CN1181381A - Synthesis and application of levofloxacin analogs - Google Patents

Abstract

A process for preparing levo-ofloxacin and its likes is disclosed with the advantages of easily available raw materials, gentle reactio conditions, high total output rate, and high optical active purity. After structural modification at positions 5 and 7 of levo-oxyxacin, the products are obtained, which has distinct antibacterial and anti-cancer activity.

Description

Synthesis and application of levofloxacin analogs

The present invention relates to the chemical synthesis of levofloxacin and its analogs and their biological activity, more specifically to the preparation method of levofloxacin and the preparation and application of levofloxacin analogs.

Fluoroquinolone compounds have achieved great success in clinical anti-infective treatment due to their high-efficiency, broad-spectrum, and low-toxicity antibacterial properties. Levofloxacin (I) developed by Japan's Daiichi Pharmaceutical Company is an example. The drug has a broad antibacterial spectrum and low toxicity. Its antibacterial activity is twice that of ofloxacin, and its water solubility is increased by 10 times, but its synthetic route is very complicated EP309789 (1989)

The synthesis of levofloxacin mainly contains the following methods:

1. Resolution method: including high-pressure liquid phase analysis EP 206283 (1986) and enzymatic analysis (K. Sakano, Agnia Biol. Chem. 1987, 51, 1265). This method is not suitable for large-scale preparation.

2. With (S)-2-aminopropanol as the chiral starting material, levofloxacin U.S.P.4777253 is obtained from tetrafluorobenzoic acid through 7 steps of reaction, (1988). This method is more efficient than the resolution method It is suitable for industrial production, but the starting materials tetrafluorobenzoic acid and (S)-2-aminopropanol are not easy to obtain.

3. From trifluoronitrobenzene and (R)-1,2-propanediol and 2-hydroxyl protected derivatives as raw materials, EP368410 (1990) and EP 273399 (1988) were obtained through multi-step reactions. There is also the problem that chiral raw materials are not easily available in this method. Moot.

4. The method of biotransformation, which is relatively complicated. (Tetrahedron Letters, 1996, 9317.)

It can be seen that the above methods all have various restrictions on the large-scale industrial production of levofloxacin with high optical purity, which brings certain difficulties to industrial production.

An object of the present invention is to find out the method that can industrialize production levofloxacin. Another object of the present invention is to carry out structural modification to levofloxacin to find new drugs with better effects.

Levofloxacin derivatives of the present invention can be represented by the following general formula:

where _R7 is

R ₁ =H, C ₁ -C ₅ alkyl; m=1,2

R ₂ = H, C ₁ -C ₅ alkyl

R ₃ ＝C ₁ -C ₅ straight chain or branched chain alkyl, CH ₂ Ph

R ₄ =H, C ₁ -C ₅ alkyl, pyridine, pyrimidine, CO(CH ₂ )nX

X = halogen, n = 1, 2

R ₅ =H, NH ₂

R ₆ = alkyl or substituted phenyl

Levofloxacin and derivatives thereof of the present invention can be made from D-mannitol as a raw material mainly through the following steps:

a. Trifluoronitrobenzene is condensed with (S)-4 under phase transfer catalysis conditions to obtain condensate (S)-5;

b. (S)-5 was deprotected to obtain (R)-6, and (R)-6 was bromoacetylated with acetic acid-hydrogen bromide to obtain (R)-7 and (S)-7;

c. Cyclization of (R)-7 and (S)-7 in sodium methoxide methanol solution to obtain cyclized product (R)-8;

d. (R)-8 was reduced with palladium/carbon, the reduced product was condensed with EMME to obtain the condensate (R)-10, (R)-10 was cyclized and acid hydrolyzed to obtain the key intermediate (S)-13;

e. Condensation of (S)-13 and HR ₇ in an appropriate anhydrous solvent to obtain the above-mentioned levofloxacin and its derivatives substituted at the 7-position;

f. (S)-13 introduces ammonia at the 5-position to obtain 23, and condenses 23 and HR ₇ in a suitable solvent to obtain the above-mentioned levofloxacin derivative substituted with 5-amino group.

g. Condensation of alkyl carbamate, alkyl or substituted phenylsulfonyl chloride, α-haloacetyl chloride and levofloxacin derivative with secondary nitrogen structure at the 7-position to obtain compound 29-60.

The preparation method of levofloxacin and derivatives thereof of the present invention has the advantages of rich and easy-to-obtain chiral starting materials, mild reaction conditions, high total yield, high optical purity, low production cost and the like. In a word the method of this patent is more suitable for industrialized production than literature method.

The present invention is implemented in more detail by the following reaction steps:

a.KOH/K ₂ CO ₃ /toluene b.HCl/CH ₃ COCH ₃ c.HBr/ACOH

d.KOH/H ₂ O e.Pd/C/H ₂ f.EMME/Heating

g.Ph ₃ P/DAD/THF h.PPE/Heating i.ACOH/HCl heating

After D-mannitol is protected by acetone under the catalysis of Lewis acid such as anhydrous zinc chloride, anhydrous aluminum chloride, etc., use sodium periodate in a polar aprotic solvent such as dichloromethane, chloroform or alcohol solvent Such as oxidation in methanol, ethanol, etc., and then reduction with a negative hydrogen ion reducing agent such as potassium borohydride, sodium borohydride, etc. to obtain chiral compound (S)-4. Trifluoronitrobenzene is condensed with (S)- in an aprotic solvent such as toluene, benzene, halogenated hydrocarbons, etc. under phase transfer catalysis conditions to obtain (S)-5. Bromic acid bromoacetylation, using inorganic bases such as sodium hydroxide, potassium hydroxide or organic bases such as methanol to obtain (R)-8, (R)-8 with Pd/C or platinum/carbon in alcohol solvents Catalytic reduction, and then condensation with diethyl ethoxymethylene malonate to obtain (R)-10, and (R)-10 was hydrolyzed by cyclization acid to obtain the key intermediate (S)-13. (S)-13 reacts with methylpiperazine and its derivatives to obtain levofloxacin and its derivatives.

The 7-position substituent has a great influence on the antibacterial activity of quinolones, which is related to the penetration ability of the drug on the bacterial cell membrane and the affinity to DNA, so changing the 7-position substituent may produce a drug with better antibacterial performance, so Condensation of (S)-13 and HR ₇ in anhydrous aprotic polar solvents such as pyridine, dimethylformamide, dimethyl sulfoxide, etc. to obtain the 7-substituted levofloxacin analogue 14- twenty one.

where _R7 is

R ₁ =H, C ₁ -C ₅ alkyl; R ₂ =H, C ₁ -C ₅ alkyl;

R ₃ ＝C ₁ -C ₅ linear or branched chain alkyl, CH ₂ Ph, CH ₂ X, X is halogen

R ₄ =H, C ₁ -C ₅ alkyl or substituted amino, pyridine, pyrimidine, CO(CH ₂ )nX

X = halogen, n = 1, 2

R ₆ = alkyl or substituted phenyl

m=1,2

Usually the introduction of substituents at the 5-position of quinolones can change the biological activity of quinolones. The method of introducing the amino group is to nitrate the 5-position of (S)-13 with nitrate-concentrated sulfuric acid, and the nitrate can be potassium salt or sodium salt. Then catalytic reduction, the catalyst can be Raney (Raney) nickel, palladium/carbon, platinum/carbon and the like. Then condense with HR ₇ in anhydrous aprotic polar solvents such as pyridine, dimethylformamide, dimethyl sulfoxide, etc. to obtain 5-amino-substituted levofloxacin derivatives 24-28.

Synthesis of 7-alkoxycarbonyl substituted quinolones comp.29-48

Synthesis of 7-sulfonyl substituted quinolone comp.49-54 Synthesis of Quinolone-Alkylating Agent Derivatives The structural formulas of the compounds involved in the present invention are shown in Table 1.

Table 1 compound 1,14-60 structural formula

Note: 1, 15, 16 are reference compounds.

