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  1. 1. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedBiochemistry II - examinationGENERAL MEDICINEDENTISTRYKhuram AhmedReband AhmedGeneral Medicine 4th semester 2009
  2. 2. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed1. Factors influencing results of laboratory examination (three phases of examination, biological and analytical factors, sample collection and handling of samples, interpretation of results, reference interval and its calculation, critical difference).Biological factors can influence the results of labority examinations. Body Weight can affect the concentration ofsome analytes, by changing their distribution volumes. The serum concentration of cholesterol, LDL-cholesterol,triacylglycerols, uric acid, insulin and cortisol positively correlates with obesity. Exercise can effect blood compositionvalues depending on the duration and intensity, and the physical condition of the patient. Exercise causes a reductionof cellular ATP which increases cellular permeability, leading to increases in serum activites of enzymes an metabolitesoriginating from skeletal muscles. Smoking may affects the level of many analytes by nicotine. Smoking increases theconcentration of cholesterol and triacylglycerol. Alcohol affects mainly the metabolism of glucose, and it increasesliver enzymes in blood. Stress affects production of hormones. Environmental factors include altitude, ambienttemperature and geographical localization.Analytical factors determine the closeness of the measured value to the true value. Precision is the ability of ananalytical method to produce the same value for replicate measurements of the same sample, i.e. agreementbetween two independant test results. Trueness is the closeness of agreement between the average value from alarge series of test results and an accepted reference value. Accuracy is closeness between the result of ameasurement and an accepted reference value.Sample collection involves many reccomendations, the patients are not allowed to eat 10-12 hours before bloodcollection. They have to exclude fat food and alcohol from their diet. Patients can drink ¼ of a litre of water in themorning before the blood collection. Type of blood collected depends on the test ordered, some specimens must becollected in tubes which have anticoagulants. Time of collection is important because concentration of somesubstances vary throughout the day. Blood collection is usually performed in the morning. Haemolysis can occur ifthere is rough handling of the sample, use of incorrect sized needle, moisture in the test tube, or centrifugation athigh speed. Transport should be carried out with blood samples at 0c, which is the temperature of thawing ice.Interpretation of results is most frequently carried out by the comparision with the reference interval. Referencevalues are required from healthy individuals and patients with relavant diseases. Reference interval includes 95% ofresults of a reference group. 5% of the results are not included (2.5% of the higest values and 2.5% of the lowestvalues). Critical difference is expressed as statistically significant difference between the two results of a givenlaboratory test measured in an individual between the giventime interval. The difference reflects the change in clinicalstate of the patient.2. The significance of (both functional and non-functional) enzyme assays in blood serum. Isoenzymes - multiple forms of LD and CK.Enzymes in blood:TYPE EXAMPLE AFTER ORGAN DAMAGE, ACTIVITY WILLPlasmatic co-agulation factors decreaseSecretory amylase, lipase increaseIntracellular ALT increase Indirect determination involves calculating catalytic concentration (ukat/l), the product of enzyme reaction is determined. It is used for most enzymes such as ALT and AST. Direct determination involves mass conc (ug/l), enzyme molecules are determined as antigens. It is used for a few enzymes, e.g. PSA. Isoenzymes are genetically determined differences in the primary structure. They catalyse the same reaction. They may have different subcellular or tissue distribution. They are usually determined by electrophoresis. Elevated blood values are a specific marker of tissue damage.General Medicine 4th semester 2009
  3. 3. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedLD – Lactate DehydrogenaseLactate + NAD(+) > Pyruvate + NADH + H(+)Is a tetramer (protein with four subunits), with two different chains (H= heart, M=muscle). It has five isoenzymes, withdiffering composition of chains: (H4) (H3M) (H2M2) (H1M3) (M4).  LDH-1 and LDH-2 are markers of myocardial infarction. Usually LDH-2 is predominant in serum. A LDH-1 level higher than the LDH-2 level suggests myocardial infarction.  LDH-3 is a marker of lung embolia.  LDH-4 and LDH-5 mark skeletal muscle diseases.CK - Creatine KinaseIs a dimer with two chains (M=muscle, H=heart). It has three isoenzymes, CK-MB, CK-BB, and CK-MM. CK-MB is themajor isoenzyme in blood. CK-MB is a marker of myocardial infarction.3. Provision of glucose in different states, the factors increasing susceptibility of glucose (glucagon, adrenaline, cortisol). Glucosuria.Glucose is the most common monosaccharise, C6H12O6. Its chemical energy is 17kj/g. Its a fuel source for tissues,especially the brain and erythrocytes. The source of glucose in blood is from dietry saccharides, gluconeogenesis, andglycogenolysis.Feature I II III IV VStage well- prolonged extreme post resorption early starvationdescription fed starvation starvationTime a 0-4 h 4-16 h 16-30 h 2-24 d over 24 dintervalOrigin of liver glycogen gluconeogenesisGlc in food gluconeogenesis gluconeogenesis gluconeogenesis liver glycogenblood b b all tissues all tissuesUtilization all brain, Ercs, Ercs, kidney, muscle, ad.t. muscle, ad.t.of Glc tissues kidney brain - limited limited limitedEnergy for Glc, ketone ketone bodies, Glc Glc Glcbrain bodies GlcGlucagon binds to receptors in liver, it activates adenylate cyclase, which increases cAMP, this activates cAMPdependant protein kinase A which leads to glucogen phosphorylation.Cortisol increases blood sugar in response to stress. Substrates from proteolysis in muscle are used ingluconeogenesis. It is also an inducer of enzymes in gluconeogenesis.Adrenaline secretion is a response to acute stress. It is involved with the breakdown of glycogen in the liver andmuscles. Also increases glycolysis in muscles.Glucosuria is when glucose concentration in urine is higher than 0.8mmol/L. Glucosuria is the excretion of glucose intothe urine. Ordinarily, urine contains no glucose because the kidneys are able to reclaim all of the filtered glucose backinto the bloodstream. Glucosuria is nearly always caused by elevated blood glucose levels, most commonly due tountreated diabetes mellitus.4. The basic metabolic disorder in diabetes mellitus: the cause of ketoacidosis or of hyperosmolarGeneral Medicine 4th semester 2009
  4. 4. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed coma.Elevated blood glucose is due to lack of insulin => few insulin-dependant Glut-4 transporters => which enables glucoseto enter muscle cells or adipose tissue.Elevated FFA is due to excess glucagon => which leads to increased lipolysis. (FFA in blood are bound to albumin)Elevated TAG is due to lack of insulin, which means there isn‘t enough Lipoprotein Lipase, LPL (insulin is inducer of it’ssynthesis).KETOACIDOSIS is a state of elevated concentration ketone bodies. It is due to excess of FA from lypolysis, B-oxidationof their carbon chains gives Acetyl CoA. Acetyl CoA is a precursor for synthesis of ketone bodies in the liver.HYPEROSMOLAR COMA is when extreme hyperglycemia and dehydration are sufficient to cause unconciousness.Diabetes Mellitus 1:Due to defficiency of insulin caused by autoimmune attack on B-cells of pancreas. Leads to hyperglycemia,ketoacidosis and hypertriglyceridemia.Diabetes Mellitus 2:This is genetic, and is due to resistance to insulin. It decreases the ability of target cells (liver, muscles, etc) to react toinsulin.5 Lipids in blood plasma and the major classes of lipoproteins (differences in the lipid and apolipoprotein content, in size, in properties and in electrophoretic mobility, the origin in enterocytes and hepatocytes).Lipids in blood:Cholesterol (free and esterified) 5mmol/lPhospholipids 2.5mmol/lTriacylglycerols 1.5mmol/lFree Faty Acids 0.5mmol/lClasses of Lipoproteins: (increasing density, decreasing size)Chylomicrons 85% TAGVLDL 50% TAGLDL 50% cholesterolHDL 50% protein Lipoproteins consist of a polar surface monolayer (phospholipids, free cholesterol, apoprotein) and a non- polar core (triacylglycerol, cholesteryl ester).LIPORPOTEIN: ORIGIN: TRANSPORT:Chylomicrons Enterocyte Exogenous TAG from GIT --> tissuesVLDL Liver Endogenous TAG from liver -- > tissuesLDL Blood Plasma Cholesteryl ester --> tissuesHDL Liver Free cholesterol --> liverCM contains predominantly TAG = neutral molecules (without charge)  They do not move in electric field6 Transformation of chylomicrons and VLDL.General Medicine 4th semester 2009
