PEGylated Norfloxacin and its antibacterial applicationTechnical field
The present invention relates to pesticide synthesis fields, and in particular to a kind of PEGylated Norfloxacin and its antibacterial application.
Background technology
Norfloxacin (Norfloxacin also known as Noroxin, Fulgram) is third generation Comprecin, is classWhite is to light yellow crystalline powder, odorless, mildly bitter flavor.Exposure is easy to moisture absorption in air, forms semihydrate.Meet it is photochromic graduallyIt deepens.In water or ethyl alcohol, solubility slightly soluble.But it is readily soluble in acid or alkaline solution.
Norfloxacin is to gram-negative bacterias such as Escherichia coli, shigella dysenteriae, Salmonella, proteus, Pseudomonas aeruginosasThere is height antibacterial activity, also has good antibacterial action to gram positive bacterias such as staphylococcus, Diplococcus pneumopniaes.But promise fluorine is huskyStar water solubility is poor, and half-life period is not grown, and side effect is larger.
Referred to as " PEG ", ethylene glycol is through high-molecular compound made of intermolecular dehydrating condensation for polyethylene glycol.According to average molecularQuality is different and property is different, from no color or smell thick liquid to waxy solid.It is liquid under 200~600 room temperature of molecular weightBody, molecular weight just gradually become semi-solid in 600 or more persons, and with the difference of average molecular weight, property is also variant.FromNo color or smell thick liquid is to waxy solid.This product is dissolved in water, ethyl alcohol and many other organic solvents.Due to polyethylene glycolSteam forces down, and stablizes to heat, acid, alkali.It does not work with many chemicals.There are good hygroscopicity, lubricity, caking property.NothingPoison, it is non-stimulated.Therefore there is critically important application prospect in fields such as medicine, material and engineerings.
Invention content
To solve the above-mentioned problems, The present invention gives a kind of technical solutions that PEG and Norfloxacin are connected.The Norfloxacin compound of PEG loads not only has good water solubility, but also substantially prolongs its half-life period, to carryIts high antibacterial and bactericidal activity.
The purpose of the present invention is to provide a kind of preparation method of PEGylated Norfloxacin and its application in antibacterial, PEGChange Norfloxacin, structural formula is：
Wherein, the molecular formula of polyethylene glycol (PEG) is：ForAmino, carboxyl, methoxyl group, halogen, sulfydryl etc., molecular weight are：1000-20000；
Further, A is methoxyl group or halogen；
Further, molecular weight polyethylene glycol 2000-10000.
The synthetic method of PEGylated Norfloxacin, it is characterized in that synthetic route as shown in Figure 1,
Step 1) polyethylene glycol is oxidized to the poly- second of end carboxyl two through TEMPO (2,2,6,6- tetramethyl piperidines-nitrogen-oxide)Alcohol；
Polyethylene glycol acyl chlorides is made in step 2) end carboxyl polyethylene glycol；
PEGylated Norfloxacin is made in step 3) polyethylene glycol acyl chlorides with Norfloxacin in the presence of alkali.
Further, which is characterized in that the step 1) be using dichloromethane as solvent under conditions of it is anti-in 0-30 DEG CTemperature is answered, is carried out under the reaction time of 6-12h.
The material ratio of the step 1) is polyethylene glycol：TEMPO:Sodium hypochlorite：Sodium chlorite=1:0.05:2:2.
Further, the reaction dissolvent of the step 1) has the buffer system of sodium bicarbonate.
Further, which is characterized in that the step 2) be using oxalyl chloride as solvent under conditions of in 60-80 DEG C of reactionTemperature carries out under the reaction time of 6-12h.
Further, which is characterized in that the step 3) be using dichloromethane as solvent under conditions of it is anti-in 0-30 DEG CTemperature is answered, is carried out under the reaction time of 4-10h.
Further, the required alkali of the step 3) is triethylamine.
The invention has the advantages that：
Norfloxacin after 1.PEG modifications, drug water solubility enhancing, increases pharmaceutical activity.
Norfloxacin Increased Plasma Half-life after 2.PEG modifications, reduces formulation rate.
Description of the drawings
Fig. 1 is the synthetic route of PEGylated Norfloxacin.
Specific implementation mode
Preparation method with reference to the conjugate of the embodiment description present invention and its antibacterial application, it does not limit this hairBright, the scope of the present invention is defined by the claims.
