Lecture 7 & 8 Medicinal Chemistry 1 PC 509 Prof. Dr/ Ghaneya Sayed Hassan ghanmoom2@yahoo.com

Antibacterial Sulfonamides Discovery: In 1932: Gerhard Domagk studied the antimicrobial effect of Prontosil Dye "brilliant red Dye" it was found to be active ≠ Streptococcal infection in mice [in vivo] but inactive on bacterial culture [in vitro]. In 1935: Jacques Tréfouël discovered the conversion of inactive prontosil dye -in vivo- into active Sulfanilamide "Lead compound or Prototype". This finding was confirmed by isolating free sulfonamide from blood & urine of patients treated with Prontosil.

Antibacterial Sulfonamides denote 3 different cases: N.B: Side effects & resistance to sulfonamides limit its use today; Penicillins were excellent alternatives for sulfonamides. Antibacterial Sulfonamides denote 3 different cases: [3] Non-aniline sulfonamides [2] Prodrugs giving active sulfonamide [1] Aniline-substituted sulfonamide Mafenide acetate Sulfasalazine Sulfanilamides All sulfanilamides are sulfonamides, but not all sulfonamides are sulfanilamides

Mechanism of action: Sulfanilamides are active BACTERIOSTATIC. Many bacteria are impermeable to folic acid, so they rely on their ability to synthesize folate from PABA “p-Amino Benzoic Acid”, Petridine & Glutamate against MAMMALS who can't synthesize folic acid, so obtained from diet & so not affected by sulfanilamides [selective chemotherapy]. Because of their structural similarity to PABA, sulfonamides act as competitive inhibitor with this substrate for the enzyme dihydropteroate synthetase. Thus  synthesis of folic acid  thymidine, purine synthesis   synthesis of DNA   multiplication & growth of m.o.

Biosynthesis of folate co-enzymes: By using sulfonamides

This mechanism is supported by: 1- PABA added to culture media antagonizes effect of sulfonamides. 2- Man can't form folic acid  so his cells are immune to sulfonamides. 3- M.O. which can utilize performed folic acid is less sulfonamides susceptible. Resistance of m.o. to sulfonamide drugs is by: The bacterial cell wall becomes more permeable to folic acid. The m.o. learns to utilize preformed folic acid. The organism develop alternate pathway for synthesis of folic acid.  PAPA synthesis by m.o. to overcome inhibition of dihydropteroate synthetase.

Adverse effects of sulfonamides therapy: (1) Gastrointestinal distress. (2) Hemolytic anemia. (3) Hepatitis. (4) Stevens-Johnson Syndrome [sever skin eruption]. (5) Crystalluria. General synthesis of Sulfa drugs:

Structure Activity Relationship [SAR]: N.B: As structure become more close to PABA  more active.

Main characters of Sulfonamides: [1] They are weak organic acids  due to SO2NH2 group [by loss of proton & stabilization of –ve charge by resonance], & this determine pka of the drug. N.B: acidity  by attachment of e-withdrawing group to N1 [2] Metabolism: By N4-acetylation  sulfanilamides excreted as it’s N4-acetate and glucuronide [both two metabolites are inactive] N4-acetate is less water soluble than parent drug   tendency of crystalluria

Crystalluria: (1) If pH of urine = pka of drug  ionized/unionized = 1 Those sulfanilamides and their acetyl derivatives are sparingly soluble in water & excreted almost in urine precipitation in kidney  crystalluria. According to the following equation: (1) If pH of urine = pka of drug  ionized/unionized = 1 (2) If pka of drug > pH of urine  unionized/ionized >1   solubility (3) If pka of drug < pH of urine  unionized/ionized <1   solubility (4) pH of urine is about 6 & pka of sulfanilamide “prototype’ is 10.4  drug present in urine in unionized form   solubility  crystalluria.

