GB2471293A

GB2471293A - The extraction of pharmacological agents from medicinal herbs using subcritical water

Info

Publication number: GB2471293A
Application number: GB0910849A
Authority: GB
Inventors: Kenneth Davison; Gary William Wheatley
Original assignee: Individual
Current assignee: Individual
Priority date: 2009-06-24
Filing date: 2009-06-24
Publication date: 2010-12-29
Anticipated expiration: 2029-06-24
Also published as: GB0910849D0; GB2471293B

Abstract

A method for preparing an extract containing pharmacologically active constituents from certain traditionally used medicinal herbs comprises extracting the botanical raw material with subcritical water and then removing the water to give a pharmaceutically acceptable extract. The extraction may be conducted at 150-180°C and/or 70-85 bar. The medicinal herb maybe Melissa officinalis (lemon balm), Salvia officinalis (sage), Thymus vulgaris (thyme), Trifolium pratense (red clover), Calendula officinalis (marigold) or Hypericum perforatum (St Johns Wort). These sub-critical water extracts may be similar in composition to those extracts obtained by means of methanolic extraction. The extract may be formulated as a self emulsifying drug delivery system in order to improve the bioavailability of the active constituents: a formulation may comprise the extract, lauroyl macrogolglyc elide and surfactant.

Description

Further Examples of Pharmacologically Active Extracts Prepared from Certain Medicinal Herbs in Traditional Use in Europe Without the Use of an Organic Solvent.

FIELD OF THE INVENTION

This invention relates to a method of producing an extract of certain herbs, used traditionally in Europe for their medicinal value, without the use of an organic solvent.

BACKGROUND TO THE INVENTION.

Higher plants are unsurpassed in their ability to produce biologically active molecules often of great complexity. These plant "secondary metabolites" have proven a fruitful source of new pharmaceuticals. Often the first use of these compounds has been in the form of traditional herbal medicines. However the structure of the active compounds is frequently so complex that synthetic chemistry can provide no economically viable route to the molecules and hence these have to be extracted from the plant material.

Conventional aqueous extraction of the biomass almost invariably results in a complex extract in which the target compound is diluted by a range of other unwanted phytochemicals derived from the plant, including proteins, sugars and other carbohydrates.

Extraction with a lower alcohol, typically ethanol or methanol, is traditionally a very popular means of obtaining a pharmacologically active preparation from a medicinal herb. The use of an alcohol has the advantage over an aqueous extraction that the highly polar compounds such as proteins, carbohydrates et cetera, which generally are inactive and serve only to add bulk and dilute the concentration of the active compounds, are not extracted.

However the obvious disadvantages of the use of an alcohol in extraction are that it is flammable, necessitating expensive precautions during processing, and also the recovery of the organic solvent for either disposal or for re-use after further purification is required. Methanol in addition is highly toxic, and residual levels must be rigorously limited in a pharmaceutical preparation. Indeed due to the high toxicity of methanol, it is frequently substituted by a mixture of the more polar water and the less polar ethanol to produce an extraction solvent of comparable polarity.

A more recent approach that has also been successfully applied to the production of a high potency extract is extraction with a sub-critical fluid (in effect a gas liquefied by the application of high pressure) most commonly carbon dioxide. One drawback of liquid carbon dioxide (LCO2) extraction is that the technique is limited to less polar compounds. It will not extract the compounds most readily soluble in methanol or aqueous ethanol, including the important classes of flavonoids, lignans and other related polyphenolic compounds. The importance of polyphenolic compounds as potential new drugs is increasingly being recognised; with anti-inflammatory activity, anti-microbial activity, estrogenic activity and a wide range of activities related to the suppression of tumour growth (including inhibition of angiogenesis and reversal of multidrug resistance) being well established.

A more recent example of a high pressure liquefied gas which is being evaluated as an extraction solvent and which promises capabilities not exhibited by LCO2 is high pressure superheated water at >150°C, termed "sub-critical water". Importantly, it is capable of solubilising more polar compounds than LCO2 extraction, including flavonoids and other polyphenolics. Sub-critical water has the unique property that the polarity of the solvent decreases in a predicable manner as temperature is increased; thus at 150°C the polarity of sub-critical water is equivalent to that of a 50:50 mixture of ethanol and water, and the polarity further decreases as the temperature is increased to 230°C to a value equivalent to that of pure methanol, A further advantage of sub-critical water extraction is that the high temperature and pressure produce high diffusion rates which promote very efficient extraction of the raw material The traditionally used medicinal herbs for which extraction with sub-critical water is described in this invention are all reported in the scientific literature to produce a methanol or aqueous alcohol extract exhibiting important pharmacological activities.