Determination of biological activity: 1. Determination of antibacterial activity The determination of antibacterial activity adopts the most suitable Mueller-Hinto (MH) broth medium for general bacteria recommended by the World Health Organization, adding 1.6% agar powder as the matrix for drug dilution. The experimental bacteria were cultured overnight in MH broth medium and diluted to control the inoculum size to 5×10 ³ -5×10 ⁴ CFU. The drug concentration was serially diluted, and added to the medium at 50-60°C and mixed evenly. Prepare double-disc petri dishes, the highest concentration is 12.5μg/ml, and the lowest concentration is 0.025. Due to the different solubility properties of various samples, DMF and ethanol are used as co-solvents for comparison, and diluted with sterile saline. The final co-solvent concentration DMF≤5.0 %, ethanol ≤ 2.5%, the blank control does not affect the growth of each test bacteria. The test bacteria suspension was inoculated in the above-mentioned petri dishes containing various compounds and different concentrations with a multi-head inoculator. Cultivate the petri dish at 35-37°C for 16-18 hours, observe the experimental results with the naked eye, continue to cultivate until 42-44 hours, and then observe the results. The lowest concentration that inhibits the growth of test bacteria is the minimum inhibitory concentration (MIC). The in vitro antibacterial activity (MIC, μg/ml) of levofloxacin derivatives is shown in Table 2. Antibacterial activity (MICμg/ml) of table 2 target compound and reference compound ^* sample number Bacteria name Bacillus subtilis Staphylococcus aureus Staphylococcus epidermidis pneumococcus Klebsiella pneumoniae Pseudomonas aeruginosa Cloacae Escherichia coli Proteus mirabilis 1 0.05 0.2 0.2 0.1 <0.025 0.025 <0.025 1.56 0.05 15 0.8 3.12 0.8 1.56 0.2 0.05 <0.025 0.4 0.05 16 0.2 0.8 0.4 0.2 0.05 0.05 <0.025 0.4 <0.025 17 ND 18 3.12 0.8 0.4 0.2 >6.25 6.25 6.25 >6.25 >6.25 19 0.2 0.2 0.2 0.2 >6.25 >6.25 3.12 >6.25 6.25 20 <0.025 0.2 0.2 0.2 0.05 <0.025 <0.025 0.8 0.2 twenty one 0.8 1.56 0.8 1.56 1.56 0.4 0.1 3.12 0.2 twenty four <0.025 0.05 0.1 <0.025 <0.025 ND <0.025 0.8 ND 25 <0.025 0.2 0.1 26 0.05 0.2 0.1 0.05 0.05 ND <0.05 0.2 ND 27 ND 28 0.4 3.12 0.8 0.4 0.4 ND 0.2 0.8 ND 29 0.1 0.4 0.4 0.2 12.5 ND 3.12 12.5 ND 30 0.1 0.4 0.2 0.1 >6.25 >6.25 6.25 >6.25 >6.25 31 <0.025 0.2 <0.025 0.1 >6.25 >6.25 6.25 >6.25 >6.25 32 0.05 0.2 0.2 0.1 6.25 ND 1.56 >12.5 ND 33 ND 34 0.2 0.4 0.2 0.1 >6.25 >6.25 3.12 >6.25 >6.25 35 0.05 0.4 0.8 0.2 >12.5 ND 6.25 >12.5 ND 36 0.4 0.8 1.56 0.4 >6.25 >6.25 >6.25 >6.25 >6.25 37 0.05 0.4 0.8 0.05 >6.25 >6.25 >12.5 >12.5 >12.5 38 0.05 0.4 0.1 0.2 >6.25 6.25 1.56 >6.25 >6.25 39 0.2 0.8 0.4 0.4 >6.25 >6.25 >6.25 >6.25 >6.25 40 0.1 0.4 0.4 0.2 >6.25 >6.25 6.25 >6.25 >6.25 41 <0.025 0.1 0.2 0.1 6.25 3.12 3.12 >6.25 3.12 41 0.8 0.8 0.2 0.8 >6.25 6.25 1.56 >6.25 3.12 42 0.025 0.8 12.5 0.8 >12.5 >12.5 >12.5 >12.5 >12.5 43 0.05 0.4 0.8 0.2 6.25 6.25 1.56 >6.25 >6.25 44 0.1 0.4 0.4 0.2 6.25 ND 1.56 >12.5 ND 45 0.05 0.8 12.5 0.2 >12.5 >12.5 >12.5 >12.5 >12.5 46 <0.025 0.4 1.56 0.1 >6.25 >6.25 3.12 >6.25 >6.25 47 0.2 0.4 0.2 0.2 >6.25 >6.25 3.12 >6.25 >6.25 48 0.2 0.4 0.4 0.4 >6.25 >6.25 >6.25 >6.25 >6.25 50 0.2 0.2 0.4 0.4 6.25 3.12 0.8 >6.25 6.25 51 0.4 1.56 0.8 0.4 0.05 ND <0.025 0.4 ND 52 0.2 0.2 0.4 0.2 <0.025 <0.025 <0.025 0.4 <0.025 53 0.1 0.4 1.56 0.4 >6.25 >6.25 >12.5 >12.5 >12.5 54 0.025 0.1 <0.025 <0.025 1.56 0.4 0.1 >12.5 0.4 55 1.56 1.56 0.8 1.56 >6.25 >6.25 >12.5 6.25 >12.5 56 <0.025 0.8 0.4 0.2 6.25 3.12 1.56 >6.25 6.25 57 0.1 0.2 0.1 0.1 0.4 ND 0.2 6.25 ND 58 0.2 0.4 0.2 0.4 >6.25 3.12 0.8 >6.25 1.56 59 0.8 6.25 12.5 6.25 >12.5 >12.5 >12.5 >12.5 >12.5 60 <0.025 0.05 0.1 <0.025 1.56 0.8 0.2 >6.25 3.12 61 <0.025 0.4 0.4 0.2 6.25 6.25 1.56 >6.25 6.25 ^* ND means that the MIC value of the compound has not been determined

2. Antitumor Activity Determination

Microculture Tetrozolium (MTT) Metabolic Microculture Tetrozolium Test

Inoculate a certain amount of cancer cells into a 96-well culture plate, dilute to 90 μl with RPMI-1640 culture solution containing 10% calf serum, add 10 μl of drug solution to each well, and the final concentrations are 10 ^-4 , 10 ^-5 , 10 ^-6 Mol/ml, each group has 3 duplicate wells. Incubate for 72 hours at 37°C, 5% CO ₂ . Add 20 μl/well MTT (5 mg/ml) solution, incubate at 37° C. in a 5% CO ₂ incubator for 4-5 hours, then add 100 μl/well triple solution (SDS-isobutanol-hydrochloric acid). Then overnight in a _CO incubator. The OD value was measured with a microplate reader, and compared with the control group, the tumor growth inhibition rate (%) was calculated. See Table 3. Table 3 partial compound tumor inhibition rate % (110r, 10r, 1r) compound tumor cells P388 HL60 N45 47 100 98.8 81 97.1 47.1 95.2 89 91 twenty three 58 98.6 98.6 85.5 97.5 97.5 97.5 84.6 84.6 24.4 59 46.4 62.3 78.3 68.8 85 82.5 23.6 20.5 43.6 60 92.9 95.2 71.4 92.3 93.3 96.2 85 87 19 61 100 96.4 88.1 96.2 96.2 93.2 91 92 50

3. Anti-mycoplasma activity assay

Anti-mycoplasma activity was determined using conventional methods. The composition of the culture medium is as follows: 1:1 bovine heart digestate, 0.5% sodium chloride, 0.15% potassium dihydrogen phosphate, 10% self-made fresh yeast extract, 0.002 phenol red, and 10% calf serum. U _U4 add 0.1% urea, pH=6.0±0.5. Add 0.1% arginine to Mh, Mo, Ms, pH=6.8～7.0. Mg and Mp were increased to 15% by adding 1% glucose serum, pH=7.6～7.8. Add 10% sucrose to spiroplasma, pH=7.2～7.4. The minimum drug concentration MIC (μg/ml) for inhibiting mycoplasma was determined by serial multiple dilution method. The measured values of MIC are shown in Table 4.

Anti-mycoplasma ^* activity MIC (μg/ml) of some compounds in table 4 sample number Mycoplasma U _U4 mh Mo Mrs. MP Mg CH-1 CR-1 sc 18 0.125 0.25 0.125 0.0625 1 1 0.125 0.125 0.0625 19 0.125 0.5 0.25 0.125 1 2 0.25 0.125 0.125 twenty four 4 2 0.25 0.25 2 2 0.5 0.25 25 4 0.25 0.5 0.5 4 8 0.5 1 31 0.25 1 0.5 0.5 2 1 0.125 0.125 0.25 30 0.25 0.5 0.5 0.5 1 2 0.125 0.135 0.0625 34 0.5 0.5 0.25 0.25 4 4 0.5 0.5 0.25 43 0.5 1 2 1 1 2 0.125 0.125 0.0625 44 0.5 0.5 0.5 0.5 1 1 0.125 0.0625 0.0625 46 1 0.5 0.5 0.5 4 8 1 1 47 2 0.25 0.25 0.5 2 4 1 0.5 57 0.25 0.25 0.25 0.5 2 2 0.125 0.125 0.0625 58 4 0.25 0.5 0.5 2 2 0.25 0.5 60 4 0.25 0.125 0.25 4 4 0.5 1 LVFX 2 1 0.5 0.5 0.25 1 0.125 0.25 0.0625

^* U _U4 - Ureaplasma urealyticum; Mh-Mycoplasma hominis; Mo-Mycoplasma oralum; Ms-Mycoplasma salivarius; Spiroplasma honeybee; CR-1 - Spiroplasma rapeseed.

