1. Łukasz Sienicki Institute of Chemistry, University of Białystok, Piłsudskiego 11/4, 15-443 Białystok, Poland University of Bialystok

2. Fig.1. Synthesis of new 1,4-disubstituted piperazines The compounds were resolved into enantiomers on:  cellulose tris(4-methylbenzoate)  amylose tris(3,5-dimethylphenylcarbamate) -C 3 H 7 -C 4 H 9 -C 5 H 11 -C 6 H 13 -C 7 H 15 -C 5 H 11 R (1)(2)(3)(4)(5)(6)(7)Compound XHHHHHCH 3 -C 3 H 7 -C 4 H 9 -C 5 H 11 -C 6 H 13 -C 7 H 15 -C 5 H 11 (1)(2)(3)(4)(5)(6)(7) HHHHHCH 3

3. Fig. 3. Synthesis of 2-(3-methylpiperazin-1-yl)pyrimidine -HCl Cl H H -HCl 18-crown-6 reflux 3h Fig. 2. Synthesis of 2-piperazin-1-ylpyrimidine +

4. Rys. 4. Synthesis of dimethyl n-alkylmalonate RC3H7C3H7 C4H9C4H9 C 5 H 11 C 6 H 13 C 7 H 15 +

5. Fig. 5. Synthesis of ethyl 2-[(4-pyrimidin-2-ylpiperazin-1-yl)carbonyl]n-alkyloate + R Compound X -C 3 H 7 -C 4 H 9 -C 5 H 11 -C 6 H 13 -C 7 H 15 -C 5 H 11 (1)(2)(3)(4)(5)(6)(7) HHHHHCH 3

6.  HPLC analyses were performed using a Shimadzu liquid chromatograph consisting of a LC- 10AS pump, variable wavelenght UV SPD-10A detector and Valco valve injector equipped with 20 µl loop.  The mobile phases were the mixtures of heksane/propan-2-ol [80:20; 90:10; 95:5; 99:1 (v/v)]. All chromatograms were recorded at room temperature with a flow rate of 1 mL min -1 [100:0 (v/v); 0,22 mL min -1 ], λ = 242 nm.

7. Tab.1. Carbohydrate chiral stationary phases The separation of compounds 1-7 into enantiomers was tested on two polysaccharide columns:  Daicel Chiralcel OJ (10 , 4,6 mm x 250 mm) column packed with immobilised on silica cellulose tris(4-methylbenzoate)  Daicel Chiralpak AD (10 , 4,6 mm x 250 mm) column packed with immobilised on silica amylose tris(3,5-dimethylphenylcarbamate) were used. NAME TYPE OF ABSORBEN ELUENT CHIRALCEL OJ cellulose tris(4-methylbenzoate) heksane/isopropanol CHIRALPAK AD amylose tris(3,5-dimethylphenylcarbamate) heksane/isopropanol R R amylose tris(3,5-dimethylphenylcarbamate) CHIRALPAK AD heksane/isopropanol cellulose tris(4-methylbenzoate) CHIRALCEL OJ ELUENT TYPE OF ABSORBENT NAME R R heksane/isopropanol amylose tris(3,5-dimethylphenylcarbamate) CHIRALPAK AD heksane/isopropanol cellulose tris(4-methylbenzoate) CHIRALCEL OJ ELUENT TYPE OF ABSORBENTU NAME R R

8. Tab. 2. Retention factors and selectivity for compounds 1-7 on cellulose tris(4-methylbenzoate) (entries 1-15) and amylose tris(3,5-dimethylphenylcarbamate) (entries 16-18) with hexane-2-propanol (v/v) mobile phases. Flow rate 1 mL min -1, λ = 242 nm. Mobile phase 0:10080:2090:1095:599:195:5 Entry123456789101112131415161718 k1k1 k2k2 α k1k1 k2k2 α k1k1 k2k2 α k1k1 k2k2 α k1k1 k2k2 α k1k1 k2k2 α 1 0,50 1,00 1,40 1,00 3,82 1,00 7,758,22 1,06 23,925,17 1,05 6,396,93 1,08 2 0,40 1,00 1,13 1,00 2,662,85 1,07 5,646,26 1,11 20,5 1,00 6,396,83 1,07 3 0,39 1,00 0,82 1,00 2,35 1,00 4,995,53 1,11 23,12 1,00 6,226,62 1,06 4 0,39 1,00 0,80,80 1,00 2,15 1,00 4,655,2 1,11 21,6626,35 1,22 6,32 1,00 5 0,330,39 1,18 0,84 1,00 1,932,13 1,10 4,154,76 1,15 13,8617,94 1,29 6,15 1,00 6 0,200,26 1,28 0,67 1,00 1,68 1,00 3,49 1,00 10,26 1,00 5,156,20 1,20 7 0,22 1,00 0,64 1,00 1,63 1,00 3,40 1,00 12,4914,21 1,14 4,985,74 1,15 Resolution of all of the compounds was achieved on cellulose tris(4-methylbenzoate) with heksan- propan-2-ol (5%, v/v) with enantioselectivity from 1.06 to 1.15. On amylose tris(3,5-dimethylphenylcarbamate) only compounds 1-3 were resolved into enantiomers.

9. Fig. 6. The relationship between alkyl chain length and retention factors for compounds 1-5 on cellulose tris(4-methylbenzoate) (solid lines) and amylose tris(3,5-dimethylcarbamate) with hexane-2-propanol 95:5 (v/v) as a mobile phase. Flow rate 1 mL min -1 ; room temperature; λ = 242 nm. On cellulose tris(4-methylbenzoate)the retention factors of the compounds enantiomers diminished together with the growing number of carbon atoms in R aliphatic substituent reflecting growing steric and/or hydrophobic interactions of the substituents and the enantioselectivity was almost constant for the whole series of compounds. On cellulose tris(4-methylbenzoate) the retention factors of the compounds enantiomers diminished together with the growing number of carbon atoms in R aliphatic substituent reflecting growing steric and/or hydrophobic interactions of the substituents and the enantioselectivity was almost constant for the whole series of compounds. On amylose tris(3,5-dimethylphenylcarbamate) the retention factors exhibited smaller dependence on the carbon chain length in R substituent (changing from 6.15 to 6.93). Thus the destabilising effect of aliphatic R chain appeared to be much stronger on cellulose (changing from 4.15 to 8.22) than on amylose phase (changing from 6.15 to 6.93).

10. Fig. 7. The schematic structure of hydrogen bonds between amylose tris(3,5-dimethylcarbamate) and 1,4-disubstituted piperazines. amylose tris(3,5-dimethylcarbamate) and 1,4-disubstituted piperazines. amylose tris(3,5-dimethylcarbamate) analyte Thus the difference in the compounds behaviour on both stationary phases could be explained by considering that carbamates may form hydrogen bonds with appropriate acceptors via the amide hydrogen bond. Accordingly, hydrogen bond formation between the NH-carbamate group of the stationary phase and a carbonyl group of the solute may thus be considered (Fig. 7). The bond does not seem to be much influenced by the steric and hydrophobic factors.