  5. 5. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedChylomicrons are produced in enterocytes, via Apo B48. It is secreted into the lymphatic system and joins the bloodvia the thoracic duct. Chylomicrons carry dietry TAG to peripheral tissues. In plasma chylomicrons recieve Apo E andApo C11 from HDL. Apo C11 activates LPL (lipoprotein lipase). LPL is attached to capillary surface in adipose, cardiac,and muscle tissue. Triacylglycerol is hydrolysed to FFA and gycerol. Apo C11 is returned to HDL. Chylomicron particlesbegin to shrink, remnants bind to APO E receptors in the liver where they are degraded in lysozymes.VLDL is produced in the liver, it transports endogenous TAG from the liver to peripheral tissues.In plasma they takeApo-C11 from HDL. Triacylglycerol is removed by LPL action. VLDL becomes smaller and more dense, it becomes IDL.IDL takes up cholesteryl ester from HDL and becomes LDL by hepatic lipase.7 Metabolism of high-density lipoproteins.General Medicine 4th semester 2009
  6. 6. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedHDL particles are made in the liver. Nascent HDL are disk shaped (bilayer of phospholipd and proteins). HDL take freecholesterol from cell membranes. Once cholesterol is taken up it is esterified by LCAT (which is made in the liver andactivated by Apo A-1). HDL becomes spherical. Spherical HDL is taken up by the liver and cholesteryl esters aredegraded.Cholesterol + Lecithin > Cholesteryl Ester + Lysolecithin8 The movements of cholesterol and its elimination. The balance of sterols and the bile acids transformation.Blood cholesterol is 5mmol/l. Its source is from food (fish, eggs, mayonaise) or biosynthesis from Acetyl CoA (incytoplasm). A small amount of cholesterol is incorporated into the cell membrane. Some is converted into hormones(steroid hormones). Some is converted into bile acids in the liver. Free cholesterol is immediatedly esterified by ACAT(Acetyl CoA Cholesterol Acyl Transferase) to esterified cholesterol. Cholesterol is eliminated in bile/bile salts.Intracellular cholesterol descreases HMG-CoA reductase (used for cholesterol synthesis), it decreases synthesis of newLDL receptors (to block LDL intake), and it enhances activity of ACAT (to help make storage).9 The metabolic interrelationships among body organs predominating in a well-fed state (absorptive phase).General Medicine 4th semester 2009
  7. 7. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedAfter a typical high saccharide meal, glucose leaves the intestine in high concentrations. Hyperglycemia stimulates thepancreas to release insulin, glucagon release is inhibited. Part of the nutrients are oxidized to meet the immediateenergy needs, exessive nutrients are stores as glycogen in liver and muscle, and as TAG in adipose tissue.During hyperglycemia, GLUT-2 transporters facilitate diffusion of glucose in to B-cells. ATP produced by glycolysiscloses the ATP-dependant K+ channel, the resulting depolarization opens voltage-gated Ca2+ channels, and increasesthe intracellular Ca2+. This is followed by exocytosis of granules containing insulin.Insulin inhibits secretion of glucagon. It supports the entry of glucose into muscle and adipocytes by GLUT-4transporters. It promotes glycogen synthesis and storage in the liver and muscle. It inhibits glycogen breakdown. Itstimulates glycolysis, and intensifies TAG synthesis in the liver.10 The metabolic interrelationships among body organs predominating after a brief fast (post- absorptive phase) and during prolonged fasting (starvation).Post-absorptive phase (early starvation):The post-absorptive phase is the time period from the first feeling of hunger, it doesn’t last more than 10-12 hours.Within one hour after a meal, blood glucose concentration declines. Release of glucagon from A-cell begins, andstimulation of insulin discontinues.Glycogen antagonises the effects of insulin:- stimulates liver glycogenolysis (inhibits glycogenesis)- supports gluconeogenesis from lactate, glycerol and amino acids- activates mobilization of fat stores  has no influence on skeletal muscle metabolism  results in maintaining fuel availability in absence of dietry glucoseGluconeogenesis occurs 90% in the liver, and 10% in the kidneys. It can be from lactate, glycerol or amino acids.General Medicine 4th semester 2009
  8. 8. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedGlycogenolysis = Glycogen > Glucose-1-phosphate > Glucose-6-phosphate > Free glucoseFatty acids act as a fuel for muscles: FFA > Acetyl CoA > Citric Acid Cycle > CO2 and energy. They come from hydrolysisof TAG by HSL (hormone sensitive lipase). They can be used for ketogenesis in the liver (a fuel for muscles/brain).Prolonged fastning (startvation):The prolonged fasting phase’s major goal is to spare glucose and to spare proteins. Tissues use less glucose, they useTAG and KB for energy instead. The brain consumes acetoacetate (30-60%) in place of glucose. After a while, KB arenot utilized in the muscles, they are saved to be used up in the brain. Sources of proteins are: intestinal epithelium,digestive enzymes, liver enzymes, and skeletal muscle contractile enzymes.11 Proteins in human nutrition, the biological value of proteins, nitrogen balance and simple methods for assessing the catabolic periods.General Medicine 4th semester 2009
  9. 9. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedFood proteins, tissue protein proteolysis, and synthesis of non essential amino acids > AMINO ACID POOL.The amino acid pool has 3 main uses:1. Synthesis of specialized nitrogenous products2. Synthesis of tissue/plasma proteins3. Deamination and utilization of carbon skeletonDigestion of proteins:Stomach: PepsinSmall Intestine: Trypsin, Chymotrypsin, Elastase, Carboxypeptidase A/B, aminopeptidaseGASTRIN is secreted by the stomach. SECRETIN is from pancreatic juice. CCK is a product of pancreatic enzymes.Endogenous protein degradation is by two methods; lysosomal or ubiqitin proteosome. Lysosomal is non-specific, noATP is required, and is for extracellular and membrane proteins. Ubiquitin proteosome requires ATP, and is fordamaged or regulation proteins.Biological Value: relative amount N used for endogenous protein synthesis from total N absorbed from foodEgg White 100%Whey Protein 100%Milk Cassein 80%Beef 80%Beans 49%Wheat Four 54%Gelatin 25%Conversion of amino acids after a meal Glutamate and glutamine are metabolic fuel for the enterocyte In the liver, AA are utilized for synthesis of proteins, glucose, and fatty acids Valine, Leucine and Isoleucine are not metabolised in the liver due to lack of aminotransferase; predominate in blood High content of NH3 in portal blood is removed by the liver by urea synthesis and is excretedCatabolic Pathway of NitrogenDietry proteins > AA in GITTransamination of AA in cells > GlutamateDehydrogenation deamination of glutamate > NH3Detoxifying NH3 > ureaNitrogen balance – the state of protein nutrition can be determined by measuring the dietary intake and output ofnitrogenous compounds. N balance = Nin – NoutThree states are distinguished:1. Nitrogen balance in equilibrium intake = output2. Positive nitrogen balance intake > output (during childhood growth and pregnancy)3. Negative nitrogen balance intake < output (response to trauma or infection or inadequate intake for requirements,there is a net loss of protein.  Growth/Prenancy Positive Effect  Metabolic stress Negative Effect  Starvation Negative Effect  Incomplete food proteins Negative Effect12 The specific functions of the liver in metabolism, proteosynthesis, and in excretion. Uptake of most nutrients is from the GITGeneral Medicine 4th semester 2009
  10. 10. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed Intensive intermediary metabolism, conversion of nutrients Controlled supply of essential compounds (glucose, ketone bodies, plasma proteins, etc) Ureosynthesis Biotransformation of Xenobiotics Excretion (cholesterol, bilirubin, hydrophobic compounds, some metals)Metabolism of saccharidesPrimary regulation of blood glucose concentration – via the glucose buffer functionUptake of glucose and storage as glycogenOR initiation of glycogenolysis and gluconeogenesisMetabolism of lipidsCompletion and secretion of VLDL and HDLKetogenesis produces ketone bodiesSecretion of cholesterol and bile acids into bile (cholesterol elimination)Metabolism of Nitrogenous compoundsDeamination of amino acids in excess of requirementsProteosynthesis of plasma proteins and blood-clotting factors – zone 1 periportal areaUptake of ammonium for ureosynthesis – zone 1 periportal areaBilirubin capturing, conjugation, and excretion  Detoxification of drugs, toxins, and excretion of some metals. Transformation of hormones – inactivation of steroid hormones, inactivation of insulin. 13 Ammonium transport, the glutamine cycle and the glucose-alanine cycle.NH3 in portal blood from: protein putrefication in GIT demaination of Gln/Glu in enterocytes In saliva from: hydrolysis of urea by oral microfloraIn venous blood from: catabolism of AA in tissues In urine from: hydrolysis of GlnGlutamine in MuscleProduced by proteolysisA product of ammonia detoxificationCarrier of NH2 group to liver where NH3 is liberatedGlutamine in enterocyteSource of energy for intestinal mucosa (Gln> 2-OG > CAC)Limited usage of glucose and fatty acids as fuel in enterocytesGlutamine in brainFormation of glutamine is a way of amonia detoxificationSynthesis occurs mainly in astroglial cellsGlutamate decarboxylation gives GABAGLUTAMATE + NH3 > (glutamine synthase / -H2O) > GLUTAMINEGlutamine in LiverPeriportal Hepatocytes: Source of ammonia for ureosynthesisPerivenous hepatocytes: a form of ammonia detoxification, released into blood to go to enterocytes and kidneysGlutamine in kidneysIs an energy sourceGlutamine and Glutamate release ammonium ions which makes the pH of urine acidicGeneral Medicine 4th semester 2009