The methanolic or aqueous alcohol extracts of Melissa of/Icinalis (lemon balm), Salvia officinalis (sage), and Thymus vulgaris (thyme) all contain significant levels of rosmarinic acid and also diterpenes including carnosol and carnosic acid, along with a range of flavonoids. These compounds have all been demonstrated to exhibit anti-inflammatory actions and various activity inhibiting the growth of cancer cells. The diterpenes and flavonoids have also been reported to inhibit P-glycoprotein and other cellular drug efflux pumps responsible for multi-drug resistance in cancer chemotherapy.

Trfoliumpratense (red clover) contains significant levels of a range of flavonoids which are present in methanolic and aqueous alcohol extracts, including biochanin A and fonnononetin. Extracts of the herb are commonly used in the management of menopausal symptoms due to the estrogenic activity associated with the isoflavones.

The isoflavone biochanin A in addition exhibits very useful pharmacological properties, including both estrogenic properties and anti-cancer properties (inhibition of angiogenesis and inhibition of P-glycoprotein and other multi-drug resistance proteins).

Preparations of the flowers of Calendula officinalis (marigold) are used in a variety products for treating skin inflammation and are also incorporated in many cosmetics.

The active constituents are not fully identified but are thought at leat in part to be triterpene alcohols typified by faradiol.

Extracts of the medicinal herb St Johns Wort (Hypericumperforatum) are best known for the well established anti-depressant action but has also traditionally been used topically as an anti-inflammatory and modem pharmacological studies have supported this use.

Thus as the subcritical water extracts of the herbs as described in this invention are demonstrated to exhibit a composition essentially similar to that of the corresponding methanolic or aqueous alcohol extract, and in most cases also to contain the actual known active compounds, then it can be expected that these extracts will exhibit comparable pharmacological activities to these and to the compounds contained therein.

A problem limiting the use of flavonoids and related polyphenolic compounds in therapeutic situations in which systemic treatment rather than local application is required is poor water solubility. This is largely responsible for the low circulating plasma levels after a single oral dose and results in the majority of the dose being excreted without absorption.

Hence this invention also describes self emulsifying formulations which increase the water solubility of these compounds, thus increasing their oral bioavailability and therapeutic efficacy.

The scope of the invention is illustrated by the following series of examples, although in not any limitative sense.

SUNMARY OF THE INVENTION.

Example 1:

The extraction apparatus consists of: * Two suitable stainless steel vessels capable of resisting high temperature and pressure connected by stainless steel tubing, the first to act as a reservoir in which sub-critical water is produced prior to introduction to the second (extraction) vessel. These are contained in a thermostatted oven.

* The first vessel is connected via an inlet valve to a high pressure pump outside the thermostatted oven.

* The extraction vessel is connected via an outlet valve to a stainless steel receiver vessel outside the thermostatted oven but which is maintained at approximately 90°C.

* A valve from the receiver vessel allows the solution that accumulates to be transferred to a suitable storage vessel.

A schematic representation of a suitable arrangement of the extraction apparatus is provided as Figure 1.

Coarsely ground lemon balm botanical raw material is packed into the stainless steel extraction vessel. The system is then filled with deionised water and the temperature and pressure of the extraction vessel raised gradually to raised to a fixed temperature in the range 150-230°C, most preferably 150-180°C and a pressure of 85 bar respectively. The extraction system is then held at these conditions for up to 15 minutes before the resulting solution is forced from the extraction vessel into the receiver by passing a quantity of sub-critical water into the system to continue the extraction. Typically a mass of sub-critical water equivalent to twenty times the mass of botanical raw material is passed through this over the course of up to two hours.

Alternatively the same extraction may be achieved in a dynamic mode by passing the same quantity of sub-critical water continuously through the botanical raw material over the course of up to two hours.

The extract is isolated from the resulting solution resulting from the extraction by removing the water by evaporation, preferably carried out under reduced pressure to reduce the temperature required.

