SYNTHESIS OF ⳑ-PROLYLGLYCINE

CAUTION: Benzyl chloroformate and ethyl chloroformate are highly lachrymatory and should be handled with great care in a fume cupboard; disposable plastic gloves should be worn.

Glycine ethyl ester hydrochloride:

• Esterify 15g (0.2 mol) of glycine in 100 ml of absolute ethanol with the aid of about 10g of dry hydrogen chloride. 

• Recrystallise the product from the minimum of absolute ethanol. The yield of glycine ethyl ester hydrochloride, m.p. 144 °C, is 18.5g (66%).

Benzyloxycarbonyl-ⳑ-proline:

• Dissolve 4.6g (0.04 mol) of ⳑ-proline in 10 ml (0.04 mol) of 4ᴍ sodium hydroxide solution, cool to °C in an ice bath, and add 7.0g (0.041 mol) of benzyl chloroformate and 10 ml (0.04 mol) of 4𝖬 sodium hydroxide solution alternately and portionwise, with shaking and cooling, during 30 minutes. 

• Allow the mixture to warm to room temperature with intermittent shaking during 1 hour, extract with 2 x 15-ml portions of ether, and acidify the aqueous phase (Congo red) with dilute (1:1) hydrochloric acid. 

• Extract the liberated oil with 4 x 10-ml portions of ether, dry over anhydrous sodium sulphate, and remove the ether to obtain 8.8g (88%) of benzyloxycarbonyl-L-proline which crystallises slowly. 

• A sample recrystallised from ether-light petroleum (b.p. 40-60 °C) has m.p. 75 °C. 

Benzyloxycarbonyl-ⳑ-proIyglycine:

• Dissolve 6.2g (25 mmol) of benzyloxy-carbonyl-ⳑ-proline in 15 ml of dry chloroform containing 2.5g (3.4 ml, 25 mmol) of dried redistilled triethylamine and cool the mixture in ice while adding 2.7g (2.4 ml, 25 mmol) of ethyl chloroformate steadily and with shaking. 

• Set the mixture aside at 0°C for 15 minutes and then stir into the semi-solid mass a suspension of 3.5g (25 mmol) of glycine ester hydrochloride in 20 ml of dry chloroform containing a further 2.5g of dry triethylamine. 

• Carbon dioxide is evolved and the solids go into solution. 

• Set the mixture aside at room temperature for 30 minutes, warm at 50 °C for 10 minutes, and then wash it successively with 15 ml of water, 10 ml of 1𝖬 hydrochloric acid, 2 x 10-ml portions of 0.5ᴍ sodium hydrogen carbonate solution and 10 ml of water. 

• Dry the solution over anhydrous sodium sulphate and remove the chloroform under reduced pressure to obtain 8.3g (99%) of benzyloxy-carbonyl-ⳑ-prolylglycine ethyl ester as an oil. 

• Dissolve the entire product in a mixture of 26 ml of 1𝖬 sodium hydroxide solution and 10 ml of acetone and set it aside at room temperature for 1 hour. 

• Concentrate the solution some-what under reduced pressure to remove most of the acetone and then acidify (Congo red) with dilute (1:1) hydrochloric acid. 

• Extract the precipitated oil with ethyl acetate, dry over anhydrous sodium sulphate and remove the solvent under reduced pressure. 

• The resulting oil crystallises slowly on cooling and scratching under a small quantity of ethyl acetate. 

• To recrystallise dissolve the product in the minimum of hot ethyl acetate, cool somewhat and add an equal volume of ether, and then seed and set aside overnight. The yield of benzyloxycarbonyl-ⳑ-prolyglycine, m.p. 122-123 °C, is 5.3g (69%). A sample crystallised from water has m.p. 125 °C.

ⳑ-Prolyglycine:

• Dissolve 4.6g (15 mmol) of the foregoing product in 75 ml of methanol and add 75 ml of water and 2 ml of glacial acetic acid. 

• Place 0.4g of palladium black into a Drechsel bottle and add the above solution, taking care to rinse all traces of catalyst down below the surface of the liquid. 

• Connect the Drechsel bottle inlet tube to a hydrogen cylinder and vent the exit tube to the external atmosphere or to a fume cupboard via a second Drechsel bottle charged with lime-water. 

• Suspend the bottom of the inlet tube just above the surface of the liquid in the reaction bottle and pass in hydrogen to displace the air. 

• Lower the Drechsel head to close the system and allow hydrogen to bubble through the reaction mixture at a rate sufficient to agitate the catalyst. 

• Continue the passage of hydrogen for about 2 hours, shaking periodically. 

• The onset of the reaction, accompanied by the liberation of carbon dioxide, is indicated by typical carbonate precipitation in the lime-water bubbler; continue to pass hydrogen until carbon dioxide evolution ceases (confirm using a fresh charge of lime-water). 

• Then filter off the catalyst (CARE - pyrophoric when dry) and evaporate the solution to dryness under reduced pressure. 

• Recrystallise the residue by dissolving it in the minimum of hot water, adding hot methanol and cooling. 

• ⳑ-Prolylglycine is obtained as the monohydrate; yield 2.4g (86%), m.p. 226-227 °C (decomp.), [𝜶]ᴅ¹⁷-21.9° (c 3.9 in H₂O). Check the homogeneity of the product by t.l.c. on silica gel using the system butan-1-ol-ethanol-acetic acid-water, 9:3:2:4; R𝑓 0.14.