Embodiment 1：The preparation of compound 1
(1) addition 100g MPEG2000, the 500ml dichloromethane in 1L flasks, 20ml saturated sodium bicarbonate solutions,0.4gTEMPO.Under ice bath temperature control, 20ml liquor natrii hypochloritis is added dropwise, it is molten to continue dropwise addition 20ml sodium chlorites after stirring 1hLiquid, is warming up to 20 DEG C of stirring 6h, and reaction finishes.92g products are obtained after crystallization for purifying.Yield：92%.Nuclear magnetic data is as follows：1HNMR (400MHz, CDCl3)：δ：3.579~3.540 (m, 182H)；2.423 (t, J=6.8Hz, 2H).
(2) 10g steps (1) product is taken, is added in 50ml oxalyl chlorides, 65 DEG C of reaction 8h are warming up to.After completion of the reaction,It is cooled to room temperature, is directly spin-dried for, 10g products are obtained after anhydrous ether mashing, it can directly in next step, without being further purified.
(3) 10g steps (2) product is added in 100ml dichloromethane, 1.6g Norfloxacins, tri- second of 1.5g is addedAmine.20 DEG C of reaction 4h.After completion of the reaction, 50ml is washed 2 times, and anhydrous sodium sulfate drying is spin-dried for.Crystallization for purifying, anhydrous ether10.5g products are obtained after mashing.Yield：91.3%.Nuclear magnetic data is as follows：1HNMR (400MHz, CDCl3)： δ：7.959 (m,1H)；7.126 (d, J=4.4Hz, 1H)；5.936 (d, J=4.4Hz, 1H)；3.679~3.560 (m, 184H)；3.423 (m,4H)；3.124 (t, J=4.8Hz, 2H)；2.373 (t, J=5.2Hz, 2H)；1.141 (s, 3H).
Embodiment 2：The preparation of compound 2
(1) the addition 100g mpeg 3s 400 in 1L flasks, 500ml dichloromethane, 12ml saturated sodium bicarbonate solutions,0.24gTEMPO.Under ice bath temperature control, 12ml liquor natrii hypochloritis is added dropwise, it is molten to continue dropwise addition 12ml sodium chlorites after stirring 1hLiquid, is warming up to 20 DEG C of stirring 6h, and reaction finishes.93g products are obtained after crystallization for purifying.Yield：93%.Nuclear magnetic data is as follows：1HNMR (400MHz, CDCl3)：δ：3.579~3.540 (m, 310H)；2.423 (t, J=6.8Hz, 2H).
(2) 10g steps (1) product is taken, is added in 50ml oxalyl chlorides, 65 DEG C of reaction 8h are warming up to.After completion of the reaction,It is cooled to room temperature, is directly spin-dried for, 10g products are obtained after anhydrous ether mashing, it can directly in next step, without being further purified.
(3) 10g steps (2) product is added in 100ml dichloromethane, 0.95g Norfloxacins, tri- second of 0.88g is addedAmine.20 DEG C of reaction 4h.After completion of the reaction, 50ml is washed 2 times, and anhydrous sodium sulfate drying is spin-dried for.Crystallization for purifying, anhydrous ether10.4g products are obtained after mashing.Yield：94.3%.Nuclear magnetic data is as follows：1HNMR (400MHz, CDCl3)：δ：7.959 (m,1H)；7.126 (d, J=4.4Hz, 1H)；5.936 (d, J=4.4Hz, 1H)；3.679~3.560 (m, 312H)；3.423 (m,4H)；3.124 (t, J=4.8Hz, 2H)；2.373 (t, J=5.2Hz, 2H)；1.141 (s, 3H).
Embodiment 3：The preparation of compound 3
(1) addition 100g MPEG5000, the 500ml dichloromethane in 1L flasks, 8ml saturated sodium bicarbonate solutions,0.16gTEMPO.Under ice bath temperature control, 8ml liquor natrii hypochloritis is added dropwise, after stirring 1h continues that 8ml sodium chlorite solutions are added dropwise,20 DEG C of stirring 6h are warming up to, reaction finishes.94g products are obtained after crystallization for purifying.Yield：94%.Nuclear magnetic data is as follows：1HNMR (400MHz, CDCl3)：δ：3.579~3.545 (m, 454H)；2.425 (t, J=6.0Hz, 2H).
