Vol. 47 No. 1/2000 87–94 QUARTERLY Synthesis and antimicrobial activity of new adamantane derivatives

A series of fourteen derivatives of adamantane was synthesised. The new compound 4-(adamant-1-ylmethoxycarbonyl)phthalanhydride obtained from 1-adamantane-methanol and trimellitic anhydride chloride appeared very useful for preparation of a number of N-substituted phthalimides. Antimicrobial activity of the newly obtained derivatives such as, for example, 4-(adamant-1-ylmethoxycarbonyl)-N-(5-carboxypentamethylene)p hthalimide or 4-(adamant-1-ylmethoxycarbonyl)-N-(L-alanyl)phthalimide was tested against Staphylococcus aureus, Bacillus sp., Micrococcus flavus and Enterococcus faecium. The minimal inhibitory concentration (MIC) for these compounds against S. aureus were 0.022 and 0.05 microg/ml, respectively.

Trimellitic acid anhydride (4-carboxyphthalanhydride) (TMAA) is used in many fields of organic chemistry.Some of ester-imides derived from this anhydride, e.g.compounds with an amino acid as N-substituent and a cholesteryl, phenyl or biphenyl moiety-containing ester group, exhibit liquid-crystal properties [1][2][3].Trimellitic anhydride is also fre-quently used as substrate for poly(ester imide)s synthesis.The combination of the rigid and flat phthalimide core with mesogenic properties of the ester structure endows such polymers with a liquid crystalline character [4][5][6][7][8].Many compounds possessing imide rings in their structure exhibit biological activity.For example, some imides are valuable substrates for the production of anti-seizure drugs [9].The most known of the clinically useful imides is the antiviral drugamantidine (1-aminoadamantane) [10].Another field where amantidine and many related derivatives are successfully employed, is the treatment of certain neurological disorders, e.g.Parkinson's disease [11].There are only few papers concerning the antimicrobial activity of adamantane derivatives [12][13][14].
In this study we show that certain easily synthesise 1-adamantanemethanol esters of various N-substituted phthalimide 4-carboxylates exhibit a distinct antimicrobial activity.It is worth to note that among structurally similar compounds N-(1-adamantyl)maleimide shows anticancer activity in mice and inhibits herpes simplex virus replication in vitro [14,15].N-Adamantylphthalimide induces tumor necrosis factor (TNF-a) enhancing activity induced by 12-O-tetradecanoylphorbol-13-acetate in human leukaemia HL-60 cells [16].We expect that a combination of both types of compounds, those with a flat phthalimide core and those with a three-dimensional "box-like" adamantane structure, will provide a broad range of active compounds for biological and pharmacological investigations.

General methods
Melting points were taken in open capillary tubes on a Gallenkamp 5 melting point apparatus and were uncorrected.The structures of products were confirmed by elemental analysis, FTIR and 1 H NMR spectroscopy.The NMR spectra were measured on a Varian Gemini 200 MHz spectrometer in CDCl 3 or d 6 -Me 2 SO solutions.Column flash chromatography was performed on silica gel 60 (Merck).FTIR spectra were recorded on a Perkin Elmer 2000 apparatus using the KBr pellet method.Adamantane and phthalic acid derivatives were purchased from Aldrich.The spe-cific rotation of compounds was determinated on a Perkin Elmer polarimeter at 20°C in chloroform (c =1 g/100 cm 3 ) using the D line of sodium.Preliminary testing of the antimicrobial activity of the newly synthesised compounds was performed by the disc diffusion method using Mueller-Hinton agar medium under standard conditions as described by NCCLS [17].Sterile filter paper discs were soaked in test compounds solutions prepared in EtOH-Me 2 SO mixture (v/v, 1 : 1).The results were read following 18 h incubation at 37°C (for M. flavus at 30°C).Compounds showing distinct antimicrobial activity in the above test were next tested for MIC (minimal inhibitory concentration) in liquid Mueller-Hinton medium according to the appropriate NCCLS protocol, using original stock solutions [18].Staphylococcus aureus NCTC 4163 and Enterococcus faecium ATCC 6057 were purchased from the National Institute of Hygiene (Warsaw, Poland) Bacillus subtilis H17rec - and Bacillus subtilis M45rec + were kindly donated by dr.T. Kada from the National Institute of Genetics (Misima, Japan) the other microorganisms used were from own collection of the Department of Pharmaceutical Microbiology, Medical University (Warsaw, Poland).

Microbiological studies
The antimicrobial activity of adamantyl derivatives of phthalimide was first tested by the agar disc-diffusion method against Gram-positive bacteria: Staphylococus aureus, Micrococcus flavus, Enterococcus faecium and certain strains of Bacillus.Gram-negative bacteria: Bordella bronchiseptica, Pseudomonas aeruginosa and strains belong to the family Enterobacteriaceae as well as the fungus Candida albicans were resistant to all tested compounds.Next, the minimal inhibition concentration (MIC) of the most active compounds was determined in liquid Mueller-Hinton medium (Table 1).A particularly strong antimicrobial activity, comparable to that of clinically used antibiotics, was observed for N-amino-acid substituted derivatives 10 and 11.Both chiral isomers, i.e. 10L, 10D, and 11L, 11D were highly active   ity of both chiral isomers 11L and 11D were observed for S. aureus ATCC 6538P and M. flavus strains.
Noteworthy are large differences in the antimicrobial activity obtained by the disc-diffusion method and MIC method with Bacillus  strains (Tables 2a and 2b).A similar discrepancy was previously observed for these aerobic bacteria in antimicrobial tests of essential oil from Tanacetum partenium [21].The high sensitivity of Bacillus strains may be useful in testing the antimicrobial activity of next adamantane derivatives, the synthesis of which we plan in the nearest future.
The genetically modified Bacillus rec + and rec -strains employed in the present study were previously shown to be useful in screening the genotoxicity of antibiotics under development [22].Results obtained in the course of the present study (details not shown) showed no genotoxicity of the newly synthesised adamantane antimicrobials.We plan to expand these studies to include other N-aminoacyl-substituted phthalimidecarboxylic acid esters of adamantane.

90 A. Orzeszko and others 2000 against
Micrococcus flavus and Staphylococcus strains.In contrast, 11L and 11D were practically inactive against Enterococcus faecium, whereas 10L and 10D showed moderate activity against this microorganism.The most distinct differences in the antimicrobial activ-Vol.47 New adamantane derivatives 91 Scheme 1.

Table 1 . Sensitivity of Gram-positive cocci strains to certain adamantyl derivatives of substituted phthalimides
*na -no activity at concentration > 50 mg/ml; **compounds 4 and 5 were inactive against bacteria strains tested.