Further illustrate the present invention with embodiment below, it does not limit the present invention. NMR spectra were measured on Bruker AM-400 and AC-100 in our analysis room, and mass spectrometry was performed on MAT-95 mass spectrometer. Elemental analysis was done by the analysis room of Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences. The model of the melting point apparatus was BUeHI510, and the thermometer was not calibrated. Silica gel for column chromatography, 200-300 mesh unless otherwise specified, is produced by Qingdao Ocean Chemical Factory.

Best Example Experiment 1 1,2,5,6-diacetonide-D-mannitol D-2 Add 22.9 g (0.167 mol) of anhydrous zinc chloride to 132 ml of dry acetone under mechanical stirring, and cool down When the temperature is below 15°C, 11.5 g of D-mannitol is added. Stir the reaction at 20°C for 13 hours, place it at 0°C overnight, filter, and stir the filtrate for 5 minutes. Pour 23ml of aqueous solution containing 23g K ₂ CO ₃ into the reaction mixture under vigorous stirring, and a large amount of white precipitates are formed immediately, continue to stir After 30 minutes, filter and wash the filter residue with chloroform (3×25ml). Ammonia water was added to the filtrate to make it alkaline, acetone and water were distilled off to obtain a white solid, which was recrystallized from carbon tetrachloride to obtain needle-like crystal D-2 (9.0 g). Yield 58.0%. mp 120°C. Experiment 2 (S)-1,2-Acetonylated glycerol (S)-47.0gD-2 was dissolved in the mixed solution of 7ml pH=6 phosphate buffer and 40ml THF, and 6.8 gNaIO ₄ . A large amount of white precipitate formed quickly, after 1 hr the pH was adjusted to 8 with 5N NaOH. Filter and wash the filter residue twice with chloroform. Evaporate THF, add 50ml of methanol, cool to below 10°C, filter, and reduce the filtrate with 1.6g KBH ₄ , add in batches so that the temperature does not exceed 10°C. After the addition, react in an ice bath for 30 min., react at room temperature for 1 hr, filter, evaporate methanol, add 20 ml of water, and extract with chloroform. Dry with chloroform, remove the solvent, and distill under reduced pressure to obtain 4.2 g of (S)-4. 96-98°C/25mmHg. Yield 60.8%. Experiment 3 (S)-3,4-difluoro-2-(1,2-acetonide propyloxy)nitrobenzene (S)-50.44g (S)-4 was dissolved in 2ml of toluene, slowly added dropwise To the 10ml toluene mixture containing 0.56gKOH, 0.45gK ₂ CO ₃ , and 0.53g trifluoronitrobenzene cooled in an ice bath, add 20mg of tetra-n-butylammonium chloride, stir for 0.5hr under ice-cooling, and stir at room temperature for 1hr, TLC ( RAC/petroleum ether=1:5) to detect the end point of the reaction. After the reaction was completed, 10ml of water was added, the toluene was separated, the water phase was extracted with toluene (2×20ml), the toluene phase was combined, dried, and the toluene was evaporated to obtain (S)-50.86g, with a yield of 100%. [α] _D ²⁰ = -7.36. MS m/e = 274 (M ⁺ -15). ¹ H-NMR (CDCl ₃ ) δppm 1.42-1.46 (6H, m), 3.20-4.40 (5H, m), 7.00 (1H, m), 7.62 (1H, m). Experiment 4 (S)-3,4-difluoro-2-(1,2-dihydroxypropyloxy)nitrobenzene (R)-62.60g (S)-5 was dissolved in 10ml of acetone, and 7ml of 1N hydrochloric acid was added , Hydrolysis at 60°C for 1 hr. The acetone was evaporated, and the residue was extracted with chloroform (5×20ml), dried and concentrated to obtain (R)-6 (2.25g). The yield was 100%. [α] _D ²⁰ =13.34. ¹ H-NMR (CDCl ₃ ) δppm 3.68-3.88 (2H, 2×m, J=5), 4.12 (1H, b), 4, 28-4.47 (2H, m, 2×m, J=6), 7.05 (1H, m, J=8), 7.05 (1H, m, J=2). Experiment 5 (S)-3,4-difluoro-2-(1-bromo-2-acetoxypropyloxy)nitrobenzene and (R)-3,4-difluoro-2-(1-acetyl Oxygen-2-bromo)nitrobenzene (S)-7 and (R)-71.91g (97.67mmol) (R)-6 and 4.68g acetic acid-hydrobromic acid were stirred for 5min. under ice-cooling, at room temperature Continue to react for 30-40min.,. Add 5ml of water, adjust to pH=7 with 5N NaOH, extract with ether (3×20ml), dry the ether solution, distill off the ether to obtain (S)-7 and (R)-7 (2.70g), yield 100 %. MS m/e = 274 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 2.12 (3H, s), 3.55-3.75 (2H, m, 2×m, J=6Hz), 4.50 (2H, m), 5.30 (1H, m), 7.05 (1H , m, J=8Hz), 7.70 (1H, m, J=2Hz). Experiment 6 (R)-3,4-difluoro-2-(1,2-epoxypropyloxy)nitrobenzene (R)-81.60g (S)-7 and (R)-7 with 3N KOH 10ml of the solution was cyclized, stirred at 0-40°C for 10min, the reaction mixture was diluted with ether, the ether layer was washed with water, dried, and the solvent was removed to obtain (R)-8 with a yield of 100%. [α] _D ²⁰ =11.93 (0.0237, CDCl ₃ ). MS m/e = 231 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 2.65-2.90 (2H, m, 2×m, J=4Hz), 3.40 (1H, b), 4.15-4.48 (2H, m, 2×m, J=6Hz), 5.30 (1H, m), 7.02 (1H, m, J=8Hz), 6.78 (1H, m, J=3Hz). Experiment 7 (R)-3,4-difluoro-2-(2-hydroxypropyloxy)aniline (R)-91.00 (R)-8 was dissolved in 60ml of absolute ethanol, and 10% Pd/C 0.48 was added g, hydrogenation at normal pressure and temperature for 3.5 hr, and the catalyst was filtered off. Ethanol was distilled off to obtain (R)-9 with a yield of 100%. It was directly used in the next reaction without purification. Experiment 8(R)-3,4-difluoro-2-(2-hydroxypropyloxy)-1-(2,2-diethoxycarbonylvinylamino)benzene(R)-100.76g(R)- 9 and 0.82g EMME were reacted at 140-145°C for 1.5hr, and then the ethanol was generated by vacuum extraction, and the reaction was continued for 0.5hr, cooled, and recrystallized with petroleum ether/ethyl acetate=1:4 to obtain (R)-10, which was recovered The rate is 95%. MS m/e = 375 (M ⁺ ). mp52-54°C. ¹ H-NMR (CDCl ₃ ) δppm 1.22-1.45 (9H, m), 3.45 (1H, d, J=4.0Hz), 3.88-4.43 (7H, m), 6.78-7.08 (2H, m), 8.48 ( 1H,d,J=14Hz). Experiment 9 (S)-diethyl-(7,8-difluoro-3-methyl-3,4-dihydro-2H-[1,4]benzoxazine-4)methylenemalonic acid Ester (S)-110.67gDAD and 0.78gPh3P were dissolved in 12ml of anhydrous THF, stirred at 0°C for 20min., 0.79g (R)-10 was dissolved in 2ml of anhydrous THF, slowly added dropwise to the above mixture, added Stir overnight at room temperature. The solvent was distilled off, and the residue was subjected to column chromatography, eluent RAC/petroleum ether=1:3, to obtain (S)-11 with a yield of 90%. ¹ H-NMR (CDCl ₃ ) δppm 1.20-1.24 (9H, m), 3.90-4.40 (7H, m), 6.71-6.78 (2H, m), 7.76 (1H, s). Experiment 10 Ethyl (S)-(-)-9,10-difluoro-2,3-dihydro-3-methyl-7-oxo-7H-pyrido[1,2,3-de][ 1,4] Benzoxazine-6-carboxylate ester (S)-121.33(S)-11 and 5g PPE were reacted at 140-145°C for 1.5hr, and the reaction was continued under reduced pressure for 0.5hr, cooled to room temperature, poured into ice In water, extract with chloroform (3×30ml), wash chloroform with 5% sodium carbonate solution (3×20ml), wash with water (1×20ml), dry, drive off the solvent, and wash the solid matter with ethanol until grayish white (S) -12, yield 85%. Without further purification, it was directly used in the next step of reverse experiment 11(S)-(-)-9,10-difluoro-2,3-dihydro-3-methyl-7-oxo-7H-pyrido[ 1,2,3-de][1,4]benzoxazine-6-carboxylic acid (S)-130.73g (S)-12, 2.27ml concentrated hydrochloric acid, 8.7ml glacial acetic acid, reflux for 3hr, cool, with or without Color needle-like crystals were precipitated, filtered, and the precipitates were washed with water, ethanol and ether in turn, and dried to obtain (S)-13 (0.56g), with a yield of 98%. mp>300°C. [α] _D ²⁰ = -64.7 (0.01, DMSO). MS m/e = 281 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm 1.85 (3H, d, 2×m, J=7Hz), 4.60-4.48 (2H, 2×m), 5.25 (1H, m), 7.87 (1H, d), 9.40 (1H, s). Experiment 12 (S)-(-)-9-fluoro-2,3-dihydro-3-methyl-10-(4-methyl-1-piperazinyl)-7-oxo-7H-pyrido [1,2,3-de][1,4]Benzoxazine-6-carboxylic acid (I) Levoflloxacin. 