  11. 11. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedMultiple functions of glutamineSynthesis of proteinsMetabolic FuelSource of nitrogen in synthesis of purines, pyrimidines, aminosugarsSource of glutamate for gaba synthesisSource of ammonium ions in urine14 Degradation of haemoglobin, formation of bile pigments.Erythrocytes are taken up by the reticuloendothelial cells by phagocytosis. These are cells of the spleen, bone marrowand Kupffer cells in the liver.Haemoglobin > (haem oxygenase) VERDOGLOBIN > (lose Fe3 and globin > BILIVERDIN > (biliverdin reductase)BILIRUBIN Conjugated bilirubin is secreted into the bile. As long as bilirubin remains in the conjugated form it cannot be absorbed into the small intestines. In the large intestines, bacterial reductases and B-glucouroniases catalyse the deconjugation and hydrogenation of bilirubin to mesobilirubin and urobilinogen. Urobilinogen is split into dipyrromethene and this condenses into intensively coloured BILIFUSCINS. Conjugated Bilirubin > (deconjugation/hydrogenation) > mesobilirubin and urobilinogen > dipyrromethens > bilifuscins 15 Metabolism and excretion of bile pigments. The main typesGeneral Medicine 4th semester 2009
  12. 12. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed of hyperbilirubinaemiaIn blood plasma, hydrophilic bilirubin is unconjugated and is transported as a complex with albumin. Unconjugatedbilirubin is non-polar. Hepatocytes convert it into a polar form by conjugation with glucouronic acid so that it may beexcreted. Glucosyluronate transferase on ER membranes add the glucouronic group to bilirubin. Conjugated bilirubinis polar and water soluble.Urobilinogens are partly excreted in the urin and partly excreted in the faeces. In air they are oxidised to a dark browncolour.Major types of hyperbilirubinaemia:Hyperbilirubinaemia > when serum bilirubin is 20-22umol/l/Icterus (jaundice) > when serum bilirubin is 30-35 umol/l/Causes of hyperbilirubinaemia:Prehepatic – increased production of bilirubinHepatocellular – due to inflammation or autoimmune diseasePosthepatic – insufficient drainage of intrahepatic or extrahepatic bile ducts16 Metabolism of iron (absorption, transfer and distribution in the body, functions, iron balance).Body contains 4-4.5g of Fe.Daily supply of iron in a mixed diet is about 10-20mg.From this, only 1-2mg are absorbed.There is no natural mechanism of eliminating excess in the body.Absorption of Iron in duodenum and jejunum: Ascorbate or fructose promote absorption aswell as Cu2+. Fe2+ is absorbed much easier than Fe3+. Gastroferrin (component of gastric secretion) is a glycoprotein that bings to Fe2+ maintiaing its solubility by preventing it from oxidising to Fe3+. Insoluble iron salts are formed from Fe3+. Phosphates, ocalate and phylate form insoluble Fe3+ complexes, this disables absorption.Transferrin:Is a plasma glycoprotein, serum concentration is 2.5-4g/l. Two binding sites for Fe ions. Biosynthesis of transferring isincreased during iron deficiency. Iron is taken up by cells through specific receptor-mediated endocytosis.Ferritin:One molecule can bing a few thousand Fe3+ ions. When it is not carrying iron it is called Apoferritin.It consists of 24protein subunits.Hepcidin:Is a hormone produced in the liver which limits accessibility of iron. Biosynthesis is stimulated in iron overload andinflammations. The same two factors stimulate hepcidin that inhibit transferin. It reduces absorption in theduodenum, inhibits Fe transport across placenta, and prevents release of recyclable iron from macrophages.17 Biochemical tests used for identification of liver injuries (detection of cell damage, cholestasis, reduced proteosynthetic capacity, etc.).General Medicine 4th semester 2009
  13. 13. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedPlasma markers of hepatocytes membrane integrity:Catalytic concentration of intracellular enzymes in blood increasesEnzyme assays of ALT is most sensitive (0.45-0.9ukat/l)Tests for decrease in liver proteosynthesis:Serum concentration of albumin, transthyretin, transferring and blood co-agulation factorsTests for excretory function and cholestasis:Serum bilirubin concentration is measuredSerum catalytic concentration of alkaline phosphatesTests for urobilinogen and bilirubin in urine18 The metabolism of xenobiotics - stage I of their biotransformation (various types of transformation, examples, mixed-function monooxygenases – function of cyt P450).Xenobiotics are hydrophobic (lipophilic) compounds present in the environment that cannot be used in normalbiological processes – they are foreign to the body. Their elimination depends on their transformation to morehydrophilic compounds. They are excreted in milk, urine, bile or sweat.Stage 1:The polarity is increased by adding a polar group (usually hydroxylation). Reactions usually take place on membranesof ER, or in the cytoplasm. The first stage may convert the xenobiotic into a more biologically active compound.Types of biotransformationsHydroxylation (aromatic systems)Dehydrogenation (alcohols, aldehydes)Sulfooxidation (dialkyl sulfides (to sulfoxides)Reduction (nitro compounds (to amines))Hydrolysis (esters)  The overall purpose of the biotransformation of xenobiotics is to reduce their nonpolar character as far as possible. The products of transformation are more polar, many of them are soluble in water. Their excretion from the body is thus facilitated.Monooxygenases:Catalyse reactions of stage 1, they have low substrate specificity. There are two types; those that contain cytochromep450 or flavin monooxygenases.Flavin monooxygenases:Important in the biotransformation of drug containing sulphurous or nitrogenous groups on aromatic rings. Itproduces sulfoxides and nitroxides.Cytochrome P450 monooxygenases:Major monooxygenases of ER, over 30 isoforms in humans. Haemoproteins, they are the most versatile biocatalysts inthe body. Highly active in liver, occur in all tissues except RBC and skeletal muscle. They are inducible/inhibited bycertain xenobiotics.19 The metabolism of xenobiotics - stage II (conjugation). Reaction types, reactant activation, products –examples).General Medicine 4th semester 2009
  14. 14. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedStage 2:Cytoplasmic enzymes catalyze conjugation of the functional groups, introduced in the first phase reactions, with apolar component (glucouronate, sulphate, Glycine, etc). These products are less biologically active.It renders xenobiotics more water soluble, to enable excretion. Transferases are cytosolic or bound in membranes ofER, and they catalyse conjugation, acetylation or methylation of polar groups added from phase 1. Reactions areendergonic (require energy), and one of the reactants must be activated.Reaction type Reagent Group in XenobioticGlucournidation UDP-Glucouronate -OHSulfation PAPS -OHMethylation S-AM -phenolic OHAcetylation Acetyl-CoA -NH220 Alcohols and phenols as xenobiotics and their transformation (ethanol and ethylene glycol, salicylates and acetaminophen).General Medicine 4th semester 2009
  15. 15. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed21 Principles of metabolism control (control of enzyme activity and of protein synthesis, control of transport across membranes, extracellular signals).General Medicine 4th semester 2009
  16. 16. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedControl of enzyme activity is a more rapid type of control than the control of enzyme synthesis. The enzyme activitiescan be changed effectively in several ways.- Activation of proenzymes by partial proteolysis of the proenzymeActive enzymes are formed from proenzyme molecules by irreversible splitting of certain parts in their polypeptidechains. This principle of activation is frequent among proteinases because it prevents unwanted breakdown ofproteins.- Allosteric conrol and cooperative effects of enzymes that consist of several identical subunitsRegulatory enzymes are frequently oligomers that consist of several identical subunits. Their saturation curves areusually sigmoid shaped. Allosteric effectors bind non-covalently at a site other than the active site and may eitherstimulate or inhibit the activity of the enzyme.- Control arising from regulatory proteins- Control by reversible covalent modification of enzymes or of their regulatory proteinsPhosphorylation, catalyzed by protein kinases. Acetylation from Acetyl CoA. Carboxylation of glutamyl in residues sidechains.Transport across membranes is regulated. For example, insulin stimulates glycolysis because it promotes the uptakeof glucose by muscle and adipose tissue. Binding of insulin to its receptor leads to rapid increase in the number ofGLUT4 transporters in the plasma membrane.Transduction of extracellular signals is important for the cell in receiving and responding to information from theenvironment. Proteins and small polar signal molecules bind on to specific membrane receptors, which results in aconformational change of the intracellular domain, resulting in the increase of secondary messenger molecule oractivation of a protein kinase. Non-polar signal molecules diffuse through plasma membrane and bind to specificproteins called intracellular receptors.22 General features of hormone synthesis, secretion, transport, and inactivation in relation to signal intensity received by the target cell.General Medicine 4th semester 2009