So, it’s present mainly in ionized form  more soluble Question: if pka of sulfisoxazole is 5, determine its risk of crystalluria. So, it’s present mainly in ionized form  more soluble   risk of crystaluria. N.B. Sulfanilamides drugs are AMPHOTERIC in nature [with acidic & basic characters] So, in when we make urine alkaline  we  solubility [ ionized form]

To solve problem of crystalluria: Drinking large amount of water   urine flow by  rate of glumerular filtration. (2) Combination therapy [triple therapy]: using mixed sulphonamides [3 sulpha drugs: Sulfadiazine + Sulfamerazine + Sulfamethazine]  only 1/3 of the amount of each drug is used giving the same bacterial action but each one is present in amount less that its solubility product  no precipitation. (3)  pH of urine by alkalinization [using NaHCO3] (4)  pka of drug by N1-substitution with electron-withdrawing group [as heterocycle or acyl group]

Assay: [1] Methods depends on primary aromatic amino group [ N4 ]: Diazotization and diazocoupling. [2] Methods depend on acidity of sulfonamide group [ -SO2NH-R] [a] Non-aqueous titration [b] Argentometric method [Back titration] [3] Bromometric method - Add known excess of standard Br2 solution in HCl  bromination of sulphonamides. - Excess Br2 determined by adding KI  I2  titrated ≠ standard Na2S2O3 e.g. Sulfadiazine

[i] Systemic Sulfonamides Classification of antibacterial sulfonamides [i] Systemic Sulfonamides - Used in treatment of systemic infections. - Classified according to rate of excretion [t1/2] into: LONG ACTING MEDIUM ACTING SHORT ACTING Taken every 24 hrs Taken every 8-12 hrs Taken every 6 hrs With slow excretion rate t1/2 = 10-24 hrs t1/2 < 10 hrs

Systemic Sulfonamides classified into: [1] N1-Acyl Derivatives Sulfacetamide Synthesis:

Sulfacetamide Sodium Water soluble, its solution with pka 5.4 Its Na salt is less alkaline than Na salts of other sulfonamides  non-irritant to mucous membrane  used as eye drops till 10 % concentration. Can be used for urinary tract infection [why?]  it's highly soluble with t1/2 = 7 hrs [rapid excretion] Sulfacetamide Sodium Sodium salt of N-[(4-aminophenyl) sulfonyl] acetamide OR [N1-acetyl sulfanilamide]

[2] N1-Heteroatomic Derivatives [i] Pyrimidine Derivative t1/2 = 7 hrs / pka = 7.2 More water soluble > sulfadiazine & sulfamerazine in acidic urine [pH=5.5], but lower activity in vitro & in vivo. Used in combination sulfa [Tri-sulfapyrimidine therapy] Short Acting Sulfamethazine N1( 4,6-dimethyl- 2- pyrimidinyl ) Sulphanilamide

Moderate Acting Long Acting Sulfadiazine Sulfamerazine t1/2 = 17 hrs / pka = 6.3 Broad spectrum, the drug of choice in UTI. NaHCO3 is co-given [why?] Uses: Na salt as 5 % solution: In Meningitis. Ag salt: topically in burns. Sulfadiazine N1(2- pyrimidinyl ) Sulfanilamide Long Acting t1/2 = 27 hrs / pka = 6.99 Similar properties to sulfadiazine, but: With more water solubility. More absorbable. Less excretion rate. Higher blood level can be obtained with a similar dose. Sulfamerazine N1(4-methyl-2- pyrimidinyl ) Sulfanilamide

Long Acting Sulfameter Sulfadimethoxine Long duration of action due to presence of OCH3   plasma protein binding   excretion rate. As a result of  excretion  may cause hypersensitivity upon accumulation. T1/2 of Sulfameter is 37-48 hrs & for Sulfadimethoxine is 40 hrs. Sulfameter N1(5-methoxy -2- pyrimidinyl ) Sulphanilamide Sulfadimethoxine N1(2,6-dimethoxy-4- pyrimidinyl) Sulphanilamide

[ii] Pyridazine Derivative Sulfamethoxypyridazine acetyl N1-acetyl-N1(6-methoxy-3-pyridazinyl) Sulphanilamide N1(6-methoxy-3-pyridazinyl) Sulphanilamide PRODRUG for sulfamethoxypyridazine   bitter taste  used for pediatrics. Inactive in vitro  activated by deacetylation in intestine. t1/2 = 37 hrs  long acting [why?] , due to presence of methoxy group. With bitter taste.

Sulfamethoxazole Sulfaisoxazole [iii] Isoxazole Derivative N1(5-methyl-3-isoxazolyl) Sulphanilamide ● t1/2 = 11 hrs  short acting. ● Not rapidly absorbed as sulfisoxazole  its peak blood level is only about 50 %. Sulfaisoxazole N1(3,4-dimethyl-5-isoxazolyl) Sulphanilamide ● t1/2 = 6 hrs  short acting. ● Rapidly absorbed. ● Highly water soluble  no need for using alkalinizing agent with it. ● With bitter taste. pka of sulfisoxazole is 5

[iv] Thiadiazole Derivative Sulfaethidole Sulfaimethizole N1(5-ethyl-1,3,4-thiadiazol-2-yl) Sulphanilamide N1(5-methyl-1,3,4-thiadiazol-2-yl) Sulphanilamide IUPAC name: 4-amino-N-(5-methyl-1,3,4-thiadiazol-2-yl)- benzenesulfonamide ● Moderate duration. ● Has the lowest degree of acetylation of sulfonamides ● t1/2 = 2 hrs  short acting. ● Highly soluble used in UTI.