Alternatively the extract may be isolated by from the solution by removing the water by the process of either freeze drying or spray drying.

A TLC fingerprint representative of the extract (also termed a botanical drug substance) produced in this example is given in Figures 2a & 2b.

Example 2.

A method as described in example 1 wherein the botanical raw material is ground sage.

A TLC fingerprint representative of the extract produced in this example is given in Figures2a&3.

Example 3.

A method as described in example 1 wherein the botanical raw material is ground thyme.

A TLC fingerprint representative of the extract produced in this example is given in Figures 2a&4.

Example 4.

A method as described in example I wherein the botanical raw material is ground red clover.

A TLC fingerprint representative of the extract produced in this example is given in Figure 5.

Example 5.

A method as described in example 1 wherein the botanical raw material is milled marigold flowers.

A TLC fingerprint representative of the extract produced in this example is given in Figure 6.

Example 6.

A method as described in example 1 wherein the botanical raw material is St Johns Wort.

A TLC fingerprint representative of the extract produced in this example is given in Figure 7.

Example 7.

A Self Emulsifying Drug Delivery System (SEDDS) formulation for oral administration of the extracts containing the poorly water soluble active compounds can be prepared as follows: Extract (selected from the extracts produced in Examples 1-6) 15% w/w Lauroyl macrogoiglycerides EP (e.g Gelucire 44/14) 65-85% w/w Surfactant (e.g. Cremophor RF140 or Labrafac are suitable) 0-20% w/w.

The extract is dispersed with stirring in the molten lauroyl macrogolglycerides at 70- 80°C. The surfactant is then added and stirring continued for a furtherS minutes.

Using suitable automatic or manual equipment the molten mixture is then dispensed into two piece hard shell gel capsules which are then sealed.

Example 8.

A cream formulation for topical application of the extracts can be prepared as follows: Extract (selected from the extracts produced in Examples 1-6) 2% w/w Cetostearyl alcohol EP 7% w/w Macrogol cetostearyl ether (e.g. Cremophor A6 or A25) 3% w/w Liquid paraffin EP 12% w/w Parabens (e.g. Nipastat) 0.2% w/w Deionised water 67.8% w/w Propylene glycol EP 8% w/w The extract is dispersed in the propylene glycol at 70-80°C with stirring. Allother ingredients except the water are mixed at 80°C and then added with stirring to the water that was heated separately to 80°C. The dispersion of extract in propylene glycol is then added to this mixture maintained at 70-80°C with stirring. The formulation is then filled into tubes.

Example 9.

A hydroalcoholic gel formulation for topical application of the extracts can be prepared as follows: Extract (selected from the extracts produced in Examples 1-6) 2% w/w Ethanol EP 44% w/w Carbomer (e.g. carbopol 980 NF) 3% w/w De-ionised water 51% w/w Sodium hydroxide (aq) qs to neutralise The extract is dispersed in the ethanol at 50-60°C with stirring. The carbomer is then added slowly to the water with rapid stirring. The extract dispersion is then added to the aqueous carbomer whilst stirring. The resulting mixture is neutralised by slowly adding aqueous sodium hydroxide to produce a smooth semi-solid.

The formulation is then filled into tubes.

While the preferred embodiments of the invention have been described above, it will be recognised and understood by those skilled in the art that various modifications may be made therein, and the appended claims are intended to cover all such modifications which may fall within the spirit and scope of the invention.