84.3mg (S)-13 was dissolved in 3ml of pyridine, 0.13ml of N-methylpiperazine was added, the reaction mixture was refluxed and stirred at 120°C for 20hr, cooled, and the pyridine was driven away, 10ml of chloroform and 5ml of ether were added to dissolve the residue, and the mixture Wash with water, dry, remove chloroform, and recrystallize the residue from ethanol to obtain levofloxacin (80 mg) with a yield of 80%. mp238-240°C. [α] _D ²⁰ = -77.9°C (c = 0.2, 0.05N NaOH). MSm/e361 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm 1.45 (3H, d, J=7Hz), 2.30 (3H, s), 2.50 (4H, m), 3.30 (4H, m), 4.35-4.58 (2H, dd, J=7Hz), 4.90 (1H, d, J=6Hz), 7.58 (1H, m, J=13Hz), 8.89 (1H, s). Experiment 13 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(4-aminomethylpiperidinyl)-7-oxo-7H-pyrido[1,2,3 -de][1,4]Benzoxazine-6-carboxylic acid 14112mg (S)-13 was dissolved in 3ml of pyridine, and 68.5mg of 4-aminomethylpiperidine was added with stirring. The reaction mixture was refluxed overnight at 120-130°C, pyridine was evaporated, the residue was dissolved in ethanol, the ethanol was evaporated under reduced pressure, acetone was added to the remaining residue, insoluble matter was filtered off, part of the acetone was evaporated, and crystallization was obtained to obtain 120 mg of the product. Yield 80%. mp175-178°C. MS m/e = 375 (M ⁺ ). ¹ H-NMR (DMSO-D ₆ ) δppm 1.05-1.55 (5H, m), 1.45 (3H, d, J=7Hz), 2.45 (2H, d), 2.95-3.30 (4H, m), 4.32-4.55 (2H, dd), 4.88 (H, d), 7.60 (1H, d,), 8.80 (1H, s). Experiment 14 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(piperazinyl)-7-oxo-7H-pyrido[1,2,3-de][1 , 4] Benzoxazine-6-carboxylic acid 15448 mg (S)-13 was dissolved in 10 ml of pyridine, and 208 mg of piperazine was added under stirring. The reaction mixture was refluxed overnight at 120-130° C., pyridine was evaporated, the residue was dissolved in ethanol, the ethanol was evaporated under reduced pressure, and the residue was recrystallized with ethanol to obtain 360 mg of the product with a yield of 65%. mp>250°C. MS m/e = 347 (M ⁺ ). ¹ H-NMR (DMSO-D ₆ ) δppm 1.48 (3H, d, J=7Hz), 2.50 (4H, m), 3.32 (4H, m), 4.38-4.60 (2H, dd), 4.92 (H, d , J=7Hz), 7.58 (1H, d,), 8.89 (1H, s). Experiment 15 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(3-methylpiperazinyl)-7-oxo-7H-pyrido[1,2,3- de][1,4]Benzoxazine-6-carboxylic acid 16112mg (S)-13 was dissolved in 3ml of pyridine, and 60mg of 2-methylpiperazine was added under stirring. The reaction operation is the same as experiment 14. The residue was recrystallized from acetone-water to obtain 57.2 mg of the product, with a yield of 40%. mp225-228°C. MS m/e = 361 (M ⁺ ). ¹ H-NMR (DMSO-D ₆ ) δppm 1.00 (3H, d, J = 6Hz), 1.48 (3H, d, J = 6Hz), 2.50 (3H, m), 3.30 (4H, m), 4.53- 4.60 (2H, dd), 4.92 (H, d, J=7Hz), 7.60 (1H, d,), 8.89 (1H, s). Experiment 16 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(4-(2-aminoethyl)piperazinyl)-7-oxo-7H-pyrido[1 , 2,3-de][1,4]benzoxazine-6-carboxylic acid 17112mg (S)-13 was dissolved in 3ml of pyridine, and 103mg of 1-(2-aminoethyl)piperazine was added under stirring. The reaction operation is the same as experiment 14. The residue was added with water to precipitate a white solid, which was filtered and recrystallized from water to obtain 80 mg of the product with a yield of 51%. mp 203-228°C. MS m/e = 390 (M ⁺ ). ¹ H-NMR (DMSO-D ₆ ) δppm 1.48 (3H, d, J=6Hz), 2.32-2.27 (8H, m), 3.35 (4H, m), 4.35-4.55 (2H, dd), 7.60 ( 1H, d, J = 13 Hz), 8.89 (1H, s). Experiment 17 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(4-(2-pyridylpiperazine)-7-oxo-7H-pyrido[1,2, 3-de][1,4]benzoxazine-6-carboxylic acid 18112mg (S)-13 was dissolved in 3ml pyridine, and 260mg1-(2-pyridyl)piperazine was added under stirring. The reaction operation was the same as in Experiment 14 18 was obtained _by recrystallization from ethanol with a yield of 41.3 ^{% .} 3.45(4H,m), 3.90(4H,m), 4.30～4.52(3H,m), 6.80(2H,m), 7.31(1H,d), 8.76(1H,m), 8.30(1H,d) , 8.65(1H, s). Experiment 18(S)-9-fluoro-2,3-dihydro-3-methyl-10-(4-(2-pyrimidinepiperazine)-7-oxo-7H- Pyrido[1,2,3-de][1,4]benzoxazine-6-carboxylic acid 19112mg (S)-13 was dissolved in 3ml of pyridine, and 264mg of 1-(2-pyrimidinyl)piperidine was added under stirring The reaction operation is the same as that in Experiment 14. Chloroform-acetone recrystallization yields 85 mg of the product with a yield of 50%. mp290-292°C. MS m/e=425 (M ⁺ ) _. (3H, d, J=6Hz), 3.25(4H), 3.90(4H, m), 4.40～4.62(2H, m,), 4.95(H, d), 6.71(1H, d), 7.61(1H, d, J=11Hz), 8.40 (2H, d), 9.00 (1H, s). Experiment 19 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(3,5- Dimethylpiperazinyl)-7-oxo-7H-pyrido[1,2,3-de][1,4]benzoxazine-6-carboxylic acid 2084.1mg(S)-13 dissolved in 3ml In pyridine, add 60mg3 under stirring, 5-dimethylpiperazine.Reaction operation is the same as experiment 14.Residue ethanol-acetone recrystallization, obtains product 30mg, yield 27%.mp240-243 ℃.MSm/e=375 (M ⁺ ) ^.1 H-NMR (CF ₃ COOD) δppm: 2.20 (6H, d), 2.50 (3H, d), 4.28 (3H, m), 4.58 (4H, m), 5.31 ~ 5.48 (2H, dd), 5.78(H,d), 8.72(1H,d), 9.95(1H,s). Experiment 20(S)-9-fluoro-2,3-dihydro-3-methyl-10-hopiper Azinyl-7-oxo-7H-pyrido[1,2,3-de][1,4]benzoxazine-6-carboxylic acid 21112mg(S)-13 was dissolved in 5ml pyridine, under stirring Add 60 mg of homopiperazine. The reaction operation is the same as experiment 14. DMF-ethanol recrystallized to obtain 82 mg of product, yield 57%. mp>280°C. MS: m/e = 347 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm1.70 (3H, d), 2.53 (2H, m), 3.60-.4.00 (8H, m), 4.50-4.72 (2H, dd), 5.50 (1H, d) , 7.95 (1H, d), 9.18 (1H, s). Experiment 21 (S)-9-fluoro-2,3-dihydro-3-methyl-8-nitro-7-oxo-7H-pyrido[1,2,3-de][1,4] Dissolve 22200 mg (S)-13 of benzoxazine-6-carboxylic acid in 4 ml of concentrated sulfuric acid, add 400 mg of KNO3 when the internal temperature is lower than 5°C, and react for 30 minutes after addition, and then react at room temperature for 1 hr. The reaction mixture was poured into ice water, and the precipitate was filtered, washed with water, dried, and recrystallized from chloroform-ethanol to obtain 0.21 mg of the product with a yield of 91%. Experiment 22 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-7-oxo-7H-pyrido[1,2,3-de][1,4]benzene Oxazine-6-carboxylic acid 230.22g22 and 180mg ethanol and 20mg Pd/C (10%) were hydrogenated at normal pressure for 3hr. After the reaction is complete, heat it to dissolve it, filter the catalyst while it is hot, and precipitate out a precipitate after cooling down slightly. After removing ethanol, 0.14 g of the product was obtained with a yield of 78%. mp>280°C. MS m/e = 296 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm 1.48 (3H, d, J=7Hz), 4.25-4.51 (2H, dd), 4.81 (1H, d), 8.98 (1H, s). Experiment 23 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-10-(4-methylpiperazinyl)-7-oxo-7H-pyrido[1, 2,3-de][1,4]benzoxazine-6-carboxylic acid 24100 mg 23 and 40 mg N-methylpiperazine were dissolved in 5 ml pyridine. The reaction operation is the same as experiment 14. Recrystallized from DMF-ethanol to obtain 70 mg of the product with a yield of 55%. mp>260°C decomposes. MS m/e 376 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm 1.61 (3H, d), 3.05 (3H, s), 3.28-3.78 (8H, m), 4.40-4.60 (2H, dd), 4.78 (1H, d), 8.98 (1H, s). Experiment 24 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-10-piperazinyl-7-oxo-7H-pyrido[1,2,3-de] [1,4] 25,100 mg of benzoxazine-6-carboxylic acid was reacted with 117 mg of anhydrous piperazine, and the reaction operation was the same as in experiment 14. 