  17. 17. Masaryk University Biochemistry II Exam Questions Reband Ahmed & Khuram Ahmed Hormone synthesis: Protein and peptide hormones are synthesized on the rough ER and in different endocrine cells. They are first “secreted” as large proteins which are biologically inactive – prohormones, which start to get smaller in the ER. Prohormones are transferred to the golgi apparatus for packaging into secretory vesicles. In these vesicles, enzymes cleave the prohormones to produce smaller, biologically active hormones and inactive fragments. >Vesicles are stored within cytoplasm or in the cell membrane until their secretion is needed => exocytosis. Stimulus of exocytosis can be increased by depolarisation of the plasma membrane => Hormone secretion. Hormone secretion - feedback control of hormone secretion -ve feedback control - ensure proper level of hormone activity at the level of the target tissue; After a stimulus causes release of the hormone, conditions or products resulting from the action of the hormone tend to suppress its further release – prevents over secretion or over activity. +ve feedback control – occurs when the biological action of the hormone causes different additional secretion of the hormone; e.g. Luteinizing hormone is secreted as result of the stimulating effect of estrogen from the anterior pituitary before ovulation. LH increases when estrogens increases in the ovaries. Transport of hormones into blood:  Water-soluble hormones are dissolved in the plasma and transported from their sites of synthesis to target tissues, where they diffuse out of the capillaries, into the intestinal fluid, and eventually to target cells.  Steroid and thyroid hormones circulate bound to plasma proteins. Inactivation of hormones – there are two main factors increasing or decreasing the concentration of hormones in blood: 1. rate of hormone secretion into the blood and 2. rate of removal of hormone from the blood – metabolic clearance rate. Metabolic clearance rate = rate of disappearance of hormone from plasma (conc. of hormone / ml of plasma). Ways of clearance: => metabolic destruction by the tissues, => binding with the tissues, => excretion by the liver into bile, => excretion by the kidneys into urine Hormones can be degraded of their target cells by enzymatic processes that cause endocytosis of the cells membrane hormone- reseptor complex the hormone is then metabolized in the cell, and receptors are recycled back to the cell membrane.23 Membrane receptors cooperating with G-proteins (types of receptors and G-proteins, corresponding intracellular messengers). General Medicine 4th semester 2009
  18. 18. Masaryk University Biochemistry II Exam Questions Reband Ahmed & Khuram Ahmed Types of membrane receptors 1. Ion channel receptors mediated by neurotransmitters in synapses – quick responses. 2. G-protein linked receptors – “G” because they bind GDP and GTP - result in specific ligand binging in: Stimulate/inhibit phospholipase C Stimulate/inhibit phosphodiesterase Stimulate/inhibit phosphodiesterase 3. Receptors with enzyme activity – granylate cyclase 4. Receptors activating non-receptor tyrosine kinase activity G-proteins (response in a few minutes) - GTP/GDP binding proteins - Freely membrane bound (can move along the inner surface) - Participate in various types of second messenger production - All have a similar structure and mechanism of activation - Heterotrimers consist of subunits A, B, and Y G-protein linked receptors All have some common structural features: 1) extracellular parts are slightly glycosylated, have accessory binding sites for agonist 2) membrane parts: 7 a-helical segments span the membrane, connected by intra and extracellular hydrophilic loops 3) intracellular parts, which have the bingind site for a specific G-protein type G-protein activation - Resting state = a-unit has GDP attached - Hormone binds to extracellular part, makes a complex with the receptor, and GDP is phosphorylated to GTP - The a-GTP interacts with the effector enzyme – activate/inactivated enzyme which causes an increase or decrease in secondary messenger signal EXAMPLE: receptors with adenylate cyclase system - membrane bound receptor that catalyses ATP > cAMP + PPi - cAMP is a secondary messenger - Gs-protein stimulates adenylate cyclase, so the cAMP increases - cAMP activates PKA, which is used in phosphorylation reactions - Gi-protein inhibits AC – opposite effect Gq-protein stimulates phospolipase C Gt-protein stimulates cGMP phosphodiesterase24 Plasma membrane phosphatidylinositols and the phosphoinositide cascade, the role in signal transduction. Inositol sources: exogenous (plant food) and endogenous (Glucose-6-phosphate) General Medicine 4th semester 2009
  19. 19. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedCascade:Signal molecule binds to the receptorThe receptor activates the G-proteinActivated G-protein (a-unit and GTP) activates the effector = phospolipase CPhospholipase C catalyses the hydrolysis of PIP2 > DG + IP3DG and IP3 are secondary messengersDG activates PK C – phosporylations in the presence of Ca2+IP3 opens Ca2+ channels in ER > cytosol Ca2+ concentration increasesCa2+ is associated with calmodulinCalcium-calmodulin complexes activate calmodulin dependant kinasesPhosphorylated intracellular proteins carry out a biological response to the signal molecule  Enzymes for glycogenolysis and gluconeogenesis are activated by phosphorylation.  Enzymes for glycogen synthesis, glycolysis, FA synthesis and cholesterol synthesis are inactivated by phosphorylation.Phosphatidylinositol:Phosphatidate is esterified with myo-inositolPIP2 is a part of membranes25 Protein kinases (main classes) and phosphoprotein phosphatases, regulation of their activity.Reversible phosphorylation of proteins is intracellular and ATP is the phosphate donor. Phosphorylation is catalysed byhighly specific protein kinases. Protein kinases are the largest family of homologous enzymes, there are over 550human types.General Medicine 4th semester 2009
  20. 20. Masaryk University Biochemistry II Exam Questions Reband Ahmed & Khuram Ahmed There are two sites where proteins can be phosphorylated: 1. On the serine/threonine residues (alcoholic groups) 2. Tyrosine residues (phenolic hydroxyl) They are both at specific positions in the polypeptide chain. The signal that activates PK is amplified causing phosphorylation of numerous protein molecules.  Dephosphorylation of phosphoproteins is carried out by PHOSPHOPROTEIN PHOSPHATASES, and it involves the hydrolysis of the ester bond. Because protein kinases have profound effects on a cell, their activity is highly regulated. Kinases are turned on or off by phosphorylation (sometimes by the kinase itself - cis-phosphorylation/autophosphorylation), by binding of activator proteins or inhibitor proteins, or small molecules, or by controlling their location in the cell relative to their substrates.26 Insulin (synthesis, regulation of secretion, fate, insulin receptor and results of its activation). Oral glucose tolerance test. Synthesis: General Medicine 4th semester 2009
  21. 21. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedIn B-cells, islets of langerhans, within the pancreas. Preproinsulin is produced in the endoplasmic reticulum. It is asingle peptide. Cleavage of the single peptide and formation of disulphide bonds makes Proinsulin. This passes to thegolgi, where it is placed in to vesicles called B-granules. After cleavage of the C-peptide, mature insulin is formed inthe B-granules. It has two peptide chains held together by disulphide bridges.Secretion:Secreted in response to increase in blood glucose levels. Stimulates glycolysis, lipogenesis, and glycogen synthesis andstorage in the liver. Inhibits gluconeogenesis, glycogenolysis and lipolysis.Degredation:Insulin binds to receptor (in liver or kidney) and enters the cell by endocytosis of the insulin-receptor complex.Insulase acts on the complex, breaking it down.Regulation of secretion: 1. Increased blood glucose levels is a signal for increased secretion 2. Increased amino acids in plasma after ingestion of proteins also increases secretion 3. Gastrointestinal horomone secretin, released after ingestion, causes anticipatory riseReceptor: Transmembrane receptor, activated by insulin Belongs to tyrosine-kinase receptors Insulin binds to receptor Starts many protein activation cascades, translocation of GLUT4 to plasma membraneoGTT – oral glucose tolerance test:Used when increased concentration of fasting glucose is found in the serum/plasma. It tests the effectiveness ofglucose metabolism.Procedure:  Blood sample is taken after overnight fasting (10-14 hours)  75g of glucose in 300ml tea  Blood sample is taken every 1-2 hours after drinking the tea Normal values 0 hours 1 hour 2 hours Normal <6 <11 <8 Impaired >6 >11 8-11 Diabetes mellitus >7 >11 >1127 Intracellular hormones receptors, their activation and consequences.Lipophilic hormones diffuse through the plasma membranes to bind to receptors in the cytoplasm or in the nucleus oftarget cells. The hormone-receptor comlex under goes activation reaction.General Medicine 4th semester 2009