Claims

CLAIMS1. A method of preparing an extract containing pharmacologically active constituents from certain traditionally used medicinal herbs, comprising reduction of the particle size of the raw material as appropriate, extracting the botanical raw material with sub-critical water, and then removing the water from the resulting solution to produce a pharmaceutically acceptable extract.
2. The method according to claims 1-4 wherein the extraction is earned out at a temperature in the range 150-230°C, and most preferably in the range in the range 150-180°C.
3. The method according to claim 1-4 wherein the extraction is carried out at a pressure in the range 15-100 bar sufficient to maintain the water in the liquid phase, and most preferably at 70-85 bar.
4. The method according to claims 1-4 wherein the medicinal herb is selected from the group consisting of the traditionally used parts of Melissa officinalis (lemon balm), Salvia officinalis (sage), Thymus vulgaris (thyme), Trfolium pratense (red clover), Calendula officinalis (marigold) and Hypericum perforarum (St Johns Wort) and also including species closely related to the foregoing.
5. The method according to claims 1-4 wherein the water is removed to yield the extract by evaporation.
6. The method according to claims 1-4 wherein the water is removed to yield the extract by freeze drying or spray drying.
7. The method according to any preceding claim wherein the sub-critical extraction of Melissa officinalis herb or a closely related species produces an extract with a TLC fingerprint substantially as illustrated in Figure 2a, which is essentially similar to the corresponding 50% aqueous ethanol extract and exhibits a significant spot due to rosmarinic acid. The component at Rf = 0.22 which is more abundant in the SWE extract than in the conventional solvent extract, is much reduced when the temperature of the sub-critical water extraction is reduced from 190°C to 150°C as illustrated in Figure 2b.
8. The method according to any preceding claim wherein the sub-critical extraction of Salvia officinalis herb or a closely related species produces an extract with a TLC fingerprint substantially as illustrated in Figure 2a, which is essentially similar to the corresponding 50% aqueous ethanol extract and exhibits a significant spot due to rosmarinic acid. The component at Rf= 0.22 which is more abundant in the SWE extract than in the conventional solvent extract, is much reduced when the temperature of the sub-critical water extraction is reduced from 190°C to 150°C as illustrated in Figure 3.
9. The method according to any preceding claim wherein the sub-critical extraction of Thymus vulgaris herb or a closely related species produces an extract with a TLC fingerprint substantially as illustrated in Figure 2a, which is essentially similar to the corresponding 50% aqueous ethanol extract and exhibits a significant spot due to rosmarinic acid. The component at Rf 0.22 which is more abundant in the SWE extract than in the conventional solvent extract, is much reduced when the temperature of the sub-critical water extraction is reduced from 190°C to 150°C as illustrated in Figure 4.
10. The method according to any preceding claim wherein the sub-critical extraction of Trfolium prarense herb or a closely related species produces an extract with a TLC fingerprint substantially as illustrated in Figure 5, which is essentially similar to the corresponding 50% aqueous ethanol extract.
11. The method according to any preceding claim wherein the sub-critical extraction of Calendula officinalis herb or a closely related species produces an extract with a TLC fingerprint substantially as illustrated in Figure 6, which is essentially similar to the corresponding 50% aqueous ethanol extract.
12. The method according to any preceding claim wherein the sub-critical extraction of Hypericum ofJIcinalis herb or a closely related species produces an extract with a TLC fingerprint substantially as illustrated in Figure 7, which is essentially similar to the corresponding 50% aqueous ethanol extract.
13. The use of a botanical drug substance as claimed in any of the preceding claims that consist essentially of botanical drug substances.
14. The use of a botanical drug as claimed in claim 13 further comprising excipients.
15. The use of a botanical drug as claimed in claim 13 wherein the botanical drug substances comprise total extracts derived from the botanical raw materials.
16. The use of a botanical drug as claimed in claim 13 wherein the botanical drug substances comprise more refined fractions derived from the total extracts of the botanical raw materials.
17. The use of a botanical drug as claimed in claim 13 wherein the botanical drug substances are standardised extracts.
18. The method according to any preceding claim wherein the pharmacologically active extract is formulated in a self emulsifying drug delivery system to improve the oral bioavailability of the active constituents.
19. The method according to clami 17 wherein the self emulsifying drug delivery system is based on the formulation described in Example 7.
20. The method according to claims 18-19 wherein the pharmacologically active extract as described in any preceding claim is selected from the list Melissa officinalis (lemon balm), Salvia officinalis (sage), Thymus vulgaris (thyme), Trfo1ium pratense (red clover) Calendula ofJIcinalis (marigold) and Hypericumperforatum (St Johns Wort) and alsospecies closely related to the foregoing.
21. The method according to any preceding claim wherein the pharmacologically active extract is formulated in a topical vehicle formulated to increase efficacy of the active constituents.
