Figure 1. The schema of the appliance for extraction under high pressure
(HPEP - High Pressure Extraction Plant, NOVA - Swiss)
Investigation of antimicrobial activity of the extracts Hypericum olympicum L. obtained by the extraction with CO2 under high pressure was carried out. Ethanolic solutions of the investigated extracts show good antimicrobial activity. Extract obtained under higher-pressure shows better antimicrobial effect. Application of HPE for manufacture of antimicrobial parapharmaceutical preparationswill depend upon the results of technoeconomic analysis.
Herbal species of the genus Hypericum L. have been known for a long time as being medicinal. The best known and the most investigated species of this genus is H. perforatum L. However, intensive and systematic investigations of other herbal species of the genus Hypericum L. have been carried out in the factory of chemical and pharmaceutical products "Zdravlje", Leskovac and at the Technological faculty in Leskovac. Within those investigations antimicrobial activity of the extracts H. olympicum L. obtained by the extraction with CO2 under high pressure was analysed.
Extraction under high pressure (HPE) by very compressed gasses as with solvents, becomes more often the subject of investigation of many research groups in the countries with mainly English and German spoken languages. Essential difference of HPE and conventional methods of extraction lies in the use of the solvents that require high pressure and specific temperature conditions in order to maintain in liquid state. Carbon dioxide has special position in HPE (1). Due to its suitable physical parameters (tc = 31°C, pc = 7.38 MPa), its inert nature as well as its physiological harmlessness, CO2 is a well-known means for extraction in food and pharmaceutical industry (2).
St. John's wort Hypericum olympicum L. used for the investigation in this paper has been collected on the Rujan mountain (Southeast Serbia, Yugoslavia). Herbal material has been dried at room temperature, in thin layer, and then ground in an electric mill and sifted through a sieve with a hole of 1 mm.
Extraction with CO2 under high pressure was carried out on the appliance HPEP - High Pressure Extraction Plant, NOVA - Swiss (Figure 1) under the following conditions:
1. p = 100 bar, t = 40°C, t = 2 h (extract 1)Mass of investigated samples of the drug was 50.0 g, pressure CO2 97.725 dm3/h (at 20°C), while the conditions in the separator were p = 15±1 bar and t = 22±1°C.
2. p = 300 bar, t = 40°C, t = 2 h (extract 2)
Antimicrobial activity of the extracts was investigated by the method of diffusion on cellulose disc, with the following 10 test microorganisms: Bacillus subtilis S, Escherichia coli 95, Pseudomonas aeruginosa, Klebsiella pneumoniae, Staphylococcus aureus ATCC 6538, Sarcina lutea ATCC 9341, Salmonella enteritidis, Aspergillus niger, Candida albicans ATCC 24493 and Saccharomyces cerevisiae ATCC 9763.
The yields of the extraction of St. John's wort H. olympicum L. with CO2 under high pressure are given in the Table 1.
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t = 40°C t = 2 h |
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t = 40°C t = 2 h |
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It can be seen from the Table 1 that the increase of pressure of CO2 causes the increase extraction yield.
Obtained extracts are dissolved in ethanol (c = 0.01 g/cm3), and their antimicrobial activity by the method of diffusion on cellulose disc was investigated (Table 2).
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| Escherichia coli 95 |
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| Bacillus subtilis S |
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| Sarcina lutea ATCC 9341 |
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| Salmonella enteritidis |
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| Staphylococcus aureus ATCC 6538 |
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| Klebsiella pneumoniae |
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| Pseudomonas aeruginosa |
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| Saccharomyces cerevisiae ATCC 9763 |
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| Candida albicans ATCC 24493 |
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| Aspergillus niger |
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Extract obtained by the extraction with CO2 under high-pressure (extract 2) shows better antimicrobial effect. The extracts show the best antimicrobial effect on fungus Candida albicans, while the culture Saccharomyces cerevisiae is resistant to both extracts.
With the changes of the conditions for extraction process of HPE could be optimised. Final decision on use of HPE for manufacture of antimicrobial preparations will be given through economic analysis and by monitoring the quality of obtained extracts.
Ethanolic solutions of H. olympicum L. HPE extracts show good antimicrobial effect.
Extract obtained under higher pressure shows better antimicrobial effect.
The extracts show the best antimicrobial effect on fungus Candida albicans, while the culture Saccharomyces cerevisiae is resistant to both extracts.
Use of HPE for manufacture of antimicrobial parapharmaceutical preparations will depend on the results of technoeconomic analysis.
Bunzenberger G., Lack E. and Marr R. (1984): CO2-Extraction: Comparasion of Super- and Subcrtical Extraction Conditions. German Chemical Engineering 7, 25-31.
Gährs H.J. (1984): Applications of Atmospheric Gases in High Pressure Extraction. Berichte der Bunzen-Gesselschaft für Physikalische Chemie 88, 894 - 897.