(2) 10g steps (1) product is taken, is added in 50ml oxalyl chlorides, 65 DEG C of reaction 10h are warming up to.After completion of the reaction,It is cooled to room temperature, is directly spin-dried for, 10g products are obtained after anhydrous ether mashing, it can directly in next step, without being further purified.
(3) 10g steps (2) product is added in 100ml dichloromethane, 0.64g Norfloxacins, tri- second of 0.6g is addedAmine.20 DEG C of reaction 6h.After completion of the reaction, 50ml is washed 2 times, and anhydrous sodium sulfate drying is spin-dried for.Crystallization for purifying, anhydrous ether10.0g products are obtained after mashing.Yield：94.0%.Nuclear magnetic data is as follows：1HNMR (400MHz, CDCl3)： δ：7.959 (m,1H)；7.126 (d, J=4.4Hz, 1H)；5.936 (d, J=4.4Hz, 1H)；3.679~3.560 (m, 456H)；3.423 (m,4H)；3.124 (t, J=4.8Hz, 2H)；2.373 (t, J=5.2Hz, 2H)；1.141 (s, 3H).
Embodiment 4：The preparation of compound 4
(1) addition 100g MPEG10000, the 500ml dichloromethane in 1L flasks, 4ml saturated sodium bicarbonate solutions,0.08gTEMPO.Under ice bath temperature control, 4ml liquor natrii hypochloritis is added dropwise, after stirring 1h continues that 4ml sodium chlorite solutions are added dropwise,20 DEG C of stirring 6h are warming up to, reaction finishes.91g products are obtained after crystallization for purifying.Yield：91%.Nuclear magnetic data is as follows：1HNMR (400MHz, CDCl3)：δ：3.579~3.540 (m, 910H)；2.423 (t, J=6.8Hz, 2H).
(2) 10g steps (1) product is taken, is added in 50ml oxalyl chlorides, 80 DEG C of reaction 12h are warming up to.After completion of the reaction,It is cooled to room temperature, is directly spin-dried for, 10g products are obtained after anhydrous ether mashing, it can directly in next step, without being further purified.
(3) 10g steps (2) product is added in 100ml dichloromethane, 0.32g Norfloxacins, tri- second of 0.3g is addedAmine.20 DEG C of reaction 4h.After completion of the reaction, 50ml is washed 2 times, and anhydrous sodium sulfate drying is spin-dried for.Crystallization for purifying, anhydrous ether10.0g products are obtained after mashing.Yield：96.9%.Nuclear magnetic data is as follows：1HNMR (400MHz, CDCl3)： δ：7.959 (m,1H)；7.126 (d, J=4.4Hz, 1H)；5.936 (d, J=4.4Hz, 1H)；3.679~3.560 (m, 912H)；3.423 (m,4H)；3.124 (t, J=4.8Hz, 2H)；2.373 (t, J=5.2Hz, 2H)；1.141 (s, 3H).
Embodiment 5：The antibacterial experiment in vitro of each target compound
1) test strain be chicken colibacillosis, swine escherichia coli, pig staphylococcus aureus, chicken staphylococcus aureus,Streptococcus；
2) bacteriostatic test is done with Beating holes method, measures inhibition zone size.Bacteriostatic diameter >=16mm is Gao Min, antibacterial straight11~15mm of diameter is quick in being, bacteriostatic diameter≤10mm is muting sensitive or drug resistance.
3) preparation of bacterium solution：Bacterial strain is accessed into broth bouillon, sets 37 DEG C of cultures for 24 hours.
4) preparation of antibacterial liquid：1mg drugs are dissolved in 20ml sterile waters, are shaken up, 5% concentration is made.
5) tablet culture：Cultured solution of broth and antibacterial liquid are pressed 1:1 mixing is placed on culture medium, then takes 0.1ml'sBacterium solution is added thereto, and sets 37 DEG C of cultures for 24 hours.Do two groups of parallel tests.
1 unit of table (mm)
As seen from table, PEGylated Norfloxacin compound and Norfloxacin all have gram-negative bacteria and gram positive bacteriaSensibility, 5 kinds of bacterial strains all show as hypersensibility to each compound.Chicken colibacillosis has highest sensibility, pig to compound 2Staphylococcus aureus has highest sensibility to compound 1.Each PEGylated Norfloxacin compound is relative to NorfloxacinBacteriostatic activity have advantage.