  22. 22. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed The hormones bound to their transport proteins in the blood, attach to the megalin transport protein and passes into the cytoplasm. In lysosomes the hormone is released from its binding protein via hydrolysis and the hormone binds with its intracellular receptor. The intracellular hormone-receptor complex binds to DNA sequence HRM (hormone response element) – works as enhancer supporting initiation of transcription on the promoter.  Gene transcription effect and production of target mRNA - Amount of specific protein changed - Metabolic processes are influenced  Low density lipoprotein-related protein 2 also known as LRP2 or megalin is a protein which in humans is encoded by the LRP2 gene. Function: LRP2 is multiligand binding receptor found in the plasma membrane of many absorptive epithelial cells. LRP2 is a member of a family of receptors with structural similarities to the low density lipoprotein receptor (LDLR). LRP2 functions to mediate endocytosis of ligands leading to degradation in lysosomes or transcytosis. LRP2 (previously called glycoprotein 330) together with RAP (LRPAP1) forms the Heymann nephritis antigenic complex. LRP2 is expressed in epithelial cells of the thyroid (thyrocytes), where it can serve as a receptor for the protein thyroglobulin (Tg).28 The role of hypothalamic and pituitary hormones – a brief survey, functions.Hypothalamus – affects the endocrine system, controls emotional behaviour. Most hypothalamic hormones go topituitary via hypophyseal portal system. It maintains homeostasis, including blood pressure, heart rate andtemperature regulation.Hypothalamic hormones control the release of the anterior pituitary gland hormones and the hormones of theposterior pituitary gland are synthesized in the magnocellular neurons in the hypothalamus.The pituitary gland secretes hormones regulating homeostasis, including trophic hormones that stimulate otherendocrine glands. It is connected to the hypothalamus by the medial eminence.Name Location FunctionCorticotropin-releasing hormone paraventricular nuclues with ADH, stimulates anterior pit. To secrete ACTHDopamine arcuate nucleus inhibits anterior pit. Secreting prolactinGonadotropin-releasing hormone arcuate nucleus stimulates anterior pit. To secrete LH and FSHGrowth hormone releasing hormone arcuate nucleus stimulates anterior pit. To secrete GHVasoprissin (ADH) paraventriculat nuclues with CRH, stimulates anterior pit. To secret ACTHACTH, adrenocorticotropic hormone, polypeptide – secretion of glucocorticoids.Beta Endorphins, polypeptide – inhibits perception of pain.Prolactin, polypeptide – milk production in mammary glands.TSH, thyroid stimulating hormone, glycoprotein – secretion of thyroid hormones.Growth hormone, glycoprotein – promotes growth and lipid/carb metabolism.29 Synthesis of thyroid hormones (description, localization, secretion and its control).Thyroxine (T4) – tetraiodothyronine and it’s active form triiodothyronine(T3)From tryosineTakes place in thyroid gland-follicular cellsT4 has a longer haf life than T3General Medicine 4th semester 2009
  23. 23. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedT4:T3 is 20:1 in blood, bound to transport protein (thyroxine-blinding globulin)A small amount is free and biologically activeT4 deiodinaes to T3 when neededIodothyrodines are the only organic molecules in the body that contain iodineT4 and T3 are lipophilic – cross the cell membrane easilyThyroid-stimulating hormone regulates their synthesis at every step. It is a glycoprotein, from the anterior pituitary. Itincreases basal metabolsm, heat generation and o2 consumption.PRECURSOR: thyroglobulinOVERVIEW: iodide anions are oxidized by thryoperoxidase (TPO) and incorporated to tyrosyl residues of thyroglobulin.Tyrosine is converted to thryoglobuin in thyroid follicular cells.Thyroglobulin reacts with I2 to form monoiodotyrosine and diiodotyrosine (MIT/DIT).Thyroxine is formed when two molecules of DIT combine.T3 is formed when a molecule of MIT and DIT combine.30 Intracelullar Ca2+ distribution - calcium channels, carriers, Ca2+-dependent proteins (e.g. calmodulin) and enzymes, relations to cell functions.Distribution: 2+Whole Ca = 1-1.3kgIt is located in the bones (99%) and body fluis (ICF 0.9% ECF 0.1%)Blood plasma concentration is (2.5mmol/l): 2+50% free ionized Ca BIOLOGICALLY ACTIVE 2+32% Ca bound to albumin 2+8% Ca bound to globulins 2+10% Ca bound in complexes with anions CHELATED 2+Ca functions:It is a bone componentSignalling substance, second messengers in transduction pathways -cause exocytosis -muscle contraction -co-factors in blood coagulationStored in the SER, which keeps the cytoplasm levels low – good function in sarcoplasmic reticulum 2 -for the release and uptake, SER membranes contain signal controlled Ca channels with energy 2+ Dependant Ca ATPase 2+Ca -Calmodulin: 2+Calmodulin is a small protein found in all animal cells, which can bind 4 Ca ions1. Hormone binds receptor in the cell membrane2. Via G-Proteins, this has 2 actions 2+ -mobilises intracellular Ca storesGeneral Medicine 4th semester 2009
  24. 24. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed 2+ - opens Ca channels in the cell memrane3. Activated G-protein activates phospholipase -PLC catalyses the hydrolysis of PIP2 to DG and IP3 -DG activated PKC which phosphorylates enzymes 2+ -IP3 opens Ca channels in ER 2+4. Ca binds to calmodulin and this complex produces physiological actions5. It activates calmodulin dependant kinases – phosphorylated intracellular proteins for a biological response31 Calciferols (calciols) - structure, sources, transformations, effects, mechanism of action.The calciols are several forms of vitamin D, a family of sterols that affect calcium homeostasis. Their daily requirementis 5-20ug. D-provitamins (ergostrerol and 7-dehydrocholesterol) are widely distributed in animals and plants.Most natural foods have a low content of vitamin D3. It is present in egg yolk, butter, cows milk, beef and pork liver,animal fat and pork skin. The most important vitamin D (D2) source is fish oil, primarily liver oil. ≡ Calciol (cholecalciferol, vitamin D3) Ercalciol (ergocalciferol, vitamin D2) The calciols are 9,10-sekosteroids, in which the ring B is opened.The effects of calciols: 1. Increase absorption of Ca2+ by enterocytes 2. Regulates reabsorption and regeneration of bone tissueIn human liver, a small amount of cholesterol transforms into 7-dehydrocholesterol and from that, in dermal capillaryexposed to sun radiation, calciol (cholecalciferol, vitamin D3) is formed - by of opening of the ring B(C9-C10 bond): Cholesterol THE LIVER CELLS 7,8-Dehydrogenation Lumisterol Tachysterol 7-Dehydrocholesterol Capillaries of the SKIN A high-speed photolysis λ max = 295 nm Slow thermal conversion An intermediate (praevitamin) Calciol is slowly released into blood Calciol (vit. D3) and bound to serum DBP (D vit. binding protein).General Medicine 4th semester 2009
  25. 25. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed Calciol is an inactive precursor of calcitriol, the most potent biologically active form of vitamin D. The hydroxylation of calciols C-25 The LIVER CELLS The RENAL TUBULAR CELLS 1α 25-Hydroxylation 1α-Hydroxylation (monooxygenase, cyt P450) (monooxygenase, cyt P450) Calciol Calcidiol Calcitriol (Cholecalciferol) (25-Hydroxycholecalciferol) (1α,25-Dihydroxycholecalciferol) A CALCIOTROPIC STEROID HORMONE Calcidiol is the major circulating metabolite of calciol. Its biological half -life is rather long, approx. 20 - 30 days. The concentration of calcidiol in blood plasma informs of the body calciol saturation. Seasonal variations are observed. 25-Hydroxylation of calcidiol is inhibited by the high concentrations of calcidiol and calcitriol (feedback control), calcitonin, and the high intake of calcium in the diet. Calcitriol has a short biological half-life. 1 -Hydroxylation is stimulated by parathyrin (PTH), inhibited by calcitonin and high concentrations of calcitriol.32 Calcium and (inorganic) phosphate metabolism - distribution in the body, mineral deposits and soluble forms, the role of PTH, calcitriol, calcitonin.Calcium = 1-1.3kg (99% bone, ICF 0.9%, ECF 0.1%)Blood plasma concentration is (2.5mmol/l): 2+50% free ionized Ca BIOLOGICALLY ACTIVE 2+32% Ca bound to albumin 2+8% Ca bound to globulins 2+10% Ca bound in complexes with anions CHELATEDHormonal control of plasma caclium concentration:PARATHYRIN – secretion regulated by plasma Ca2+ concentration: secreted in HYPOCALCEMIA. Stimulates bone resportion through differentiation and activation of osteoclasts In the renal tubules, Ca2+ resorption increases and HPO42- resporption decreases Increased calcium absorption results in the intestinesCALCITONIN – secreted by the C-cells of the thyroid gland: secreted in HYPERCALCEMIA Counteracts PTH in the control of Ca metabolism Inhibits bone resorption Supports synthesis of organic matrix and mineralization of osteoid Inhibits resorption of Ca2+ AND phosphates, increasing both their excretion in this wayCALCITROL – steroid hormone, from tthe kidneysStimulates resporption of Ca+ and HPO42- from the renal tubulesIncreases blood Ca2+ concentration by increased Ca2+ mobilization from boneIncreases plasma level of both ionsHypercalcemiaPlasma concentrations above 3.5mmol/lRenal functions are imparedSoft tissue calcification and renal stones developHypocalcemiaPlasma concentration is below 2mmol/lIncreased neuromuscular excitability and tetany (carpopedal spasms)General Medicine 4th semester 2009
