WO2000066136A1

WO2000066136A1 - Water extract of chamomile, process for obtaining it and use as an antifungal agent

Info

Publication number: WO2000066136A1
Application number: PCT/IB2000/000549
Authority: WO
Inventors: Mohsen Daneshtalab; Saeed Sattari
Original assignee: Cv Therapeutics, Inc.
Priority date: 1999-04-30
Filing date: 2000-04-28
Publication date: 2000-11-09
Also published as: EP1185280A1; AU4136600A

Abstract

A process for preparing a water extract of chamomile, the process comprising extracting chamomile flowers with an alcohol extraction solvent to produce an alcohol extract and a solid residue and extracting the solid residue with a water extraction solvent to produce a water extract, and the use of the water extract as a pharmaceutical composition for the treatment of topical, systemic, mucosal and lung-invasive fungal infections, or for the preparation of a disinfectant for the treatment of superficial fungal infections.

Description

WATER EXTRACT OF CHAMOMILE , PROCESS FOR OBTAINING IT AND USE AS AN ANTIFUNGAL AGENT

This application claims priority under 35 USC § 1 19(e) of provisional application serial no. 60/131 ,880. filed April 30, 1999, incorporated by reference.

Background of the Invention

Antifungal therapy in immunocompromised patients such as those with AIDS or organ transplant is a booming business (a five billion dollars per annum sales is reported for the systemic antifungal remedies in 1997). Although some medications such as amphotericin B or azole antifungals are currently used for the treatment of systemic and deep-seated mycosis, they are prone to toxicity and emergence of resistance. Herbal medicines can be considered as the logical substitute for the synthetic products in many aspects. The herbal remedies with the history of human use are one of the beneficial alternatives for the therapy of different diseases.

Chamomile is a herbal remedy that grows freely everywhere. Its dried flowers and various parts contain flavons and coumarines, and the essential oil contains up to 50% α-bisacolol, as well as chamazulene. It is traditionally used as a nerve tonic, appetite stimulant, sleep aid, digestive aid, and flu remedy. It 0 relieves bladder, cold, asthma, colitis, fever, headaches, hemorrohids, muscle cramp, and pain. It is also effective in the treatment of rheumatism, worms, and jaundice. At present, different products of chamomile are on the market, which are all originated from its essential oil (which is obtained through a steam- distillation process) or methanolic extracts.

Summary of the Invention

In this invention we would like to present a new technology and new application of chamomile as an anti-infective remedy. Namely, we have successfully identified that an aqueous extract of chamomile has an impressive activity against one of the most pathogenic fungi (Aspergill s sp.), which is a fatal organism which targets, among others, immunosupressed patients, such as those with AIDS or having recent organ transplant. This is the first reported indication of effectiveness of chamomile against pathogenic fungi responsible for systemic and nosocomial infections. This invention includes the following:

• a process for the preparation of a water extract of chamomile, the process comprising extracting chamomile flowers with an alcohol extraction solvent to produce an alcohol extract and a solid residue and extracting the solid residue with a water extraction solvent to produce the water extract;

• a water extract of chamomile produced by the process;

• the use of the water extract as a pharmaceutical composition or for the preparation of a pharmaceutical composition for the treatment of topical, systemic, mucosal and lung-invasive fungal infections, preferably against Aspergillus species, such as A. niger and A. fumigatus;

• the use of the water extract for the preparation of a disinfectant for the treatment of superficial fungal infections; and

• a formulation comprising the water extract, for use to treat systemic, invasive or superficial fungal infections.

Description of the Preferred Embodiments

All of the following percentages are weight-percentages, unless otherwise indicated.