65 mg of product was obtained, yield 53%. mp215 ℃ decomposition. MS m/e = 362 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm 1.75 (3H, d), 3.60-4.00 (8H, m), 4.48-4.72 (2H, dd), 4.90 (1H, d), 8.45 (1H, s). Experiment 25 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-10-(3-methylpiperazinyl)-7-oxo-7H-pyrido[1, 2,3-de][1,4]Benzoxazine-6-carboxylic acid 26100mg23 was reacted with 136mg 2-methylpiperazine, and the reaction operation was the same as in Experiment 14. Recrystallization from water gave 50mg of the product with a yield of 39%. mp215 ℃ decomposition. MS m/e = 376 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm 1.85 (3H, d), 2.00 (3H, d), 3.80-4.20 (7H, m), 4.48-4.72 (2H, dd), 4.90 (1H, d), 8.45 (1H, s). Experiment 26 (S)-9-fluoro-2,3-dihydro-3-methyl-amino-10-(4-(2-aminoethylpiperazinyl)-7-oxo-7H-pyrido[ 1,2,3-de][1,4]benzoxazine-6-carboxylic acid 27100mg 23 reacted with 176mg1-(2-aminoethyl)piperazine, and the reaction operation was the same as in Experiment 14. Acidify to pH= with hydrochloric acid 1, standing, filtering, acetone-water recrystallization to obtain 40mg 27, yield 27%. mp236-237°C. MSm/e 405 (M ⁺ ). ¹ H-NMR (D ₂ O) δppm 1.56 (3H, d ), 3.50-4.00 (12H, m), 4.28-4.45 (2H, dd), 4.65 (1H, d), 8.50 (1H, s). Experiment 27 (S)-9-fluoro-2,3-dihydro -3-Methyl-8-amino-10-homopiperazinyl-7-oxo-7H-pyrido[1,2,3-de][1,4]benzoxazine-6-carboxylic acid 28120mg 23 and 85 mg of homopiperazine were reacted, operation experiment 14. DMF-ethanol recrystallization gave 100 mg of 28, the yield was 66%. mp>230°C decomposed. MS m/e 376 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm1.65 (3H, d), 2.30 (2H, m), 3.50-3.80 (8H, m), 4.38-4.62 (2H, dd), 4.80 (1H, d), 7.00 (1H, s). Experiment 28 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(4-ethoxycarbonylpiperazinyl)-7-oxo-7H-pyrido[1,2,3- de][1,4]Benzoxazine-6-carboxylic acid 29 and 15 equimolar benzoic acid were dissolved in 20ml of anhydrous tetrahydrofuran, cooled to -15°C, added 42μl of triethylamine, and then 14μl of ethyl chloroformate After reacting at -15°C for 1 hr, add 52 mg of 15, continue the reaction for 30 min., and detect the end point of the reaction by TLC. After the reaction, acidify to pH=3-4 with 1N hydrochloric acid, evaporate THE, and extract the residue with chloroform, dry, The solvent was removed, and the solid was recrystallized from ethyl acetate to obtain 30 mg of 29, with a yield of 48%. mp235-237°C. MSm/e 419 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 1.3 (3H, m, J =7Hz), 1.58(3H, d), 3.35(4H, m), 3.65(4H, m), 4.18(2H, m), 4.35-4.51(3H, d), 7.75(1H, d, J=13Hz ), 8.62 (1H, s). Experiment 29 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(3-methyl-4-ethoxycarbonylpiperazine)-7 -Oxo-7H-pyrido[1,2,3-de][1,4]benzoxazine-6-carboxylic acid 3040mg 16 was suspended in 15ml sodium dry tetrahydrofuran, added 0.032ml triethylamine, in ice-cooled A solution of 0.011 ml of ethyl chloroformate in 5 ml of THF was added dropwise, stirred for 2-3 hrs after the addition, and then reacted at room temperature for 2-3 hrs, and the end point of the reaction was detected by TLC. After the reaction, adjust to acidic (pH=3-4) with 1N hydrochloric acid, distill off THF, and leave the residue as a column layer (eluent: dichloromethane/methanol=25:1). 30 mg of 30 was obtained, yield 63%. mp252-253°C. MS m/e = 433 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 1.35 (6H, m,), 1.65 (3H, d), 3.15-3.55 (7H, m), 4.0 (1H, m), 4.18 (2H, m), 4.40 ( 2H, m), 7.70 (1H, d, J=12Hz), 8.62 (1H, s). Experiment 30 (S)-9-Fluoro-2,3-dihydro-3-methyl-10-(4-isobutoxycarbonylpiperazinyl)-7-oxo-7H-pyrido[1,2, 3-de][1,4]benzoxazine-6-carboxylic acid 3150mg 15 (0.144mmol) was dissolved in a saturated solution of sodium bicarbonate, cooled to 0°C, and 38μl of isobutyl chloroformate in 2ml of ethylene glycol was added dropwise Dimethyl ether solution, continue to react for 1 hr after adding, and react for 2 hr at 18-20°C. Acidify to pH=3-4, filter, dry the precipitate, and recrystallize from methanol to obtain 25 mg of 31, yield 39%. mp252-253°C. MS m/e = 447 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 0.95 (6H, 2×s,), 1.59 (3H, d, J=6Hz), 1.95 (1H, m), 3.35 (4H, m), 3.60 (4H, m ), 3.89 (2H, d, J = 2Hz), 4.40-4.51 (3H, m), 7.75 (1H, d, J = 12Hz), 8.65 (1H, s). Experiment 31 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(4-methyloxycarbonylpiperazine)-7-oxo-7H-pyrido[1,2,3 -de][1,4]Benzoxazine-6-carboxylic acid 3260mg (0.173mmol) 15 was suspended in 15ml of sodium dry tetrahydrofuran, added 0.048ml of triethylamine, and 0.027ml (0.35mmol) of chlorine was added dropwise under ice-cooling Methyl formate in 5 ml THF. The operation is the same as experiment 29. Column chromatography of the residue, eluent: dichloromethane/methanol=20:1. 20 mg of 32 was obtained, yield 29%. mp237-239°C. MS m/e 405 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 1.65 (3H, d, J=6Hz), 3.30 (4H, m), 3.60 (4H, m), 3.70 (3H, s), 4.40-4.60 (3H, m) , 7.70 (1H, d, J = 12Hz), 8.65 (1H, s). Experiment 32 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-10-(4-isobutoxycarbonylpiperazine)-7-oxo-7H-pyrido[ 1,2,3-de][1,4]benzoxazine-6-carboxylic acid 3350mg 25 (0,138mmol) reacted with equimolar isobutyl chloroformate, the operation was the same as in experiment 30, column chromatography (elution Agent: hexyl acetate/petroleum ether=1.5:1). 25 mg of the product was obtained, and the yield was 39%. mp260-262°C. MS m/e 462 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 0.95 (6H, 2×s,), 1.55 (3H, d, J=6Hz), 1.98 (1H, m), 3.32 (4H, m), 3.65 (4H, m ), 3.90 (2H, d, J=2Hz), 4.18-4.35 (3H, m), 8.45 (1H, s). Experiment 33 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-10-(4-ethoxycarbonylpiperazine)-7-oxo-7H-pyrido[1 , 2,3-de][1,4]benzoxazine-6-carboxylic acid 3450mg 25 (0,138mmol) was reacted with 0.026ml ethyl chloroformate, the operation was the same as experiment 30. Recrystallized from ethanol to obtain 35mg of the product with a yield of 61%. mp281-282°C. MS m/e = 434 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 1.36 (3H, m, J = 8Hz), 1.60 (3H, d, J = 6Hz), 3.30 (4H, m), 3.50 (4H, m), 4.20 (2H, m), 4.35-4.50 (3H, m), 8.45 (1H, s). Experiment 34 (S)-9-Fluoro-2,3-dihydro-3-methyl-8-amino-10-(3-methyl-4-ethoxycarbonylpiperazine)-7-oxo-7H- Pyrido[1,2,3-de][1,4]benzoxazine-6-carboxylic acid 3550mg 26 (0.138mmol) was reacted with 0.026ml ethyl chloroformate, the operation was the same as experiment 30. Recrystallized from ethanol to obtain 30 mg. Yield 50%. mp 240-242°C. MS m/e 448 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 1.30 (3H, m), 1.48 (3H, d), 3.15-3.45 (7H, m), 3.95 (2H, m), 4.05-4.40 (3H, m), 8.40 (1H, s). Experiment 35 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-10-(3-methyl-4-isobutyloxycarbonylpiperazine)-7-oxo- 7H-pyrido[1,2,3-de][1,4]benzoxazine-6-carboxylic acid 3650mg 26 (0,133mmol) reacted with 0.038ml isobutyl chloroformate, the operation was the same as experiment 30, column layer Analysis (eluent: dichloromethane/methanol = 30:1). 43 mg of the product was obtained, and the yield was 68%. mp224-226°C. MS m/e 476 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 0.95 (6H, 2×s,), 1.35 (3H, m), 1.95 (1H, m), 3.15-3.50 (7H, m), 3.95 (2H, m), 4.40 (3H, m), 8.50 (1H, s). Experiment 36 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(3-methyl-4-isobutyloxycarbonylpiperazine)-7-oxo-7H-pyrido [1,2,3-de][1,4]Benzoxazine-6-carboxylic acid 3750mg26 (0,138mmol) was reacted with 0.036ml isobutyl chloroformate, and the operation was the same as in experiment 30. 