the drug for the receptors lower is the Kd value. Mathematically Kd is k2/k1 where k2 is the rate of dissociation of the drug from the receptor and k1 is the rate of association of the drug for the receptor. Receptor (R) and Drug (D) interact in a reversible manner to form a drug-receptor (RD) complex. Where R = Receptor D = Drug (L for ligand is sometimes used in these equations) k1 = the association rate constant and has the units of M-1min-1 k2 = the dissociation rate constant and has the units of min-1. k2 is sometimes written as k-1. If an agonist binds to the receptor, then the interaction of the agonist (D) and the receptor (R) results in a conformational change in the receptor leading to a response. If an antagonist binds to the receptor, then the interaction of antagonist (D) and receptor (R) does not result in the appropriate conformation change in the receptor and a response does not occur. For drugs that follow the law of simple mass action the rate of formation of the complex can be defined by the following equation d[RD]/dt refers to the change in the concentration of [RD] with time (t). Note: the square brackets refer to concentration. This equation indicates that the rate at which the drug receptor complex (RD) is formed is proportional to the concentration of both free receptor (R) and free drug (D). The proportionality constant is k1. The rate of dissociation can be defined by the following equation -d[RD]/dt is the decrease in drug-receptor complex with time This equation indicates that the rate at which the drug-receptor complex (RD) dissociates back to free drug and free receptor is proportional to the concentration of the drug receptor complex. The proportionality constant is k2. When the drug and the receptor are initially mixed together, the amount of drug-receptor complex formed will exceed the dissociation of the drug-receptor complex. If the reaction is allowed to go for a long enough, the amount of drug-receptor complex formed per unit time will be equal to the number of dissociations of drug-receptor complex per unit of time, and the system will be at equilibrium. That is equilibrium has occurred. Equilibrium can be defined as or k1[R][D] = k2[RD] This equation can be rearranged to give Kd is the dissociation equilibrium constant. Kd has units of concentration as shown in the following equation. Simple competitive antagonism: simple competitive antagonism is the most important type of the antagonism. In this type of antagonism the antagonist will compete with available agonist for same receptor site. Sufficient antagonist will displace agonist resulting in lower frequency of receptor activation. Presence of antagonist shifts agonist log dose response curve to right. A schild plot for a competitive antagonist will have a slope equal to 1 and the X-intercept and Y-intercept will each equal thedissociation constantof the antagonist. This can be explained in equation as: Occupancy for agonist [RA] = [A] OR [A]/ KA [RT] KA+ [A] [A]/ KA +1 In presence of competitive antagonist (B) [RA] = [A]/ KA [RT] [A]/ KA + [B]/ KB + 1 Occupancy reduced according to [B] and KB To obtain same occupancy, must increase [A] to [A`] r = [A] / [A] = [B] / [B] Schild equation: r = [B] / KB +1 Where r depends on [B] and KB Applying log on both sides log (r-1) = log[B] log KB Aim: The main aim of the experiment is to measure the equilibrium dissociation constant (KB) for atropine at acetylcholine muscuranic receptors and to determine the drug receptor interactions. Objectives The main objectives of the experiment are as follows Ø To measure the equilibrium dissociation constant for atropine at acetylcholine muscuranic receptors Ø To demonstrate the reversible competitive antagonism of atropine at acetylcholine muscuranic receptors Ø To determine the equilibrium dissociation constant (KB) for atropine at acetylcholine muscuranic receptors by using schild plot. MethodIsolation and mounting of Guinea-pig ileum in organ bath Guinea-pig was first sacrificed and then the ileum was collected and transferred into physiological salt solution maintained at 370C. The food particles present in the ileum was expelled out through running Krebs solution through the lumen. Then tissue was tied with a thread at both the ends where one was tied to the mounting hook and the other was attached to the transducer. 1) Preparation of serial dilutions of drug The drugs used in the experiment were acetylcholine (Ach) and atropine. To determine the simple competitive antagonism of atropine at Ach muscuranic receptors serial dilutions of Ach were carried out. Ach was given as 110-2M and from the above concentration of the drug the following concentrations were prepared to the organ bath concentration such as 110-6M, 310-6M, 110-7M, 310-7M, 110-8M, 310-8M, 110-9M and 310-9M Ach. Then atropine was diluted to 110-8M (organ bath) from the given 110-2M concentration. 