In a preferred extraction procedure, flowers (dried or fresh, preferably dried) of Matricaήa recutita (chamomile) are ground and macerated with an alcohol, preferably a C_rC₆ alcohol, and most preferably MeOH or EtOH. It is preferred that the alcohol be at least 85 % alcohol, most preferred absolute alcohol . After separation of the alcohol extract, the solid residue is extracted with water. Preferably, the alcohol extraction procedure removes at least 50% of the essential oils from the chamomile flowers, more preferably at least 60% , 70% , 80% , 90% or 95 % of the essential oils. It is most preferred that essentially all (e.g. , at least 98 %) of the essential oils or all (i.e. , 100%) of the essential oils are removed via the alcohol extraction procedure.

Preferably, the water extract is thereafter concentrated, optionally under reduced pressure. If desired, the water may be evaporated to near dryness, to obtain a brown sludge, or evaporated to dryness, to obtain a brown powder. The preferred residual water content in the extract of the invention is less than 40% , more preferably less than 30 % , 20% , 10% or 5 % . The residual water concentration of the extract (i.e. , the amount of water remaining in the extract) depends on the particular need. It is preferred to evaporate the water to near dryness or dryness, to obtain a brown sludge or dry powder, since these forms are easiest to store. However, it may also be useful to simply concentrate the extract in liquid form, if long-term storage is not necessary. Preferably, the liquid water extract is kept under refrigeration (preferably at a temperature under 15°C). The liquid water extract remains active for 30 days or more, especially if kept under refrigeration.

The sludge or powder extract of the invention may be formulated into a solid-based dosage form for therapeutic use (such as tablet, capsule, etc.), or the sludge or powder may be dissolved in a suitable carrier for liquid formulation (such as water, saline solution, etc.). Alternatively, the extract may be used as is in its concentrated liquid form (i.e. , without evaporating the extract to near dryness or dryness). The concentrated liquid form, or the sludge or powder combined with a suitable carrier, may be formulated for therapeutic use or for se in application to surfaces as a disinfectant.

As desired, before formulation into a therapeutic dosage form or for use in application to surfaces, as noted above, the extract of the invention may be subjected to one or more further extraction procedures to further purify the active components in the extract. For example, the brown sludge or powder may again be extracted one or more times with an alcohol (such as MeOH), and after alcohol removal, the solid residue may again be extracted one or more times with water. The further extraction procedure may be repeated a plurality of times before the extract is formulated into a suitable therapeutic dosage form, or combined with a suitable carrier for application to surfaces.

The constituents of the extract of the invention may be determined by NMR and LC-MS-MS methods, which are well known to those of skill in the art, and will therefore not be described in detail.

The water soluble extract of the invention is selectively most effective against Aspergillus sp . (the major pathogenic fungi affecting immunosuppressed patients, such as those with AIDS, or recent recipients of organ transplant, and those under cancer chemotherapy). The water soluble extract of the invention may be used as a prophylactic, as well as a treatment of topical, systemic, mucosal and/or lung-invasive aspergillosis in, for example, immunosuppressed patients. The water soluble extract may also be used as a disinfectant on surfaces. The easy process for the large scale preparation of the water soluble extract of the invention, as well as the desirable accessibility of raw material (chamomile flowers), makes this product an economically feasible nutraceutical/pharmaceutical. Since the water extract is preferably in a stable- solid form, preparation of different dosage forms, such as capsules, tablets, droplets, syrup, infusable or injectable forms, are feasible (each of these can be prepared with standard procedures known in the art). Due to a long history of utilization of chamomile products in humans, no major toxic effects are expected, even from large dosages. In the meantime, due to its water solubility, the extract of the invention provides a very favorable pharmacokinetic profile, even while using different routes of administration. The extract of the invention may be administered to a patient in an amount of 0.1 to 1000 mg/kg per day in one or more doses, optionally in a liquid carrier, depending on age, sex, infirm, etc. Suitable dosages are well within the skill of the ordinary artisan.

EXAMPLES

In a 5 liter separatory funnel, dried-ground chamomile flowers (750 g) were macerated in methanol (2 liters) overnight. The methanol fraction was separated, and the process was repeated four times. The total fractions were concentrated under reduced pressure to obtain 150 g of deep-green thick oil (extract A). The remaining solid material was air-dried. The remaining air-dried material was macerated in water (2 liters) overnight, and the water-soluble fraction was separated. The process was repeated four times. The collected fractions were concentrated under reduced pressure to obtain 150 g of a deep-brown solid (extract B).