40 mg of 37 was obtained by recrystallization from ethanol, with a yield of 63%. mp205-206°C. MS m/e 461 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm0.98 (6H, 2×s), 1.45 (3H, m), 1.53 (3H, d), 1.95 (1H, m), 3.15-3.50 (7H, m), 3.90 (2H, m), 4.35-4.55 (3H, m), 7.65 (1H, d, 12), 8.60 (1H, s). Experiment 37 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-10-(4-methoxycarbonylpiperazine)-7-oxo-7H-pyrido[1 , 2,3-de][1,4]benzoxazine-6-carboxylic acid 3850mg 25 (0,138mmol) reacted with 0.021ml (0.27mmol) methyl chloroformate and 0.038ml triethylamine, and the operation was the same as in Experiment 29 . Eluent: dichloromethane/methanol=20:1. 15 mg of 38 was obtained, yield 26%. mp286-288°C. MS m/e 420 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm1.45 (3H, m), 3.28 (4H, m), 3.50 (4H, m), 3.75 (3H, s), 4.15-4.40 (3H, m), 8.45 (1H , s). Experiment 38 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(3-methyl 4-methoxycarbonylpiperazine)-7-oxo-7H-pyrido[1 , 2,3-de][1,4]Benzoxazine-6-carboxylic acid 3950mg 16 (0.138mmol) reacted with 0.021ml (0.27mmol) methyl chloroformate and 0.038ml triethylamine, the operation was the same as in Experiment 29 . Developing solvent: dichloromethane/methanol=20:1. 20 mg of product was obtained, yield 35%. mp195-198°C. MS m/e 419 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 1.30 (3H, m), 1.68 (3H, d), 3.10-3.45 (7H, m), 3.65 (3H, m), 4.30-4.55 (3H, m), 7.70 (1H, d, J = 12Hz), 8.65 (1H, s). Experiment 39 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-10-(3-methyl-4-methoxycarbonylpiperazine)-7-oxo-7H -Pyrido[1,2,3-de][1,4]benzoxazine-6-carboxylic acid 4050mg 26 (0,133mmol) reacted with 0.021ml (0.27mmol) methyl chloroformate and 0.038ml triethylamine , the operation is the same as Experiment 29. Eluent: dichloromethane/methanol=20:1. 25 mg of the product was obtained, and the yield was 43%. mp>280°C decomposes. MS m/e 434 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm1.28 (3H, m), 1.55 (3H, d), 3.10-3.50 (7H, m), 3.75 (3H, m), 4.10-4.45 (3H, m), 8.49 (1H, s). Experiment 40 (S)-9-Fluoro-2,3-dihydro-3-methyl-10-(4-trichloroethyloxycarbonylpiperazine)-7-oxo-7H-pyrido[1, 2,3-de][1,4]Benzoxazine-6-carboxylic acid 4150mg 15 (0.144mmol) reacts with 0.020ml (0.15mmol) trichloroethyl chloroformate and 0.038ml triethylamine, the operation is the same as the experiment 29. Developing solvent: dichloromethane/methanol=30:1. 37mg of 41 was obtained, yield 49%. mp238-240°C decomposition. MS m/e 521 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 1.51 (3H, d), 3.35 (4H, m), 3.60 (4H, m), 4.30-4.55 (3H, m), 4.95 (2H, s), 7.72 (1H , d), 8.65 (1H, s). Experiment 41 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-10-(4-trichloroethoxycarbonylpiperazine)-7-oxo-7H-pyrido [1,2,3-de][1,4]benzoxazine-6-carboxylic acid 4250mg 25 (0.138mmol) and 0.020ml (0.15mmol) trichloroethyl chloroformate and 0.020ml triethylamine reaction, The operation is the same as experiment 29. Eluent: dichloromethane/methanol=30:1. 23 mg of 42 was obtained, yield 31%. mp>260°C decomposes. MS m/e 536 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) 5ppm1.55 (3H, d, J=6Hz), 3.35 (4H, m), 3.70 (4H, m), 4.15-4.20 (2H, dd), 4.40 (1H, d) , 4.80 (2H, s), 8.45 (1H, s). Experiment 42 (S)-9-Fluoro-2,3-dihydro-3-methyl-10-(4-ethoxycarbonylhomopiperazine)-7-oxo-7H-pyrido[1,2, 4350mg21 of 3-de][1,4]benzoxazine-6-carboxylic acid was reacted with 0.025ml ethyl chloroformate, and the operation was the same as experiment 30. Eluent: dichloromethane/methanol=10:1. 40mg43 was obtained, yield 67%. mp203-205 ℃ decomposition. MS m/e 433 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 1.28 (3H, m), 1.65 (3H, d), 2.10 (1H, m), 3.45 (4H, m), 3.65 (4H, m), 4.25 (2H, m) , 4.30-4.45 (2H, dd), 4.52 (1H, d), 7.70 (1H, d), 8.65 (1H, s). Experiment 43 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(4-methoxycarbonylhomopiperazine)-7-oxo-7H-pyrido[1,2, 3-de][1,4]Benzoxazine-6-carboxylic acid 4450mg21 (0.138mmol) was heated and dissolved with 4ml DMF, after cooling, 0.048ml triethylamine was added, and then 0.022ml methyl chloroformate was added with a syringe, React under cooling for 30 min., and then react at room temperature for 3 hr. DMF was removed under reduced pressure, and the residue was dissolved in dichloromethane. Column chromatography (dichloromethane-methanol=30:1) gave 40 mg of 44, yield 77%. mp210-211°C decomposition. MS m/e 419 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm1.65 (3H, d), 1.95 (2H, d), 3.25-3.65 (8H, m), 3.70 (3H, s), 4.30-4.45 (3H, m), 7.65 (1H,d), 8.65(1H,s). Experiment 44 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(4-trichloroethoxycarbonylhomopiperazine)-7-oxo-7H-pyrido[1, 2,3-de][1,4]Benzoxazine-6-carboxylic acid 4550mg 21 (0.138mmol) was heated and dissolved with 4ml DMF, after cooling, 0.048ml triethylamine was added, and 0.022ml chloroformic acid tris Ethyl chloride, reacted under cooling for 30min. Then react at room temperature for 3 hrs, drain off DMF with an oil pump, and dissolve the residue with dichloromethane. Column chromatography (dichloromethane-methanol=20; 1). 20 mg of product was obtained, yield 27%. mp215-217°C. MS m/e 535 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm1.65 (3H, d), 2.05 (2H, d), 3.45 (4H, m), 3.75 (4H, m), 4.30-4.55 (3H, m), 4.78 (2H , s), 7.65 (1H, d), 8.61 (1H, s). Experiment 45 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-10-(4-ethoxycarbonylhomopiperazine)-7-oxo-7H-pyrido[ 1,2,3-de][1,4]Benzoxazine-6-carboxylic acid 4640mg28 (0.106mmol), 0.036ml triethylamine, 0.019ml ethyl chloroformate react, the operation is the same as experiment 30. Column chromatography (eluent: chloroform-methanol = 30:1), and then ethanol recrystallization gave 25 mg of product ² with a yield of 53%. mp240-241°C. MS m/e 448 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 1.25 (3H, d), 1.62 (3H, d), 2.05 (2H, d), 3.45 (4H, m), 3.65 (4H, m), 4.20 (2H, m) , 4.30(1H, m), 8.35(1H, s). Experiment 46 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-10-(4-trichloroethoxycarbonylhomopiperazine)-7-oxo-7H-pyridine And[1,2,3-de][1,4]benzoxazine-6-carboxylic acid 4740mg 28 (0.106mmol), 0.020ml triethylamine, 0.011mmol triethyl chloroformate react, the operation is the same as experiment 30 . Column chromatography (eluent, chloroform-methanol=30:1), and then ethanol-chloroform recrystallization gave 35 mg of the product. Yield 60%. mp>280°C decomposes. MS m/e 551 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 1.55 (3H, d, J=6Hz), 2.00 (1H, m), 3.50 (4H, m), 3.70 (4H, m), 4.15-4.30 (2H, dd) , 4.35 (1H, d), 4.80 (2H, s), 8.45 (1H, s). Experiment 47 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(4-benzyloxycarbonylpiperazinyl)-7-oxo-7H-pyrido[1,2,3 -de][1,4]Benzoxazine-6-carboxylic acid 4840mg 15 (0.115mmol) was reacted with 0.024ml benzyl chloroformate (0.12mmol) and 0.035ml triethylamine, and the operation was the same as Experiment 29. Column chromatography (eluent: dichloromethane/methanol=30:1). 46 mg of product was obtained, yield 83%. mp218-220°C. MS m/e 481 (M ⁺ ). ¹ H-NMR (CDCl3) δppm 1.65 (3H, d), 3.30 (4H, m), 3.65 (4H, m), 4.13-4.30 (2H, dd), 4.45 (1H, d), 5.15 (2H, s ), 7.27-7.35 (5H, m), 7.75 (1H, d, J=11Hz), 8.60 (1H, s). Experiment 48 (S)-(-)-9-fluoro-2,3-dihydro-3-methyl-10-(4-p-aminobenzenesulfonyl-1-piperazinyl)-7-oxo-7H -Pyrido[1,2,3-de][1,4]benzoxazine-6-carboxylic acid 4950mg 15 (0.144mmol) was dissolved in 4ml of 0.1N sodium hydroxide solution, added para After adding 35 mg (0.15 mmol) of nitrobenzenesulfonyl chloride, continue to react for 30 minutes, then react at room temperature for 3-4 hours, acidify with 1N hydrochloric acid to pH = 3-4, cool and filter, wash the precipitate with water, and dry. Recrystallized from DMF-ethanol to obtain 50 mg of light yellow crystals with a yield of 65%. 