[2] Topical Sulfonamides Sulfadiazine Silver [Silvadene®, Dermazine] Applied in water-miscible cream [Dermazine]  active topically ≠ Pseudomonas species  used in burn therapy [that Pseudomonas is responsible for failure of therapy]. Slightly soluble, Not penetrate cell wall but act on external cell structures. Prepared by mixing equimolar amount of AgNO3 & Na sulfadiazine [both dissolved in water].

Sulfacetamide (Klaron®, Ovace®) Sulfacetamide 10% topical lotion, is approved for the treatment of acne and seborrheic dermatitis. (e.g., seborrheic dermatitis, seborrhea sicca [dandruff]); also indicated for the treatment of secondary bacterial infections of the skin due to organisms susceptible to sulfonamides

Mafenide acetate (Sulfamylon) 4-(Aminomethyl)benzenesulfonamide Not true sulfanilamide compound Not inhibited by PABA [its M.O.A. involves different mechanism than true sulfonamides]. Effective ≠ Clostridium welchii in topical use for infected wounds. Not effective orally. Used alone or with antibiotics in treatment of slow healing infected wounds. If used in large quantities  metabolic acidosis. So, a series of new organic salts was prepared. The acetate derivative in ointment base is the most efficient.

[3] Non-absorbable Sulfonamides [i] Topical Sulfonamides: Sulfadiazine Ag & Mafenide acetate. [ii] Intestinal Sulfonamides They are Prodrugs designed to be poorly absorbed “from small intestine”  In large intestine  cleavages  free sulfonamide "active". Used in treatment of intestinal infections, ulcerative colitis & reduction of bowel flora. Sulfa guanidine N1-amidino sulfanilamide Poorly absorbed [with additional basic group]  not absorbed from GIT  high local concentration Synthesis:

N4-substituted Sulfonamides Phthalyl sulphacetamide Phthalyl sulphathiazole Succinic sulphathiazole N4-acetyl-N4-phthalyl sulfanilamide 2-(N4-phthalyl sulfanilamide) thiazole 2-(N4-succinyl sulfanilamide) thiazole They have additional acidic group  poorly absorbed from GIT. They are prodrugs  activated in vivo by slow hydrolysis giving  local concentration.

5-[4-(2-pyridyl sulfamoyl) phenyl azo] salicylic acid Sulfasalazine Water insoluble, broken in body giving: 5-amino saicyclic acid [anti-inflammatory] + Sulfapyridine [carrier]  so, USED IN ULCERATIVE COLITIS. 5-[4-(2-pyridyl sulfamoyl) phenyl azo] salicylic acid Synthesis:

Trimethoprim Combination of sulfonamides with Dihydrofolate Reductase Inhibitor Remember  :  THFA synthesis   purine & pyrimidine bases synthesis   DNA synthesis  stop growth of bacteria. Trimethoprim

Advantages of this combination: It's potent anti-bacterial by inhibition of DHFR enzyme   stop bacterial growth. Selective in action  100.000 times more active ≠ bacterial DHFR relative to mammalian DHFR. [used for UTI] Combined with Sulfamethoxazole as both have the same pharmacokinetic properties [t1/2 = hrs like that of sulfamethoxazole]  excreted at about the same time. important condition for combination between two drugs] Trimethoprim combined with Sulfamethoxazole [Sutrim] [Septazole] [Septrin] The combination of a sulfonamide with trimethoprim causes a sequential blockade of folic acid synthesis. Advantages of this combination: (1) Synergism due to sequential blockage. (2) Avoid development of resistance. (3) Broader spectrum of activity.

(Sulfadoxine/pyrimethamine) Fansidar (Sulfadoxine/pyrimethamine) Combination of Antibacterial Sulfadoxine [500 mg] + antimalarial Pyrimethamine [25 mg] Inhibit folic acid synthesis by two different ways. With schizonticidal effect. Used for prophylaxis & treatment of chloroquine-resistant P.falciparum