2) Determination of Organ bath concentration The volume of physiological salt solution (pss) was 20 ml, and each time the volume of drug introduced into organ bath was 20Â µl.Therefore if 20Â µl of 110-2M drug was introduced into the organ bath then it gives 110-5M organ bath concentration. Mathematical calculation of organ bath concentration: In organ bath we have 20ml of pss which is equal to 20103 Â µl of pss, if 20 Â µl of 110-2 M Ach was introduced then the organ bath concentration 20Â µl→XM 20ml→10-2M = 20 Â µl x 10-2 M 20x 103 Â µl = 110-5M (organ bath concentration). The isolated guinea- pig ileum was mounted onto the organ bath and set up for recording isometric tension of the tissue using chart software in a Mac book. Step-1 Calibration of the experimental apparatus: The chart 5 software was calibrated and the sampling rate was adjusted to 10 samples per second with a maximum input voltage to 10 mV. The baseline was set to zero and then trace was started from the baseline zero then the force transducer was calibrated by placing 1 gram weight and after the calibration the trace produced was stopped for the moment to convert the units of tension into grams by selecting the trace produced previously. Step-2 Sensitisation of preparation: To check the viability of the tissue a response of suitable height was obtained by adding a little high concentration of the drug. Here in the experiment an appreciable recording was noted at 110-7M Ach. Step-3 The time cycle followed to construct a concentration- response curve was 0 seconds to add the drug concentrations 30 seconds to empty the organ bath and refill with fresh physiological salt solution 180 seconds next drug concentration was added to the organ bath. Concentration Response Curve: By making use of the above drug concentrations a concentration response curve was constructed according to the provided time cycle. 1) 20 Â µl of 110-9M Ach was added into the organ bath at zero seconds at is allowed to stand for 30 seconds, then after 30 seconds the organ bath was emptied and refilled with pss. Pss was allowed to stand for 180 seconds. During the wash period if the peak does not return to the base then it was washed twice or thrice to make sure that all the drug dissociates from the receptors before the next addition of the other drug concentration. Each concentration was repeated twice or thrice until the two consecutive responses were reported with the same peak height. 2) By following the procedure and time cycle, the concentration response curve was constructed with different concentrations of acetyl choline such as 110-9M,310-9M, 110-8M, 310-8M, 110-7M, 310-7M, 110-6M and 310-6M Ach (organ bath concentration). Step-4 Equilibration of Acetylcholine receptors with acetylcholine After step-2 the preparation was washed several times until the peak returned to the base line. Then atropine (110-8M organ bath concentration) was added to the preparation and then set aside for 40 minutes to allow atropine to equilibrate with acetylcholine muscuranic receptors. Step-5 Concentration response curve in the presence of atropine The concentration response curve with acetylcholine was repeated again in the presence of atropine by following the time cycle and procedure, which was same as same step 2.Therefore in step 3 with each addition of acetylcholine concentration atropine was added simultaneously. Step-6 Analysis: i) The graph pad prism in the Mac book was used to plot concentration response curves in the absence and presence of atropine. Log concentration (acetylcholine) Vs response in grams ii) From the above plot EC 50 values of acetylcholine in the presence and absence of atropine were obtained. Then the distance between the two curves control and response for the atropine presence was denoted by ‘r, where ‘r was called as shift. iii) The shift was calculated mathematically as r= EC 50 of response in the presence of atropine EC 50 of Ach in the absence of atropine iv) From the value of the shift, schild plot was plotted as log concentration of atropine presence against log(r-1). v) From the schild plot the dissociation constant KB for atropine at acetylcholine muscuranic receptors was determined. Results: As explained above in the procedure serial dilutions of acetylcholine was added to the organ bath, where Ach has produced concentration dependent contractions of the guinea pig ileum as shown in the fig 1. As shown in 1 the serial dilutions of acetylcholine are added into the organ bath from 110-7M to 310-6M Ach. Here in the trace it was clearly shown that contractions produced by the acetylcholine have been increased with respect to the concentrations. In step-2 the preparation was washed and added with 110-8M atropine and set aside for 40 minutes to equilibrate the acetylcholine receptors. In the trace it is clearly shown that, the contractions produced by serial dilutions of Ach from 110-8M to 310-4M in the presence of 110-8M atropine. When Trace 1 and Trace 2 are compared it is evident that the contractions produced by Ach alone (trace 1) were greater than the contractions produced Ach in the presence of atropine (trace 2) which proves the simple competitive antagonism by atropine at muscuranic receptors. A graph is plotted to the log concentration response curve produced by Ach alone against Ach in the presence of atropine. (graph is attatched to the report) From the graph it is known that with the increase in the concentration of Ach, response have been increased when compared to Ach in the presence of atropine and also there is a shift towards right which shows the simple competitive antagonism produced by atropine. From the results produced by Ach alone against Ach in the presence of atropine the fractional difference which is called as shift can be obtained as follows Mathematical Calculation shift ‘r = EC50 of response after atropine (or) in the presence of atropine EC50 of control (or) Ach in the absence of atropine = 2.5110-6 = 8.36 3.0 x10-7 r-1 =8.36 -1=7.36 log(r-1)=log (7.36) =0.86 Partial dissociation constant (PKB) or PA2 is measured to confirm the simple competitive antagonism, where pKB values play an important role in classifying receptors. Therefore PKB =log(r-1) -log [atropine] =0.86 -log (110-8) =0.86 (-8) =0.86+ 8 =8.86 From the above results log EC50 values for control (Ach alone) and Ach in the presence of atropine were given as 3.0e-007 and 2.51e-006 respectively. This shows the molar concentration of Ach which produces 50% of the maximal possible response is higher than the molar concentration response produced by Ach in the presence of atropine. If the antagonist is competitive, the dose ratio equals one plus the ratio of the concentration of antagonist divided by its Kd for the receptor. (The dissociation constant of the antagonist is sometimes called Kb and sometimes called Kd) MathType Equation A simple rearrangement gives: MathType Equation Here we have plotted a graph with log (antagonist) on the X-axis and log (dose ratio -1) on the Y-axis. If the antagonist has shown simple competitive antagonism then the slope should be 1.0, X-intercept and Y-intercept values should be both equal the Kd of the antagonist obtained. If the agonist and antagonist are competitive, the Schild plot will have a slope of 1.0 and the X intercept will equal the logarithm of the Kd of the antagonist. If the X-axis of a Schild plot is plotted as log(molar), then minus one times the intercept is called the pA2 (p for logarithm, like pH; A for antagonist; 2 for the dose ratio when the concentration of antagonist equals the pA2). The pA2 (derived from functional experiments) will equal the Kd from binding experiments if antagonist and agonist compete for binding to a single class of receptor sites. From 5 and 6 it is evident that no concentrations of atropine have showed competitive antagonism perfectly. Therefore from the above results it is known that the concentrations of atropine has not shown simple competitive antagonism fairly. Discussion: Reversible competitive antagonism: The binding of drug to a receptor is fully reversible which produces a parallel shift of the dose response curve to the right in the presence of an antagonist. The mechanism of action of acetylcholine at muscuranic receptors: In various gastrointestinal smooth muscles, acetylcholine and its derivatives produce contractions by activating muscuranic receptors. It is generally assumed that the M3 muscuranic receptor plays a key role in mediating this activity. The M3 receptor is coupled preferentially to Gq-type G proteins, resulting in the activation of phospholipase C (PLC) and the formation of ionositiol trisphosphate (IP3) and diacylglycerol (DAG) which are likely to participate in muscuranic receptor-mediated smooth muscle contractions. IP3 causes Ca2+ release from intracellular store and can also mobilize Ca2+ secondarily through Ca2+-sensitive or store-dependent mechanisms. DAG, via activation of protein kinase C, phosphorylates various proteins and can directly activate non selective cationic channels. From the above results the value of shift obtained was 0.378 which denotes the simple competitive antagonism produced by the concentration of atropine used (110-8 M).From the value of shift the pKB value was calculated as 8.4.If atropine has shown simple competitive antagonism then the value of pKB should be equal to 1-X intercept. Therefore pKB=1-X intercept =1-(-8.86) =9.86 We got value of pKB as 8.86.Therefore pKB is not equal to 1-X intercept. Therefore the concentration of atropine (110-8M organ bath concentration) used by our group has not shown simple competitive antagonism effectively. The literature value of pKB is given as approximately 9 and we have obtained the value of pKB as 8.86 which does not fit with literature value. Therefore from the above observations and results i can conclude that a little more high concentration of atropine may serve to produce complete simple competitive antagonism by atropine at acetylcholine muscuranic receptors.

