MBBS pH and Buffering Sept 2023 PDF

pH and buffering Despo Papachristodoulou  pH and buffers learning outcomes Students should be able to: explain the difference between strong and weak acids in terms of dissociation properties Define and explain pKa and its importance in biological buffering U...

pH and buffering Despo Papachristodoulou  pH and buffers learning outcomes Students should be able to: explain the difference between strong and weak acids in terms of dissociation properties Define and explain pKa and its importance in biological buffering Use the Henderson-Hasselbalch equation List a number of physiological buffers Explain how proteins can be used as physiological buffers Describe the function of Haemoglobin as a buffer for Hydrogen ions produced in metabolism and explain what allows this to happen  What is pH? pH is a measure of Hydrogen ion concentration (acidity or alkalinity of a solution). H+ H+ H+ H+  First important concept: acidity depends only on free hydrogen ions Not those still bound to anions. H+ H+  blood pH Blood is in contact with nearly every body cell. Regulation of its pH is particularly critical. Normally, a very narrow range (7.35 to 7.45).  Outside these limits it may be fatal. Living range: pH 7.0-7.8 acidosis- normal range-alkalosis  Where do acids in the body come from? Some acids enter in foods most are generated by: breakdown of proteins incomplete oxidation of fats or glucose, loading and transport of carbon dioxide in the blood.  Acid –base balance It is regulated in the body by the lungs the kidneys systems in the blood known as chemical buffers  Buffering Buffers resist abrupt and large swings in the pH of body fluids by: releasing H+ (acting as acids) when the pH begins to rise binding H+ (acting as bases) when the pH drops. As the pH rises…(0H- increases) B- H+ OH- OH- B- H+ OH- B- H+ B- H+ B- H+ As the pH drops…. (H+ increases) B- B- H+ B- H+ H+ B-  How do buffers operate? you must thoroughly understand what is: a strong acid a strong base a weak acid a weak base ionisation of water  More than half of our weight is water infants are 73% or more water (low body fat and low bone mass) water is only about 45% of body mass in old age.  a healthy young man is about 60% water a healthy young woman about 50%.  Ionisation of water Pure water is a 55.6M solution Water dissociates to a very small extent H2O → H+ + OH- [H+] x [OH-] = 10-14 M2 ionic product of water at neutrality, [H+] =[OH-] = 10-7M  pH = - log [H+] when [H+] = 10 -2 M pH is 2 At neutrality when [H+] =[OH-] [H+] = 10-7M pH = – log [10-7] pH = - (-7) = 7 when [H+] is 10-2, then [OH-] is 10-12 when [H+] is 10-4, then [OH-] is 10-10  remember 101 = 10 log =1 102 = 100 log = 2 10-2 = 1/100 log = -2 blood pH is 7.4 7.4 = - log [H+] [H+] = 3.98 x 10-8 M  Dissociation of acids Acids are proton (H+ ) donors Bases are proton acceptors. Acids that dissociate completely in solution are strong acids e.g HCl→ H+ + Cl- those that dissociate incompletely (depending on pH) are weak acids e.g. H2CO3 →H+ + HCO3- Strong bases are more effective proton acceptors than are weak bases. Strong acid v weak acid dissociation Titration curve of 50 ml of 0.1 M ethanoic acid with 0.1M NaOH [CH3COO-] = 0.100 M Vol NaOH added Titration of 50 ml of 0.1 M phosphoric acid with 0.1 M KOH H3PO4→H2PO4- H2PO4- →HPO42- HPO42- →POA3-  pKa What is pKa? pKa = –log Ka Ka is the dissociation constant. And so what? What does it mean? It is the pH at which the acid is half dissociated There are equal amounts of undissociated acid and its conjugate base The lower the pKa , the stronger the acid  At the pkA B- H+ B- B- B- B- H+ H+ B- Henderson-Hasselbalch equation [ A ] pH  pKa  log [ HA] [conjugateb ase] pH  pKa  log [acid ] A convenient way to relate the pH of a solution, the pKa of a weak acid and the relative amounts of dissociated and non-dissociated (unprotonated and protonated) forms of the acid.  How does pKa relate to buffering? Buffers are mixtures of weak acids and their conjugate bases Buffering is the ability of a solution to resist a change in pH when acid or alkali is added At the pKa there are equal amounts of dissociated and non dissociated forms of the acid (conjugate base and acid) At the pKa buffering is best Titration of 50 ml of 0.1 M phosphoric acid with 0.1 M KOH H3PO4→H2PO4- H2PO4- →HPO42- HPO42- →POA3-  Why is that? If H+ are added they can be picked up by the conjugate base If OH- are added the acid can donate a proton and form H2O In this way, the pH does not change So, if you know the pKa of your acid, you know at what pH it will buffer best  At the pkA B- H+ B- 0H- B- B- H+ B- H+ H+ B-  Physiologically important buffers: In blood, saliva and other body fluids : H2CO3→HCO3- pKa 6.1 H2PO4- → HPO42- pKa 6.8 Protein → protein- protein+ → protein Henderson-Hasselbalch equation [ A ] pH  pKa  log [ HA] [conjugateb ase] pH  pKa  log [acid ] H2CO3→HCO3- pKa 6.1 H2CO3 is proportional to the pCO2 Can distinguish cause of e.g. acidosis metabolic or respiratory Amino acids can be used as buffers Titration of glycine using strong base Acids: -COOH and –NH3+ -COOH is the stronger acid pKa = 2.34 - NH3+ is the weaker acid pKa = 9.66 NOTE best buffering around the two pKa values NO buffering by the zwitterion  Which amino acids are involved in physiological buffering? Glycine is not a good candidate as it buffers best at pH 2.3 and 9.6 Clearly the alpha carboxyl and alpha amino groups are not good physiological buffers (they are involved in the peptide bond anyway) The R groups are important Haemoglobin is an important buffer in the blood. Most amino acid side chains do not buffer in the physiological range Their pKa is outside the physiological range What makes haemoglobin a good blood buffer? The presence of a large number of histidine residues.  Change of pKa of a group The pKa of histidine in Hb is different from that of free His which is 6 Neighbouring groups affect the pKa Oxyhaemoglobin pKa = 6.8 Deoxyhaemoglobin pKa = 7.8  Which is the better buffer for H+ produced in metabolism? AND WHY? Henderson Hasselbalch equation [ A ] pH  pKa  log [ HA] [conjugateb ase] pH  pKa  log [acid ] pH of blood 7.4 pKa oxyHb 6.8 pKa deoxyHb 7.8 OxyHb 7.4=6.8+0.6 Log of base /acid is 0.6 The ratio of base/acid is 3.98: 1 DeoxyHb 7.4 =7.8 -0.4 Log of base/acid is -0.4 the ratio of base/acid is 1: 2.51  Lipid soluble molecules can diffuse freely across biological membranes substances that interact strongly with water do not. The molecule shown on the next slide is aspirin. What can you predict about the diffusion of aspirin across biological membranes?  It diffuses equally well at all pH values It diffuses more easily at pH 8 than pH 2 It diffuses more easily at pH 2 rather than pH 7