40 mg of the above crystals were dissolved by heating with methanol, and 20 mg of Pd/C (10%) was catalytically hydrogenated for 8 hours. The catalyst was filtered off, the methanol was evaporated, and the residue was recrystallized with ethanol to obtain 16 mg of the product with a yield of 42%. mp>260°C decomposes. MS m/e 502 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm 1.65 (3H, d, J=7Hz), 3.38 (4H, m), 3.75 (4H, m), 4.45-4.62 (2H, dd), 4.95 (1H, d) , 7.80 (1H, d, J=9Hz), 7.85-8.00 (4H, m), 9.15 (1H, s). Experiment 49 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-10-(4-p-aminophenylsulfonylpiperazine)-7-oxo-7H-pyrido [1,2,3-de][1,4]Benzoxazinazine-6-carboxylic acid 5050 mg 25 and 35 mg p-nitrobenzenesulfonyl chloride The operation was the same as in Experiment 48. 16mg of the product was obtained, the yield was 22% (combination of two steps). mp>280°C decomposes. MS m/e = 517 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm1.55 (3H, d, J=6Hz), 3.30 (4H, m), 3.60 (4H, m), 4.45-4.62 (2H, dd), 4.75 (1H, d ), 7.75-8.00 (4H, dd), 8.95 (1H, s). Experiment 50 (S)-(-)-9-fluoro-2,3-dihydro-3-methyl 10-(4-methylsulfonyl-1-piperazinyl)-7-oxo-7H-pyridine And [1,2,3-de][1,4]benzoxazine-6-carboxylic acid 5150mg 15 (0.144mmol) and MsCl (0.15mmol) reaction, operation experiment 48. The product was recrystallized from chloroform-methanol to obtain 31 mg of the product, and the yield was 50%. mp>250°C decomposes. MS m/e 425 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm 1.75 (3H, d), 3.00 (3H, s), 3.51-4.10 (8H, m), 4.59-4.75 (2H, m), 5.10 (1H, d), 7.95 (1H, d), 9.25 (1H, s). Experiment 51 (S)-(-)-9-fluoro-2,3-dihydro-3-methyl-10-(4-benzenesulfonyl-1-piperazinyl)-7-oxo-7H-pyridine And [1,2,3-de][1,4]benzoxazine-6-carboxylic acid 5250mg 15 (0.144mmol) reacted with benzenesulfonyl chloride (0.15mmol), operation experiment 48. Yield 55%. mp>250°C decomposition; MSm/e487(M ⁺ ); ¹ H-NMR(CF ₃ COOD) δppm1.68(3H, d), 3.51-4.10(8H, m), 4.50-4.25(2H, m), 5.10 (1H, d), 7.55-8.00 (5H, m), 9.15 (1H, s). Experiment 52 (S)-(-)-9-fluoro-2,3-dihydro-3-methyl-10-(4-p-toluenesulfonyl-1-piperazinyl)-7-oxo-7H- Pyrido[1,2,3-de][1,4]benzoxazine-6-carboxylic acid 5350mg 15 (0.144mmol) reacted with p-toluenesulfonyl chloride (0.15mmol), operation experiment 48, yield 70% . mp>260°C decomposes. MSm/e501 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm 1.65 (3H, d), 2.39 (3H, s), 3.45 (4H, m), 4.00 (4H, m), 4.51-4.72 (2H, m), 5.10 ( 1H, d), 7.74-7.90 (4H, 3×d), 9.25 (1H, s). Experiment 53 (S)-(-)-9-fluoro-2,3-dihydro-3-methyl-10-(4-p-acetamidobenzenesulfonyl-1-piperazinyl)-7-oxo -7H-pyrido[1,2,3-de][1,4]benzoxazine-6-carboxylic acid 5450mg 15 (0.144mmol) and p-acetamidosulfonyl chloride (0.15mmol) reaction, the operation is the same as above, Yield 64%. mp205-207 ℃ decomposition. MSm/e544 (M ⁺ ). H-NMR (CF ₃ COOD) δppm 1.45 (3H, d), 2.10 (3H, s), 3.00 (4H, m), 3.30 (4H, m), 4.30～4.55 (2H, dd), 4.90 (1H, m), 7.58-7.75 (4H, 2×d,), 7.85 (1H, d), 8.95 (1H, s). Experiment 54 (S)-9-Fluoro-2,3-dihydro-3-methyl-10-(4-(2-bromoethylcarbonyl)piperazinyl)-7-oxo-7H-pyrido[1 , 2,3-de][1,4]benzoxazine-6-carboxylic acid 5550mg 15 (0.144mmol) was dissolved in saturated NaHCO ₃ solution, cooled to 0°C, and 2ml of 30mgβ-bromopropionic acid chloride was added dropwise After the addition of ethylene glycol dimethyl ether solution, continue to react for 1 hr, and react at 18-20 °C for 3 hr. After the reaction is completed, adjust the pH to 3-4 with 1N hydrochloric acid, filter, dry the precipitate, and recrystallize with methanol to obtain 35mg 55, yield 50%. mp221-223°C. MS m/e 482 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm2.55 (3H, d, J=7Hz), 4.10 (2H, m), 4.50 (2H, m), 4.50-5.00 (8H, m), 5.45-5.55 (2H , dd), 5.58 (1H, dd), 8.80 (1H, d), 10.00 (1H, s). Experiment 55 (S)-9-Fluoro-2,3-dihydro-3-methyl-10-(4-chloromethylcarbonylpiperazinyl)-7-oxo-7H-pyrido[1,2, 3-de][1,4]Benzoxazine-6-carboxylic acid 5650mg 15 was dissolved in 3ml dry DMF, added 0.020ml triethylamine, cooled to 0°C, added dropwise 0.012ml of chloroacetyl chloride in 2ml of ethylene glycol After the alcohol dimethyl ether solution was added, the reaction was continued for 3 hours, triethylamine hydrochloride was filtered off, DMF was evaporated to dryness, and 50 mg of the product was obtained by ethanol recrystallization, with a yield of 82%. mp202-204. MSm/e423 (M ⁺ ). ¹ H-NMR (CF ₃ COOD) δppm 1.48(3H,d), 2.48(4H,m), 3.25(4H,m), 3.60(2H,s), 4.40-4.60(2H,dd), 4.95(1H , dd), 7.65 (1H, d), 9.00 (1H, s). Experiment 56 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(3-methyl-4-chloromethylcarbonylpiperazine)-7-oxo-7H-pyrido[ 1,2,3-de][1,4]Benzoxazine-6-carboxylic acid 5750mg 16 was dissolved in 20ml of dry THF, added 0.020ml of triethylamine, cooled to 0°C, added dropwise with 0.012ml of chloroacetyl chloride After the addition of 2ml of ethanol dimethyl ether solution, the reaction was continued for 3 hrs, triethylamine hydrochloride was filtered off, steamed in THF, and recrystallized from ethanol to obtain 30 mg of the product with a yield of 50%. mp164-165°C. MS: m/e437 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm1.60 (3H, m), 1.65 (3H, m), 3.20 (4H, m), 3.40 (4H, m), 4.10 (2H, s), 4.35-4.55 (3H , m), 7.75 (1H, d, J=12Hz), 8.65 (1H, s). Experiment 57 (S)-9-fluoro-2,3-dihydro-3-methyl-8-amino-10-(3-methyl-4-chloromethylcarbonylpiperazine)-7-oxo-7H -Pyrido[1,2,3-de][1,4]benzoxazine-6-carboxylic acid 5850mg 26 was dissolved in 3ml dry DMF, added 0.020ml triethylamine, cooled to 0°C, added chlorine dropwise After adding 0.012ml of acetyl chloride in 2ml of ethanol dimethyl ether, the reaction was continued for 3hr. The operation is the same as experiment 56. Column chromatography (eluent: chloroform/methanol=30:1). 42 mg of the product was obtained, and the yield was 34%. mp269-270°C. MS m/e 452 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 1.35 (3H, d), 1.55 (3H, d), 3.20-3.60 (7H, m), 4.10 (2H, s), 4.15-4.40 (3H, m), 8.45 ( 1H, s). Experiment 58 (S)-9-Fluoro-2,3-dihydro-3-methyl-8-amino-10-(4-chloromethylcarbonylpiperazine)-7-oxo-7H-pyrido[1 , 2,3-de][1,4]benzoxazine-6-carboxylic acid 5950mg 25 was dissolved in 3ml of dry DMF, added 0.020ml of triethylamine, cooled to 0°C, added dropwise 0.012ml of chloroacetyl chloride After adding 2ml of ethanol dimethyl ether solution, the reaction was continued for 3hr. The operation is the same as experiment 56. Recrystallized from ethanol-water to obtain 30 mg of the product with a yield of 50%. mp>280°C decomposes. MSm/e438 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 1.48 (3H, d, J=7Hz), 3.45 (4H, m), 3.80 (4H, m), 4.20 (2H, s), 4.21-4.48 (3H, m) , 8.48 (1H, s). Experiment 59 (S)-9-fluoro-2,3-dihydro-3-methyl-10-(4-chloromethylhomopiperazine)-7-oxo-7H-pyrido[1,2,3 -de][1,4]benzoxazine-6-carboxylic acid 6040mg 21 (0.11mmol), 0.020ml triethylamine, 0.010ml chloroacetyl chloride. The operation is the same as experiment 56. Column chromatography (eluent: dichloromethane/methanol=20:1). 21 mg of product was obtained, yield 44%. mp217-217°C. MSm/e437 (M ⁺ ). ¹ H-NMR (CDCl ₃ ) δppm 1.62 (3H, d), 2.00 (1H, m), 3.45 (4H, m), 3.70 (4H, m), 4.10 (2H, s), 4.30-4.55 (3H , m), 7.55 (1H, d), 8.65 (1H, s).