  26. 26. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed33 Synthesis and inactivation of catecholamines, degradation products.Biogenic amines with a catechol group. Biosynthesis occurs in the adrenal cortex and CNS, from tyrosine.  Tyrosine hydroxylation is the rate limiting step.Inactivation is by MONOAMINE OXIDASE (MAO). They are found in the neural tissue, gut and liver. Inactivation is bymeans of oxidative deamination to acidic metabolites and 3-O-methylation to metanephrines. Metabolic products ofthese reactions are excreted in urine as vanillylmandelic acid, metanephrine, and normetanephrine. 34 Glucocorticoids - structure, biosynthesis, function, regulation of secretion.Are synthesized mainly in the zona fasiculatis of the adrenal cortex.Function:play crucial role in adaption of the organism to the state evoked by stress. They increase glucoseconcentration in blood by stimulating liver gluconeogenesis. They also make amino acids more easily available bysuppressing proteosynthesis and supporting breackdown of proteins. Administration of high doses of glucocorticoidscan evoke immunosuppressive effect, necessary after organ transplantations.Glucocorticoids have anti-inflammatory effects.The most important glucocorticoid is Cortisol; secretion controlled by ACTH (adrenocorticotrophic hormone).Synthesis:Cortisol – is a major glucocorticoid, synthesized from progesterone by hydroxylations at C17, 21, and 11. Secretionunder basal conditions 22-70umol/day.General Medicine 4th semester 2009
  27. 27. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed35 Mineralocorticoids - structure, biosynthesis, function, regulation of secretion, the renin- angiotensin system.Synthesis occurs in the zona glomerulosa of the adrenal cortex. The zona glomerulosa doesnt express the 17-hydroxylase, so it doesnt produce precursors of gluticoids. It is the site of aldestorone production. The synthesis andsecretion is controlled by Renin-Angiotensin system. ACTH influence is very weak.Functions:- Act on the kidney to increase reabsorption of Na+ and the excretion of K+, leading to increase in BP and volume(this is effective in keeping the water mineral balance)Cholesterol > Pregnenolone > Progesterone > Corticostreone > AldosteroneRenin-Angiotensin System:1. Decrease in blood volume causes a decrease in renal perfusion pressure = increases renin secretion.Renin is an enzyme that catalyses the conversion of angiotensinogen to angiotensin I. Then angiotensin I >angriotensin II by angiotensin converting enzyme ACE.2. Angiotenin II acts on zona gomerulosa to increase conversion of corticosterone to aldosterone3. Aldosterone increases reanal Na+ reabsorption, restores ECF volume and blood volume back to normal.Renin is produced when stimulated by:Decrease in pressure in afferent arteriolesCirculating catecholeaminesDecrease of [Na+] and [Cl-] in the tubular fluid36 Alkali cations - distribution in various compartments, approx. daily intake and output, control of the excretion (angiotensin-aldosterone, natriuretic peptides), consequences of retention or of heavy losses of electrolytes.Plasma cations + ECF: ICF:[Na+] – 140mmol/l [Na+] – 10mmol/l[K+] – 4.4 mmol/l [K+] – 155 mmol/l[Ca2+] – 2.5mmol/l [Ca2+] – 1umol/l[Mg2+] – 1mmol/l [Mg2+] – 15mmol/lDaily Intake:Na+ 500mg/dK+ 4mg/dCa2+ 20-25mmol/dGeneral Medicine 4th semester 2009
  28. 28. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedOutput:Ca2+ 17-25mmol/dAngiotensin-aldosterone:Renin, angiotensin and aldosterone work together to maintain blood pressure.Deacreased blood pressure makes kidneys release rennin by juxtaglomerular cells.Anginotensinogen>Angiotensin I>Angiotensin II>increases production of aldestoroneNa+ and H20 retention increases, which increases blood pressure and volumeNatriuretic peptidesAtrial natriuretic peptides, ANP, are secreted by atrial myocytes. ANP acts to reduce the water, sodium and adiposeloads on the circulatory system, thereby reducing blood pressure.Secreted in response to: - Atrial distention - Sympathetic stimulation - Increased [Na+] - Angiotensin IIANP decreases Na+ and H2O which decreses blood pressure and volume. At the same time, it increases K+.Brain natriuretic peptide is secreted by heart ventricles due to excessive stretching of heart muscle cells. Aswell asdecreasing blood pressure and volume, it also increases cardiac output.37 Sex hormones (structure, biosynthesis, function, sites of secretion and their regulation, inactivation).Testosterone (C17) – synthesised in Leydig cells in the testis.Oestrogen and progesterone – developing follicles of the corpus luteum in the ovaries.Adrenal Androgens – need 17a-hydroxylationANDROSTENEDIONE (precursor for testosterone)TESTOSTERONEDihydrotestosterone and estradiol are also in the circulation, from the conversion of testosterone.General Medicine 4th semester 2009
  29. 29. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedOESTROGEN:Synthesis is stimulated by LH and FSH. The precursor is an androgen (enzyme is cytochrome P450) which ishydroxylated twice on the methyl group on C19, and then hydroxylation of C2 forms a product which gives anaromatic ring at A:Three types are produced: estriol, estradiol and estrone. ESTRIOL Progesterone: Prepares the lining of the uterus for implantation of an ovum and is also essential for the maintenance of pregnancy. It is also a precursor for androgens and estrogens. Cholesterol > Pregnenolone > Progesterone > Androgens > Estrogens It is rapidly removed from the circulation; coverted to pregnanediol and conjugated to glucunnate in liver to be excreted as urine. PROGESTERONEGeneral Medicine 4th semester 2009
  30. 30. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed38 Neurons - components of an axon membrane and myelin, provision of energy and nutrient requirements, relationship of neurotransmitters to amino acids (a survey).Dendrites: have receptors for neurotransmittersPerikaryon: body – have the nuclues and is the metabolic centreAxon: for pimary active transport of Na+/K+ across the axolemma, contains voltage gated channelsAxonal transport: transport along microtubules, anterograde and reterogradeNodes of Ranvier: provides method of fastor saltatory conductionAxon terminals: synapses where neurotransmitter is released from synaptic vesicles by exocytosisMyelin:Myelin sheaths are wrapping of glial cells around the axons. In CNS glial cells are oligodendrocytes,in PNS they are Schwann cells.Energy and Nutrient Requirements:Glucose is the main nutrient, in prolonged starvation KB can provide half the energy requirements.This is why impairment of consciousness is the first sympton of hypoglycemia.Other neurotransmitters such as catecholamines are synthesized from the amino acid tyrosine which is a hydroxylateof phenylalanine.General Medicine 4th semester 2009
  31. 31. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed39 Membrane potential of a neuron, depolarization and the action potential propagation. Voltage- operated and receptor-operated (ligand-gated) ion channels.40 Adrenergic synapse (release and inactivation of the transmitter, the types of adrenergic receptors, signal transduction). Adrenergic synapses release catecholamines by endocytosis due 2+ to increased conc. of Ca in ICF ! Inactivation of the transmitters is done: - Acetylcholine => is cleaved by acetylcholinesterase - norepinephrine and epinephrine are taken upp by the postsynaptic/presynaptic membrane => reuptake.General Medicine 4th semester 2009
  32. 32. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed41 Cholinergic synapse (biosynthesis of the neurotransmitter and the release of it, two principal types of acetylcholine receptors and mechanisms of their function).Acetyl choline is the neurotransmitter. Acetyl choline formation takes place in the cytoplasm of the presynaptic axon.Choline + Acetyl Co-enzymeA Acetyl cholineInactivation of acetyl choline by acetylcholine esterase is in the synaptic cleft.Cholinergic synpase:Depolarisation causes intracellular Ca2+ concentration to increaseThis activates calcium-calmodulin dependant protein kinase > phosphorylates synapsin-1This interacts with synpatic vesicles, initiates there fusion with the presynaptic membrane and neuroT exocytosisMembranes of vesicles are recycles +Nicotinic receptors are ligand-gated ion channels, for Na influx on normal action potential producing structures i.e.nerves of muscles. 2+ neural nicotinic cholinergic receptors for Ca permeability in synaptic facilitation and learning.General Medicine 4th semester 2009
  33. 33. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed42 Acetylcholinesterase and its inhibitors (examples of organophosphate insecticides, typical signs of toxic effects, the first aid - the counteractive alkaloid).The effect of organophosphates is based on the fact that they block covalently the enzyme acetylcholine esterase,which catalyzes the hydrolytic breakdown of acetylcholine in the synaptic gap. [Acetylcholine is not sufficiently brokendown; it cumulates and causes long-term stimulation of the receptors in the postsynaptic membrane. Thereforeorganophosphate poisoning is viewed as a long-term stimulation of the motor neurons and the stimulation of theparasympathetic nervous system.]Inhibitors:Principle – esterification of serine hydroxyl in the active site of the enzyme 1) Reversible: Carbamates 2) Irreversible: Organophosphates (form a covalent bond with enzyme)Signs of toxic effects:S - salivationL - lacrimationU – urinary incontinenceD - defacationG – GI upsetE - EmesisM – MiosisFirst Aid:Atropine: blocks the parasympatheric nervous system, both vagal effects on the heart by blocking the acetylcholineaction at the muscarinic receptors.Organophosphates => very strong nerve paralyzing poisons, which can be absorbed through the skin. The mostexample of toxic insecticide, commonly used in agriculture is parathion or the most toxic mevinfos.43 Inhibitory GABAergic synapse (GABAA receptors, the effect of benzodiazepines and other ligands). Inhibitory GABAA receptor is a ligand-gated channel (ROC) for chloride anions. The interaction with -aminobutyric acid (GABA) opens the channel. The influx of Cl– is the cause of hyperpolarization of the postsynaptic membrane and thus its depolarization (formation of an action potential) disabled. Cl– The receptor is a heteropentamer (three subunit types). Besides the 1 2 binding site for GABA, it has at least 2 1 eleven allosteric modulatory sites for compounds that enhance the response 2 to endogenous GABA – reduction of – anxiety and muscular relaxation: – – – – – – anaesthetics, ethanol, and many useful drugs, e.g. benzodiazepines (hence the alternative name GABA/benzodiazepine receptors), meprobamate, and also barbiturates. Some ligands compete for the diazepam site or act as antagonists (inverse agonists) so that they cause discomfort and anxiety, e.g. endogenous peptides called endozepines. In the spinal cord and the brain stem, glycine has the similar function as GABA in the brain. The inhibitory actions of glycine are potently blocked by the alkaloid strychnine, a convulsant poison in man and animals.General Medicine 4th semester 2009