Sir Elton John

Sir Elton John was a pop singer in the 19 and 20 century. Elton was inducted in the Rock and Roll Hall of Fame in the year 1994. Elton was one of the biggest artist of that time. Elton has lots and lots of fans and makes over millions of dollars. This is Elton life from a child to an adult! Elton John had an ok childhood. Elton John was born on March 25 1947 in Pinner, United Kingdom. Elton John had 4 brothers named Geoff, Simon, Sam and Robert. Elton John's father, Stanley Dwight was in the royal air force. Elton did not have a good relationship with Stanley. Elton John not only loved music at a young age, he loved to play sports. One surprising fact about Elton John was that he taught himself how to play the piano at age 3! When Elton John was only 17 he dropped out of High School to start his dream of music. According to CNN, Elton John was married to Renate Blauel in 1984. But CNN states that Elton got into a divorce with Renate in 1988. Elton John went on Instagram and said that he was a bad husband to Renute and caused her into sadness, which made them have a divorce. Years later Elton ended up marrying David Furnish to be his long time partner. They have been married to David since 2014 to now. Elton John has too kids with David. Elton John has two kids named Zachary and Elijah! Zachary is Eltons older son that is about 8 years old as of 2018. Zachary was born in 2010 on christmas day! A surprising fact about Zachary is that his godmother is Lady Gaga! In a interview Elton said â€Å"He's gorgeous, he travels brilliantly, he so loves people, and it makes our lives – he's the icing on the cake†. Elton and David say they love the biological mother of Zachary like a sister. Elijah is Elton and David's youngest son. As of 2018 Elijah is about 5 years old. Elijah was born on January 11, 2013. At Elijah's birth he weighed 8lb 4oz. From interviews you can tell that Elton and David really loves there kids and would do anything for them. Elton John's has many songs that made him famous. Some of his songs that got him very famous are Rocket Man, Crocodile Rock and Goodbye Yellow Brick Road! Those are just a few of Elton John's hits singles. Elton John is also famous for his debut album Yellow Brick Road of 1969-1973. This album has helped make Elton John very famous and put him where he is today. Elton John makes about 500 millions dollars! Listen to this song by Elton called Rocket Man. (played from the slides) Elton John's is getting into his oldern days now. Elton john is 71 years old and is retiring from music very soon. Elton John is going on his very last tour soon. He will be calling it the † Farewell Yellow Brick Road tour†. From the title of the tour he may be performing songs from his album Yellow Brick Road! Rumors say that Elton wants to take a break from music and move on to his other passion, sports! As you can see Elton John is a very successful man. He is very lucky to get inducted into the Rock and Roll Hall of Fame. Even with an ok childhood, he is able to still have the passion for sports and music. Elton is a 71 year old man that is very successful that will go on the rest of his life not being worried about anything.

Analysis Of Mary Shelley s Frankenstein - 1589 Words

Extended Response (Q2) - Frankenstein By Mary Shelley Rachel .Corrie The perspective, from which a story is told, causes an influential response from readers to certain issues, characters and conflicts that are found in literary texts. Mary Shelley’s gothic novel, Frankenstein, was published in 1818 and tells the story of a scientist known as Victor Frankenstein who reanimates life in an unethical science experiment. In this novel Walton, Victor and the creature tell their side of the story, through which Mary Shelley uses the effect of a frame narrative so that it provides readers with extensive information about characters such as their intentions, emotions, and thoughts, which allows for each reader to create a unique and individual response to the novel. Robert Walton is an indirect narrator of the story, he tells Victor Frankenstein s story through letters to his sister, Margaret Saville. Through the letters, Walton is able to tell Frankenstein’s story about his creation, although the letters are written from Walton about Frankenstein it shows the personal connection between the two characters. When Walton first meets Frankenstein, readers can see why he wants to share the legacy of Frankenstein as he describes him to be gentle wise, intelligent, and that Walton is â€Å"happy to have possessed as the brother of his heart†. By placing these letters, in the beginning of the story, it becomes intriguing and exciting for readers as they explore the storyShow MoreRelatedAnalysis Of Mary Shelley s Frankenstein1411 Words   |  6 PagesIn the early 1800s Mary Shelley set pen to a paper and started to develop a novel that little to her knowledge would become world renowned. In 1818 she finished and published the novel to sel l to the European public. The novel caught the world off guard in the way that a female was able to write about such harsh, dark, and evil things in a European society whose authors like John Locke and Charles Montesquieu preached enlightenment, self exploration, and individualism all in an optimistic enablingRead MoreAnalysis Of Mary Shelley s Frankenstein1615 Words   |  7 Pagesa whole and how accurate a depiction they might think it to be, they will miss out on many of the qualities of the painting that reside below the immediately apparent surface level. Mary Shelley’s Frankenstein is a text dedicated to expounding upon the dangers of such superficial analysis. In Frankenstein, Mary Shelley openly condemns the surface level and appearance oriented methodology under which the human mind operates. The very protagonist of the novel is inspired solely by reputation and howRead MoreAnalysis Of Mary Shelley s Frankenstein 1758 Words   |  8 PagesFrankenstein was published over 200 years ago. 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The Unnecessary Destruction of Hiroshima - 598 Words