Dried powder of extract B was subjected to a second MeOH extraction procedure. This second MeOH extract (extract C) was dried. The MeOH- insoluble part was dissolved in water. The resulting solution was dried to obtain a brown powder (extract D). Alternatively, a concentrated solution of extract B was prepared and stored in a refrigerator for 30 days (extract F).

An antifungal assay was carried out using the microdilution method. The fungal strains used were Candida albicans ATCC 14053, Cryptococcus neoformans PLM 489. Saccharomyces cerevisiae PLM 454, and Aspergillus niger PLM 1140. The methanol extracts generally exhibited activity against C. albicans, C. neoformans, and S. cerevisiae. with no activity against . niger. In contrast, the water extracts of the invention generally exhibited no activity against the three strains sensitive to the methanol extracts, while exhibiting strong activity against A. niger. The results are presented in the following Table.

Table 1 : Antifungal activity of several extracts of chamomile against several species of fungus

(minimum inhibitory concentration, expressed as mg/ml, except as noted)

Legend

N.A. : not active

(A) MeOH extract of dried chamomile flowers

(B) Water extract of solid material which remained insoluble after MeOH extraction (dried)

(C) MeOH-soluble part of extract B

(D) Water-soluble part of remaining material of extract B, after preparation of extract C (E) Equal mixture of extracts C and D

(F) Extract B, kept in solution form under refrigeration for 30 days

(G) and (H): standards for comparison

Relevant Literature

Mann C, Staba EJ (1986). The Chemistry, Pharmacology, and Commercial Formulations of Chamomile; in Herbs, Spices, and Medicinal Plants: Recent Advances in Botany, Horticulture, and Pharmacology; Craker LE, and Simon JE Editors, Vol. 1, 235-280, Oryx Press, Phoenix, Arizona, USA. This publication is incorporated by reference.

Claims

We claim:

1. A process for preparing a water extract of chamomile. the process comprising

(a) extracting chamomile flowers with an alcohol extraction solvent to produce an alcohol extract and a solid residue and

(b) extracting the solid residue produced in step (a) with a water extraction solvent to produce a water extract.

2. The process of claim 1 , wherein in said step (a), the alcohol extract contains at least 50% by weight of the essential oils contained in the chamomile flowers.

3. The process of claim 1 , wherein in said step (a), the alcohol extract contains at least 90 % by weight of the essential oils contained in the chamomile flowers.

4. The process of claim 1 , wherein in said step (a), the alcohol extract contains at least 98 % by weight of the essential oils contained in the chamomile flowers.

5. The process of any of claims 1-4. further comprising repeating steps (a) and (b) at least once with the water extract produced in step (b).

6. The process of any of claims 1-5, wherein the chamomile flowers are ground in or before said step (a).

7. The process of any of claims 1-6, wherein the alcohol is a C,-C₆ alcohol.

8. The process of any of claims 1-7, further comprising drying the water extract produced in step (b) to produce a concentrated water extract having a residual water content of less than 40 % by weight.

9. A water extract of chamomile, produced by the process of any of claims 1-8.

10. Use of a water extract as claimed in claim 9, in combination with one or more pharmaceutically acceptable carriers and/or excipients, for the preparation of a pharmaceutical composition for the treatment of topical, systemic, mucosal and lung-invasive fungal infections.

11. Use of a water extract as claimed in claim 9 as a pharmaceutical composition for the treatment of topical, systemic, mucosal and lung-invasive fungal infections.

12. Use of claims 10 or 11 , wherein the fungal infection is caused by an Aspergillus species.

13. Use of claim 12, wherein the Aspergillus species is A. niger or A. fumigatus.

14. Use of a water extract as claimed in claim 9, in combination with one or more carriers, for the preparation of a disinfectant for the treatment of superficial fungal infections.