  34. 34. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed44 Retinol and its derivatives - the biological role, biochemistry of visual excitation (activation of transducin, consequences in decrease of cGMP with hyperpolarization and in decreased Ca2+ stimulating guanylate cyclase).Retinol (Vitamin A)Primary alcohol containing B-ionone ring and unsaturated side chain. Found in animal tissues as a retinyl ester with along chain fatty acid.Retinal – component of phodopsin of rod cells in the retinaAldehyde from retinol oxidation, both can be interconverted.Retinoic Acid – takes part in cell regulation of gene expressionIs an acid from the oxidation of retinal. It can’t be reduced in the body to give retinol or retinal.B-carotene: is from plant food, can be oxidatively cleaved to give two molecules of retinal.Retinoids are essential for vision, reproduction, growth and maintinance of epithelial tissues. Retinoic acid mediatesmost of the actions of the retinoids except vision, which is mediated by retinal.Sources of vitamin A: CARROTS, liver, kidney, egg yolk and butter.! Rhodopsin is found in rods (photoreceptors). It is a light sensetive chromoprotein. Opsin part contains retinal.Absorption of a photon triggers isomerisation of retinal. This leads to allosteric conformational change of rhodopsin,which binds to G-protein-TRANSDUCIN. A signal cascade follows and rods release less neurotransmitter (glutamate).Bipolar neurons register this change and transmit it to the brain for light.In the dark, rod cells have a high concentration of cGMP (synthesized by guanylate cyclase), which binds to an ion + 2+channel to open it and allow Na and Ca to enter, causing depolarization and release of glutamate neurotransmitter. +! Decrease of cAMP => Na channels closesGeneral Medicine 4th semester 2009
  35. 35. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed45 Distribution of body water, factors influencing the distribution of body water and its excretion (ADH, aldosteron, natriuretic peptides), consequences of retention or of dehydration.Distribution of body water:Total body water: 60% - ECF is 20% (1/3) ICF 40% (2/3)ECF: ¼ blood plasma and ¾ interstitial fluidHigher in new borns and adult malesLowest in females and fat peopleFactors Influencing the distribution:AGE: highest in newborns, lowest in old femalesGENDER: higher in males, lower in femalesWEIGHT: fat has 2% water content, whereas other tissues have 73% water content (more fat=less water)ADH (anti-diuretic hormone or vasopressin):from the posterior lobe of the pituitary, increases water permeability of the distal tubules and collecting duct.Aldosterone:Decrease in blood volume causes decrease in renal perfusion pressure which causes an increase in renin secretion.Renin converts angiotensinogen to angiotensin I, and then ACE converts it to angiotensin II, which acts on the zonaglomerulosa to increase conversion of corticosterone to aldosterone. Aldosterone increases renal resporption of Na+and so increases blood volume back to normal.Natriuretic peptides:Atrial natriuretic peptides, ANP, are secreted by atrial myocytes. ANP acts to reduce the water, sodium and adiposeloads on the circulatory system, thereby reducing blood pressure.Secreted in response to: - Atrial distention - Sympathetic stimulation - Increased [Na+] - Angiotensin IIANP decreases Na+ and H2O which decreses blood pressure and volume. At the same time, it increases K+.Brain natriuretic peptide is secreted by heart ventricles due to excessive stretching of heart muscle cells. Aswell asdecreasing blood pressure and volume, it also increases cardiac output.General Medicine 4th semester 2009
  36. 36. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed46 Osmotic and oncotic pressure of blood plasma, plasma osmolality (values of the main parameters, empirical relations for a rough estimate of plasma osmolality) and osmolality regulation.Osmotic pressure: hydrostatic pressure produced by a concentration gradient between two solutions on either side ofa semipermeable membrane.Oncoptic pressure: a form of osmotic pressure exerted by protein in blood plasma that tends to pull water in to thecirculatory system.Plasma osmolarity: a measure of the concentration of substrates in blood (Na+, K+, Cl-, urea, glucose, etc). The units itis measured in is ‘osmoles of solute per kg of solvent’ – mmol/kg H2O.RANGE: 275-299 mmol/kgH2O CRITICAL VALUE: 250 mmol/kgH2OUrine osmolarity = 500-850 mmol/kgH2O.Osmolarity Regulation:Body osmolarity is controlled by regulating the amount of water in the body through changes in the thirst and renalwater excretion. This controls body volume.If Na+ is high in the body, body water will be increased to reduce the osmolarity back to normal. The body volume willthen also increase.If the body volume is too low, ADH is released which promotes water resorption in the kidneys.Body osmolarity is sensed by osmoreceptors in the hypothalamus, which influences thirst and ADH secretion. Increasein osmolarity leads to an increase in thirst, and an increase in ADH secretion, which decreases renal water excretion.47 Electrolyte status of blood plasma. Relation of ion concentrations to acid-base balance (buffer base and strong ion difference, anion gap).Cations Molarity ChargeNa+ 142 142K+ 4 4Ca2+ 2.5 5Mg2+ 1.5 3 Total charge: 154Anions Molarity ChargeCl- 103 103HCO3- 25 25Proteins 2 18HPO42- 1 2SO42- 0.5 1Organic 4 5 Total Charge: 154Strong Ion Difference:SID = [Na+] + [K+] - [Cl-] = 38-46mmol/l (proportional to buffer base of serum)SID composition = HCO3- + HPO42- + Prot-Strong ions don’t hydrolyse in aqueous solution.Increased strong ion difference leads to long vomiting due to loss of Cl-.General Medicine 4th semester 2009
  37. 37. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedAnion Gap:Aproximate extent of unmeasured anionsAG = [Na+] + [K+] - [Cl-] - [HCO3-] = 12-18mmol/lAG compostition: HPO42 + Prot- + SO42- + OACauses of increased AG: - Kidney insufficiency - Diabetes, starvation - Poisoning by methanol - Lactoacidosis - Severe dehydration48 Transport of CO2 in blood: pCO2 in arterial and venous blood, [HCO3–], carbaminohaemoglobin, physically dissolved CO2, the ratio HCO3- / CO2+H2CO3 ).There are 3 forms of CO2 transport in blood:HCO3- = 85%Protein carbamates = 10%Physically dissolved = 5% (CO2 is more soluble in blood than O2)pCO2 of arterial blood: 4.6 – 6 kPa venous blood: 5.3 – 6.6 kPa[HCO3-] is the only method which communicates with the external environment. It is a buffer system found inerythrocytes.CO2 + H2O > H2CO3 (carbonic acid) > HCO3- (bicarbonate) + H+^first step catalysed by carbonic anhydrase[HCO3-]/[CO2+H2CO3] = 20:1Concentration of buffer base is 20x more than the concentration of the buffer acid. It shows that it is 20x moreresistant to acids.Carbaminohaemoglobin: Hb + CO2 - A reversible reaction - covalently bound to the N-terminus of heams (not iron!) - can also bing to the amino groups on the polypeptide chains of plasma proteinsGeneral Medicine 4th semester 2009
  38. 38. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed49 The acidic products of metabolism (H+-producing processes, approximate daily amounts of formed non-volatile acids, the origin of metabolic acidosis and alkalosis).Main acidic products:Lungs = CO2 – = 25, 000 mmol/dKidneys = H+ (NH4+ and H2PO4) = 80mmol/dKidneys = HCO3- = 1mmol/dH+ producing process:Non electrolyte > acid > anion- + H+e.g. anaerobic glycolysis: glucose > 2 lactate- + 2H+H+ consuming reactions:Anion- + H+ > non-electrolytee.g. gluconeogenesis from lactate: 2 lactate + 2H+ > glucoseProduction of CO2:Decarboxylation reactionse.g. Oxidative decarboxylation of pyrvate > Acetyl CoAAcidic Catabolytes: - Aerobic metabolism of nutrients > CO2 - Aerobic glycolysis > lactic Acid - KB production > acetoacetic acid/B-hydroxybutyric acid - Catabolism of cysteine > sulphuric acid - Catabolism of purine bases > uric acid - Catabolism of DNA/RNA > HPO42- + H+Metabolic Acidosis:Increased production of endogenous H+ - lactoacidosis, ketoacidosis...Intake of exogenous H+ - metabolites from methanol, administration of HCl...Loss of HCO3- and Na+ - diarrhea, burns, renal tubular disordersExcessive infusion of NaCl solution – dilution of plasmaMetabolic Alkalosis:Loss of Cl- and H+ - by vomitingIntake of HCO3- - excessive use of baking soda or alkaline mineral waterLoss of Cl- and K+ - by diureticsHypoalbuminemia – liver damage, severe malnutrition, kidney diseaseGeneral Medicine 4th semester 2009