22. The method according to claim 20 wherein the topical vehicle is based on the formulation described in Examples 8 & 9.
23. The method according to claim 21-22 wherein the pharmacologically active extract as described in any preceding claim is selected from the list Melissa officinalis (lemon balm), Salvia officinalis (sage), Thymus vulgaris (thyme), Trfo1ium pratense (red clover), Calendula officinalis (marigold) and Hypericumperforatum (St Johns Wort) and also species closely related to the foregoing.Amended Claims have been filed as follows:CLAIMS1. A method of preparing an extract containing pharmacologically active constituents from certain traditionally used medicinal plants, comprising reduction of the particle size of the raw material as appropriate, extracting the botanical raw material with sub-critical water, and then removing the water from the resulting solution to produce a pharmaceuticallyacceptable extract.2. The method according to claims 1-6 wherein the extraction is carried out at a temperature in the range 150-230°C, and most preferably in the range in the range 150-180°C.3. The method according to claim 1-6 wherein the extraction is carried out at a pressure in the range 15-100 bar sufficient to maintain the water in the liquid phase, and most preferably at 70-85 bar.4. The method according to claims 1-6 wherein the medicinal plant is selected from the group consisting of the traditionally used parts of Melissa officinalis (lemon balm), Trfolium pratense (red clover), Calendula officinalis (marigold) and Hypericum perforatum (St Johns Wort), and also including species closely related to any of the foregoing.5. The method according to claims 1-6 wherein the water is removed to yield the extract by evaporation.6. The method according to claims 1-6 wherein the water is removed to yield the extract by freeze diying or spray diying.7. The method according to any preceding claim wherein the sub-critical water extraction of Melissa officinalis (lemon balm) botanical raw material or a closely related species produces an exiract with a TLC fingerprint substantially as illustrated in Figure 2, which is essentially similar to the corresponding 50% aqueous ethanol extract and exhibits a significant spot due to rosmarinic acid. The component at Rf = 0.22 which is more abundant in the sub-critical water extract than in the conventional solvent extract, is much reduced when the temperature of the sub-critical water extraction is creduced from 190°C to 150°C.8. The method according to any preceding claim wherein the sub-critical water extraction of Trfolium pratense (red clover) botanical raw material or a closely related species produces an extract with a TLC fingerprint substantially as illustrated in Figure 3, which is essentially similar to the corresponding 50% aqueous ethanol extract **S.* ...,: 9. The method according to any preceding claim wherein the sub-critical water * extraction of Calendula officinalis (marigold) botanical raw material or a closely related species produces an extract with a TLC fmgerprint substantially as illustrated in Figure 4, * : : which is essentially similar to the corresponding 50% aqueous ethanol extract * * 10. The method according to any preceding claim wherein the sub-critical water extraction of Hypericum perforazum (St Johns Wort) botanical raw material or a closely :... related species produces an extract with a TLC fingerprint substantially as illustrated in Figure * 5, which is essentially similar to the corresponding 50% aqueous ethanol extract S..I11. The use of a botanical drug substance as claimed in any of the preceding claims that consist essentially of botanical drug substances.12. The use of a botanical drug as claimed in claim 11 further comprising excipients.13. The use of a botanical drug as claimed in claim 11 wherein the botanical drug substances comprise total extracts derived from the botanical raw materials.14. The use of a botanical drug as claimed in claim 11 wherein the botanical drug substances comprise more refmed fractions derived from the total extracts of the botanical raw materials.15. The use of a botanical drug as claimed in claim 11 wherein the botanical drug substances are standardised extracts.16. The method according to any preceding claim wherein the pharmacologically active extract is formulated in a self emulsifying drug delivery system to improve the oral bioavailability of the active constituents.17. The method according to claim 16 wherein the self emulsifying drug delivery system is based on the formulation described in Example 5.18. The method according to claims 16-17 wherein the pharmacologically active extract as described in any preceding claim is selected from the list of subcritical water extracts of Melissa officinalis (lemon balm), Trfolium pratense (red clover), Calendula ofJicinalis (marigold) and Hypericum peiforatum (St Johns Wort), and also including species closely related to any of the foregoing.19. The method according to any preceding claim wherein the pharmacologically active extract is formulated in a topical vehicle formulated to increase efficacy of the active constituents.20. The method according to claim 19 wherein the topical vehicle is based on the formulations described in Examples 6 & 7 21. The method according to claim 19-20 wherein the pharmacologically active extract as described in any preceding claim is selected from the list of subcritical water extracts of Melissa officinalis (lemon balm), Trfolium pratense (red clover), Calendula officinalis (marigold) and Hypericum perforatum (St Johns Wort), and also including species closely related to any of the foregoing. * S S... * . S.... * . *S S. * S * S..S