Claims (6)

1. A levofloxacin derivative with the following structural formula where _R7 is

R ₁ =H, C ₁ -C ₅ alkyl; m=1,2 R ₂ = H, C ₁ -C ₅ alkyl R ₃ ＝C ₁ -C ₅ straight chain or branched chain alkyl, CH ₂ Ph R ₄ =H, C ₁ -C ₅ alkyl, pyridine, pyrimidine, CO(CH ₂ )nX X=halogen, n=1,2 R ₅ =H, NH ₂ R ₆ = alkyl or substituted phenyl 2. according to the described levofloxacin of claim 1 and derivative thereof, it is characterized in that they are prepared by following intermediate (S)-5, (R)-6, (S)-7, (R)-7 get: 3. according to claim 1 described levofloxacin and derivatives thereof, it is characterized in that its preparation method: a trifluoronitrobenzene is condensed with (S)-4 under phase transfer catalysis conditions to obtain condensate (S)-5; b(S)-5 is deprotected to obtain (R)-6, and (R)-6 is bromoacetylated with acetic acid-hydrogen bromide to obtain (R)-7 and (S)-7; c. Cyclization of (R)-7 and (S)-7 in sodium methoxide methanol solution to obtain cyclized product (R)-8; d(R)-8 was reduced with palladium/carbon, and the reduced product was condensed with EMME to obtain the condensate (R)-10, which was cyclized and acid hydrolyzed to obtain the key intermediate (S)-13; e. Condensation of (S)-13 and HR ₇ in an appropriate anhydrous solvent to obtain the above-mentioned levofloxacin and its derivatives substituted at the 7-position; f. (S)-13 introduces ammonia at the 5-position to obtain 23, and condenses 23 and HR ₇ in a suitable solvent to obtain the above-mentioned levofloxacin derivative substituted with 5-amino group. g. Condensation of alkyl carbamate, alkyl or substituted phenylsulfonyl chloride, α-haloacetyl chloride and levofloxacin derivative with secondary nitrogen structure at the 7-position to obtain compound 29-60. 4. according to the described levofloxacin derivative of claim 1, it is characterized in that they have the effect of resisting Gram-negative and positive bacteria, and can be used as antibacterial drugs. 5 according to claim 1 said levofloxacin derivatives, characterized in that they have anti-tumor activity, can be used as anti-anti-tumor drug application. 6. according to the described levofloxacin derivative of claim 1, it is characterized in that they have anti-mycoplasma activity, can be used as anti-mycoplasma drug application.