  39. 39. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed50 Buffering systems in blood, blood plasma (components, concentrations), the main buffer bases in interstitial and intracellular fluids.Three main buffering systems: pKa 6.0 – 8.0Buffer Blood Plasma RBCHCO3-/H2CO3 + CO2 50% 33% 17%Protein/Protein-H+ 45% 18% 27%HPO42-/H2PO4- 5% 1% 4%TOTAL BUFFER BASES (mmol/l) 48+3 42+3 56+3Buffer capacity depends on concentration of both components and the ratio of both components. The best capacity iswhen buffer base concentration equals buffer acid concentration.pH = pKa + log [BB]/[BA]Hydrogen Carbonate Buffer:This is the only buffer system which communicates with the external environment.CO2 + H2O > (carbonic anhydrase) H2CO3 >(dissociates) H+ + HCO3-Effective concentration of carbonic acid (mmol/l) is 0.22 x pCO2 (0.22 is the solubility coefficient of CO2)H+ + HCO3- > H2CO3 > H2O + CO2OH + H2CO3 > H2O + CO2Hydrogen Phosphate Buffer:H2PO4- > HPO42- + H+ pKa = 6.8Found in ICF, bones, and urine.[H2PO42-] : [H2PO4-] is 4:1 in blood plasma.Protein Buffer:H-protein > H+ + proteinHistidine is the main amino acid of blood proteins.General Medicine 4th semester 2009
  40. 40. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed51 The role of the kidney and of the liver in acid-base balance.Kidneys: + -They excrete acidic species (NH4 , H2PO4 , uric acid, etc) -They reabsorb basic species (mainly HCO3 )Ammonia Excretion: + +NH4 enter tubular cells in the form of glutamine > (glutaminase) NH4 + glutamate +NH4 enters the urine by the K+ channelNH3 can freely diffuse through the tubular membrane +30-50mmol/d of NH4 excretedOther amino acids also give NH3 (alanine, serine, Glycine, etc)Proton Excretion: +Renal tubule cells can secrete H even though there is a concentration gradient from the blood to the urine + -CO2 + H2O  H2CO3  H + HCO3 - - - + -HCO3 goes back to the blood via Cl /HCO3 antiport or Na /HCO3 antiport + + +H enters the urine by secondary active transport in Na /H antiport +[Na ] gradient is the driving force for the proton excretion52 Blood acid-base parameters (reference values, changes of the values in acute disturbances and in the course of their compensation).Ph = 7.40 + 0.04pCO2 = 4.6 – 6.0 kPaOxygen parameters:pO2 = 12-13.3 kPa3O2 saturation of Hb by O2 is 94-99%Total Hb = 2.15-2.65mmol/lTissue hypoxia of any origin leas to lactic acidosis.HCO3- 24+3mmol/lBase Excess 0+3mmol/lBB serum 42+4mmol/l (lower because it doesnt include RBC which have haemoglobin)BB blood 48+3mmol/lCompensation: the process which occurs when one body system replaces the disturbed function of another, so thatthe ratio of [HCO3-] / pCO2 gets closer to normal (20:1)Correction: the process which occurs when the disturbed system itself returns the acid-base parameters to normal.Metabolic Acidosis:In acude disorders: [HCO3-], pH and [HCO3-]/0.22pCO2 decrease – pCO2 is normalCompensation: done by lungs via hypoventilation to reduce pCO2Correction: done by the kidneys, they increase reabosrbtion of HCO3-Metabolic Alkalosis:In acute disorders: [HCO3-], pH and [HCO3-]/0.22pCO2 increase – pCO2 is normalCompensation: done by the lungs via hypoventilation to increase pCO2Corrction: done by the kidneys, drecrease resorption of HCO3-General Medicine 4th semester 2009
  41. 41. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedRespiratory Acidosis:In acute disorders: [HCO3-], pH and [HCO3-]/0.22pCO2 normal – pCO2 is increasedCompensation: done by kidneys by proton excretion and resorption of HCO3-Correction: done by the lungs via hypervention to restore pCO2Respiratory Alkalosis:In acute disorders: [HCO3-], pH and [HCO3-]/0.22pCO2 normal – pCO2 is decreasedCorrection : is done by the lungs via hypoventialtion to restore pCO253 Filtration of the plasma through the glomeruli (composition and permeability of the filtration medium, glomerular filtration rate – creatinine clearance, glomerular proteinuria).Composition and permeability:There is a layer of fenestrated endothelial cells, which are negatively charged. They have pores with a diameter of 50-100nm. Large (Mr>60 000) and negatively charged proteins can’t pass through. Microproteins (Mr<6000 -10 000) passthrough easily.There is then a basement membraine, made of mainly collagen. It allows free movement of electrolytes, water, andsmall molecules. It contains sialic acid in glycoproteins, which have a negative charge and so repulse anionic proteins.The final layer is a layer of pedicles with slit membranes between them, the pores have a diameter of 5nm.Glomerular Filtration Rate (GFR) – Creatinine Clearance:Is the volume of blood plasma that is completely cleared of creatinine in one second.GFR = Vp = Vu x (Cu/Cp) Units: ml/sCorrected GFR is clearance values normalised to a standard body surface area. Creatine excretion is proportional tothe surface area of the glomeruli filtration system which is assumed proportional to the body surface area.SA = 0.167 x √ w x hPhysiological range of GFRcor= 1.3-2.6 ml/s/1.73m2It is age and gender dependant.Glomerular Proteinuria:The normal glomerular barrier to plasma proteins is disrpted, proteins with molecular mass higher than 60 000 arepresent in the urine. Proteinuria is when there is more than 300mg of total urinary protein per 24 hours.General Medicine 4th semester 2009
  42. 42. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram Ahmed54 Reabsorption and secretion in the renal tubules (water, electrolytes - natriuresis, low molecular compounds – glucose, amino acids, uric acid, tubular proteinuria), the term fractional excretion E/F.Sodium Reabsorption:Reabsorption is 95-99.5%. Aldosterone increases the Na+ reuptake, especially in the distal tubule. Atrial Natriureticpeptide will decrease the Na+ uptake.Proximal tubule: 65% reabsorption Na+/H+ antiport Na+/Glc antiport Na+/aa antiportAscending limb of loop of henele: 25% reabsorption Na+/K+/2Cl- symportDistal tubule: 4% reabsorption Na+/Cl+ symportPotassium Reabsorption:Reabsorption is 80-95%, secretion is up to 200%.Proximal tubule: 65% reabsorption Paracellular transportAscending limb of loop of henele: 10-20% reabsorption Na+/K+/2Cl- symportCollecting Tubule: SECRETION increase by aldosteroneChloride Reabsorption:Reabsorption is 95-99.5%Proximal tubule: 55% reabsorption Paracellular transportAscending limb of loop of henele: 20% reabsorption Na+/K+/2Cl- symportDistal tubule: 20% reabsorption Na+/Cl+ symportWater Reabsorption:Reabsoprtion is 70-80% mostly in the proximal tubule by aquaporins. There is high permeability in the descending limbof the Loop of Henle. The distal tubule and collecting duct have AQP-2 which are dependant on ADH. This determinesthe final concentration of urine.Urea Reabsorption:In the proximal tubule 5-% is reabosorbed. The collecting duct is permeable to urea. Here urea diffuse back tointerstitial fluid, the descending limb (urea recycling) and the vasa recta.Amino acids and glycerol are reabsorbed with Na+ symport (secondary active transport).Tubular proteinuria is when there is 150mg/g of protein in urine. It occurs when reabsorption of low Mr proteins inthe proximal tubule is disrupted.General Medicine 4th semester 2009
  43. 43. Masaryk University Biochemistry II Exam QuestionsReband Ahmed & Khuram AhmedFractional Excretion: EF = Vu/GFRPortion of water excreted into urine, from the total volume of the glomerular filtrate. Physiological range is 0.985 –0.997. A decreased value indicates Diabetes.55 Nitrogenous urinary constituents (substances from which they are derived, average daily excretions, the major factors on which the excreted amounts depend).Nitrogen Compounds Metabolite Origin Excretion (mmol/day) % of total NUrea detoxification of NH3 by liver 330-600 80-90%Creatinine Creatine catabolism 5-18 3-4% +NH4 Glutaminase + GHD reactions 20-50 3-5%Uric Acid Purine Bases catabolism 1-1.5 1-2%Free amino acids Proteolysis in tissues 4-14 1-2%! Amount of excretion depends on the intake of proteins in the diet and utilization of nitrogen by the body.56 Nitrogenous low-molecular constituents of blood plasma (the parent compounds and main factors influencing concentrations of urea, creatinine, urate, ammonium, amino acids). Amino Acids Source: dietry proteins Urea 3-8mmol/l Source: ammonia detox in liver Ammonia Source: deamination of amino acids Creatinine 70-125umol/l Source: creatine catabolism in skeletal muscles, increased in case of skeletal muscle damage Uric Acid: 200-420umol/l Source: purine bases catabolism57 Digestion in the mouth and in the stomach (constituents of the saliva, the gastric secretion, secretion of HCl, humoral control of hydrochloric acid output).Saliva is excreted by the salivary glands. The pH of salive is 7.0. 1-1.5l is excreted per day. It is slightly alkaline, andcontains 98% water, salts, glycoproteins and lubricants, antibodies and enzymes. Also contains amylase, lipase andlysozyme.Gastric secretion is secreted in the stomach. The pH is 1.0 and 2-3l is excreted per day. Gastric juice is neutral orslightly basic, when HCl is added the pH becomes 1-2. Mucus protects the stomach lining. HCl denatures proteins andkills bacteria. Intrinsic factor is a glycoprotein needed for reabsorption of vitamin B 12. TAG lipase cleaves fats.Humoral Control of HCL output: +Vagal stimulation increases H secretion directly and indirectly. Directly by stimulating parietal cells, and indirectly by +innervating G-cells to stimulate gastrin secretion, which then stimulates H secretion by endocrine action. +Gastrin also stimulates H secretion by interacting with receptors on the parietal cells. It is secreted after eating ameal.Histamine is released from ECL (enterochromaffin-like-cells) in the gastric mucosa and diffuses to nearby parietal cells +to stimulate H secretion.General Medicine 4th semester 2009