Buffer solutions المحاليل المنظمة

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1 Buffer solutions المحاليل المنظمة Presented by Dr. Mohammad Saadeh The requirements for the Pharmaceutical Biochemistry I Philadelphia University Faculty of pharmacy

2 Understanding ph balance The human body is designed to maintain a ph balance in its fluids, tissues and systems. As most biochemical reactions essential to life take place in an aqueous environment, however, it is our blood plasma and interstitial fluids surrounding the cells that are most sensitive to acid-alkaline imbalance. When these fluids are maintained within a narrow ph range of 7.35 to 7.45, our body s immune system is operating in optimal conditions and is able to fight off illness and disease.

3 Concentration Molar concentration (M) It is the most commonly expressed in unit of moles of solute per litter of solution. M=(No. of mole)/(volume(litter)) No. of mole=wt/ M.wt Equivalent concentration (N) N=(Wt/eq.wt) x (1/volume(litter)) Eq.wt= Mwt/f eq Equivalence factor (F eq) = number of [H + ] or [OH - ] or No. of electrons that oxidizing or reducing agent can accept or donate.

4 The water ionization constant (K w ) H 2 O + H 2 O <==> H 3 O + + OH K w = [H 3 O + ] [OH ]= 1.x ph = -log [H 3 O + ] Calculate the (N)concentration of HCL, Specific gravity=1.19, percent assay (36%) and the Mwt of HCL=36.46g/mole Eq.wt= M.wt/f eq =36.46/1=36.46 M=(sp.gr)(percnt assay)(1000)/m.wt M=(1.19)(36/100)(1000)/36.46= N Then, diluted solution can be prepared by N1XV1=N2XV2

5 Buffer solution A buffer is a solution that resist change in ph following the addition of an acid or base. A buffer can be created by mixing a weak acid (HA) with it is conjugate base (A ). Maximum buffering capacity occurs at ph equal to the pka, but a conjugate acid-base pair can still serve as an effective buffer when the ph of solution is within approximately ±1 ph unit of the pka.

6 Derivation of the equation HA <===> H + + A Ka=[H + ] [A ]/[HA] -logka = -log[h + ][A ]/[HA] ph = pka+log[a ]/[HA] (The Henderson-Hasselbalch equation)

7 Buffer solution

8 Ionization of amino acids Ionic states of amino acids deped on α-carboxyl and α- amino group and any ionizable groups in the side chains. ph<pka (ionizable group) =amino acid protonated state predominate (true acid, proton donor) ph>pka (ionizable group) = amino acid unprotonated form of that group predominates (conjugate base, proton acceptor). The ionic states of amino acids side chains affect the 3D structure of proteins.

9 Ionization of amino acids Also, the ionic properties of amino acids helps in understanding enzyme mechanisms since the ionizable amino acid residues are involved in enzyme catalysis. pka values of the ionizable side chaines can be determined from the amino acid titration curve. If the amino acid has two ionizable groups it has 2 pka values. If it has three ionizable groups it has three pka values. Each pka value is associated with a buffering zone where it has low ph change effect.

10 At Its Isoelectric ph (pi), an Amino Acid Bears No Net Charge The isoelectric (PI) is the form of a molecule that has an equal number of positive and negative charges and thus is electrically neutral. pi is also the ph midway between the pka values on either side of the isoionic species. PI is calculated from : PI=(pK1+Pk2)/2

11 Properties of amino Acids I

12 Properties of amino Acids II

13 Titration curves of glycine Titration curve of glycine resembles acetic acid (pka=4.8) at stage 1but the carboxyl group of glycine is over 100 times more acidic. Fully protonated Zwitterion Fully deprotonated Buffering Zone 2 Corresponds to the -NH3+ ph=pka Maximum capacity Titration ends at ph12 where (-1) form is the predominant. Buffering Zone 1 Corresponds to the -COOH ph=pka Maximum capacity Equimolar concentration of the proton donor (+1) and proton acceptor(0) species are present pi: Isoelectric point (dipolar) pi= (pk2+pk1)/2 pi= ( )/2 pi= 11.94/2 pi=5.97

14 Titration curves of acidic a.a. Fully protonated Zwitterion Fully deprotonated Titration curve of glutamate is more complex, it has three stages corresponding to three possible ionization steps, thus have three pka. Two pka for two COOH and one pka for -NH3 Note: R is for the R group The isoelectric point reflects the nature of R group present. pi: Isoelectric point pi= (pkr+pk1)/2 pi= ( )/2 pi= 3.22

15 Titration curves of basic a.a Fully protonated Zwitterion Deprotonated Histidine has two positively charged groups when it is protonated. Histidine only has R group with pka =6, which provides significant buffering power extra and intracellular fluids. The Ip reflects the pka values of amino and imidazole groups. pi: Isoelectric point pi= (pkr+pk2)/2 pi= (6+9.17)/2 pi= 7.59

16

17 Ionization behavior of peptides Peptides contain one free a-amino acid and one free a- carboxyl group and they get ionized as in free amino acids. On the other hand, a amino acids and a carboxyl groups in all non terminal amino acids are covalently joined within the peptide bond, which don t ionize, so do not participate to the total acid base behavior of peptides. However, the R group of some amino acids can ionize and contribute to the overall acid base behavior.

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