Optimizacija kromatografskih metod pri večstopenjski kemijski sintezi desmetoksisulfazecina

Sever, Vid (avtor), Gobec, Stanislav (mentor), Krajnc, Alen (komentor)

Naraščajoča odpornost bakterij na antibiotike predstavlja eno največjih groženj javnemu zdravju. β-Laktami, ki ostajajo najpogosteje uporabljen strukturni razred antibiotikov, zaradi svoje prekomerne in pogosto neustrezne uporabe izgubljajo svojo učinkovitost napram bakterijam, ki so razvile učinkovite mehanizme odpornosti proti njim. Posebej problematična je odpornost na β-laktame posredovana z β-laktamazami. Medtem ko so proti serinskim β-laktamazam na voljo učinkoviti zaviralci, zdravljenje okužb, ki jih povzročajo bakterije z metalo-β-laktamazami, ostaja omejeno predvsem na uporabo monobaktamov, ki jih metalo- β-laktamaze (še) ne morejo hidrolizirati. Edini predstavnik te skupine v klinični uporabi je aztreonam, medtem ko je sulfazecin edini znani monociklični β-laktam naravnega izvora z protibakterijskim delovanjem, ki ga v naravi proizvaja bakterija Pseudomonas acidophila. Njegov biosintetski prekurzor je desmetoksisulfazecin. V magistrski nalogi smo sintetizirali desmetoksisulfazecin po večstopenjski sintezni poti, pri čemer je bil glavni poudarek na razvoju in optimizaciji kromatografskih metod za učinkovito izolacijo ključnih intermediatov in končnega produkta zadostne čistosti za nadaljnje študije. C3 stranko verigo smo pripravili iz komercialno dostopnega Z-D-Glu-Obzl, ki smo ga najprej spojili z D-alanin terc-butil estrom. Temu je sledila odstranitev terc-butilne zaščitne skupine in optimizacija metode kromatografske izolacije diastereoizomerno čiste karboksilne kisline 2. Vzporedno smo pripravili tudi monociklični β-laktamski gradnik 9 s prosto amino skupino, ki smo ga sintetizirali v sedmih korakih iz L-serina. Po uspešni tvorbi amidne vezi med fragmentoma 2 in 9, odstranitvi zaščitnih skupin s hidrogeniranjem ter optimizaciji kromatografskih pogojev smo uspešno izolirali desmetoksisulfazecin 11. Optimizacija kromatografskih pogojev je omogočila učinkovitejše ločevanje polarnih intermediatov in izolacijo končnega produkta zadovoljive izomerne čistosti. Kemijsko strukturo desmetoksisulfazecina smo potrdili z NMR in masno spektrometrijo, končnemu produktu pa smo ovrednotili tudi minimalne inhibitorne koncentracije (MIK) proti izbranim bakterijskim sevom.

• Jezik: Slovenski jezik
• Ključne besede: ß-laktamski antibiotiki, ß-laktamaze, desmetoksisulfazecin, monobaktami
• Vrsta gradiva: Magistrsko delo/naloga
• Organizacija: FFA - Fakulteta za farmacijo
• Leto izida: 2026
• PID: 20.500.12556/RUL-181208
• Datum objave v RUL: 27.03.2026

Sekundarni jezik

• Jezik: Angleški jezik
• Naslov: Optimisation of chromatographic methods in multistep chemical synthesis of desmethoxysulfazecin

Izvleček

The increasing prevalence of multi-drug-resistant bacteria represents one of the major threats to public health. β-Lactams remain the most widely used structural class of antibiotics; however, their effectiveness is decreasing due to excessive and often inappropriate use, which has led to the emergence of bacterial resistance mechanisms. Particularly problematic is resistance to β-lactams mediated by β-lactamases. While effective inhibitors are available against serine β-lactamases, the treatment of infections caused by bacteria expressing metallo-β-lactamases remains largely limited to the use of monobactams, which metallo-β-lactamases are unable to hydrolyse. The only clinically used representative of this class is aztreonam, whereas sulfazecin is the only known naturally occurring monocyclic β-lactam with antibacterial activity, produced by the bacterium Pseudomonas acidophila. Its biosynthetic precursor is desmethoxysulfazecin. In this master’s thesis, desmethoxysulfazecin was synthesised via a multistep synthetic route. Emphasis was, however, on the development and optimisation of chromatographic methods enabling efficient isolation of desmethoxysulfazecin in sufficient purity for further studies. The requisite C3 side chain was prepared from commercially available Z-D-Glu-Obzl, which was first coupled with D-alanine tert-butyl ester. This was followed by removal of the tert-butyl protecting group and optimisation of chromatographic conditions for the isolation of diastereomerically pure carboxylic acid 2. In parallel, the monocyclic β-lactam building block 9 bearing a free amino group was prepared in seven steps starting from L-serine. After successful formation of the amide bond between fragments 2 and 9, removal of protecting groups by hydrogenation, and optimisation of chromatographic purification conditions, desmethoxysulfazecin (11) was successfully isolated. Overall, optimisation of chromatographic conditions enabled more efficient separation of polar intermediates and isolation of the final product with satisfactory stereoisomeric purity. The chemical structure of desmethoxysulfazecin was confirmed by NMR and mass spectrometry, and the minimum inhibitory concentrations (MIC) of the final product against selected bacterial strains were also determined.

• Ključne besede: ß-lactam antibiotics, ß-lactamases, desmethoxysulfazecin, monobactams