On August 6th, 1945, at 8:15 Japan time, the United States dropped an atomic bomb named â€Å"Little Boy† on Hiroshima, Japan. Little Boy was dropped from the Enola Gay, from 31,000ft and detonated at 1,900ft above Hiroshima. Dropping the atomic bomb destroyed the city, ruined and negatively changed people’s lives, and killed thousands of people. Some say it was necessary to drop it because of the lives it saved. It didn’t save lives. It was unnecessary to drop the atomic bomb because of the amount of destruction and lives it ruined. The atomic bomb caused a lot of unnecessary destruction when it was dropped on Hiroshima. It had destroyed houses and work buildings in Hiroshima, such as the hospital in which Dr. Fujii lived and worked. Dr. Fujii’s hospital was smashed â€Å"all around him in a mad assortment of splintered lumber and materials for the relief of pain† (John Hersey, 11). Buildings were blown apart, caught fire, and completely obliterated. Two-thirds of Hiroshima were demolished. Out of the 90,000 buildings in Hiroshima, 60,000 buildings were destroyed within a three mile radius of the epicenter. The havoc could have been avoided by not dropping the atomic bomb. Many people had their lives unfairly changed and even ruined from the atomic bomb. The victims of radiation poisoning and burns had to deal with disfiguring ailments. They had dark burns across their bodies. Their hair burned off. Skin drooped from the radiation and blast. The bomb also irradiated the people nearbyShow MoreRelatedThe Atomic Bomb Of Hiroshima And Nagasaki1364 Words   |  6 PagesJapanese city, Hiroshima. Three days later on the 9th of August America dropped another bombed called, ‘Fat Man’ on the Japanese city of Nagasaki. A surrender was received and accepted by America on the 15th of August and the war against Japan had ended. Harry S. Truman, the man responsible for dropping the bombs claims it ended the war more efficiently and was in fact the best option but many suspicions arose as news from Japan came to light and the utter destruction of Hiroshima and Nagasaki wasRead More Drop The Bomb? Essay656 Words   |  3 Pages U.S. History - Grech Atomic Bombs Dropped on Japan, Justified? nbsp;nbsp;nbsp;nbsp;nbsp;On August 6th and 9th of 1945 U.S. bombers dropped atomic bombs on the Japanese cities of Hiroshima and Nagasaki, causing utter destruction and many deaths. These bombs were dropped as the Pacific battles of World War II were coming to an end. Soon after Japan surrendered, ending the war. But, was the use of atomic warfare necessary? Was it too harsh and cruel to the JapaneseRead MoreUnited States Should Not Drop The Nuclear Bombs On Japan.1369 Words   |  6 Pagesweapons on the Japanese cities Hiroshima and Nagasaki respectively. Those two bombs brought severe damages to these two cities including over 200,000 innocent lives. Many people believe that Japan deserve to be bombed due to many reasons, primarily because Japan first attacked United States at Pearl Harbor. However, it is still wrong and unnecessary for United States to drop two bombs respectively on Hiroshima and Nagasaki as the two bombs not only bring destructions to Japan, but also cause furtherRead MoreThe Atomic Bombing Of Hiroshima871 Words   |  4 PagesAn Unnecessary Action The atomic bombing of Japan was an unjustified decision that many considered was inhumane. Through the use of atomic bombs, many innocents were harmed in one way or another. Berger described the atomic bombing of Hiroshima as a terrorist act and that it was evil. While Berger s argument was extreme, the U.S. still remains unjustified because they did not know exactly how much damage the bomb would actually cause. Bombing Japan, the U.S. harmed thousands of civiliansRead MoreHarry Trumans Decision to Drop the Atomic Bomb1314 Words   |  6 Pagesbombs on Hiroshima and Nagasaki justified? A. Plan of Investigation Research Question: To what extent was Harry Truman’s decision to drop atomic bombs on Hiroshima and Nagasaki justified? This investigation assesses President Harry Truman’s decision to drop atomic bombs on both Hiroshima and Nagasaki. It will determine whether or not his decision was justified. This investigation will scrutinize the reasons that made Harry Truman feel inclined to drop atomic bombs over Hiroshima and NagasakiRead MoreThe Tragedy Of Hiroshima And Nagasaki1422 Words   |  6 PagesPapas 1 Hiroshima â€Å"For those who were there and who survived to recall the moment when man first turned on himself the elemental forces of his own universe, the first instant was pure light; blinding intense heat ... if there was a sound no one heard it. 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