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Antibacterial in vitro effects of preparations from Anthroposophical Medicine

BMC Complementary and Alternative MedicineBMC series – open, inclusive and trusted201616:372

https://doi.org/10.1186/s12906-016-1350-3

Received: 21 January 2016

Accepted: 14 September 2016

Published: 22 September 2016

Abstract

Background

Medications from Anthroposophical Medicine (AM) are clinically used for the treatment of infections within a whole medical system but have not yet been evaluated regarding antibacterial effects. The aims of this study was to investigate antibacterial activity of AM medications in cell culture.

Methods

Screening of AM drug registers for preparations used to treat any kind of infection and being available in dilutions ≤ D2 and without alcoholic content. Selected medications were screened for antimicrobial activity against Bacillus subtilis, Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa using the agar diffusion method. For antimicrobial active preparations growth kinetics (drop plate method) and minimal inhibitory concentrations (MIC, macrodilution method) were determined.

Results

Thirty-three preparations matched the selection criteria and were chosen for own experiments. One of them (Berberis Decoctum D2) exhibited bactericidal activities against Bacillus subtilis and Staphylococcus aureus, including methicillin resistant strains. The MIC could be determined as 5 mg/ml. The effects could be related to the content of berberine in the extract. No activity towards gram-negative bacteria was found. The other tested extracts had no antibacterial effects.

Conclusion

Berberis Decoctum D2 which is used in AM to treat infections exhibits bactericidal effects on Staphylococcus aureus, including methicillin resistant strains.

Keywords

Staphylococcus MRSA Berberis radix Berberine Parenteral use

Background

Antimicrobial resistance is one of the major problems of modern medicine. Considering the fast development of resistance to antibiotics in bacterial species such as Mycobacterium tuberculosis [1] or Staphylococcus aures [2], it becomes obvious that there is necessity to combat further resistance expansion [3]. However there is a concerning stagnation in the development of new antimicrobial agents [4] although more and more mechanisms of bacterial resistance are being discovered. For the purpose of developing new antimicrobial agents natural substances should be considered as a promising source [5]. Besides traditional antibiotics of microbial origin like penicillin, there is clear evidence that plant-derived preparations have antimicrobial potential [6]. They are able to synergize traditional antibiotics and therefore reduce the required dosage for infection control [7, 8] and there is evidence that plant-derived preparations are able to modify mechanisms of resistance [5, 9]. Examples for commonly used phytotherapeutics in infection control are the root extracts from Pelargonium sidoides in the treatment of the common cold [10] or Horseradish and Tropaeolum in the treatment of urinary or respiratory infections [1113]. Another focus of current research is the role of bacteria for the health of the human body. Referred to as the Human Microbiome it is evident today that those billions of microbes, especially located in the human gut, play a decisive role in the strengthening of health and a balanced immune system [14]. The pathogenesis of the metabolic syndrome [15] as well as allergic and autoimmune disorders such as bronchial asthma [16] or inflammatory bowel diseases [17, 18] is nowadays considered to be at least in part related to the human microbiome and especially to the gut microbiota. Antibiotics are well-known for their hurtful effects on this sensitive system of intestinal microflora. It is not uncommon that patients need microflora reconstruction after broad-spectrum antibiotic treatment. In this regard it is particularly interesting to examine substances with selective effects on the gut microflora or on bacterial strains in general: substances which have the characteristic of combatting harmful microbes but spare the physiological flora or even supporting and protecting it. Therein lies another possible role of plant-based antimicrobials in contrast to conventionally used antibiotics. Usually their potency isn’t as resounding that they would eradicate the majority of microbes of intestinal microflora and therefore provide niches for selectivity and protective potential.

Drugs from Anthroposophical Medicine (AM) have to our knowledge not yet been explored regarding their antimicrobial effects. Their manufacturing process is sophisticated and often different from manufacturing process in phytotherapy [19]. They are amongst others used to treat infectious diseases, despite they are not claimed to possess antimicrobial effects. They are instead designed to improve the self-healing capacity of the body in these diseases [20]. Nevertheless they are a possible source of new types of antimicrobial compounds. Preparations of AM are traditionally being used in German-speaking areas. The drug compendia contain approximately 1500 different medicinal products including plant-based, animal-based, mineral and combined preparations [21, 22]. The aim of this study was to investigate the antimicrobial potential of plant-based preparations of AM.

Methods

Compendia of drugs from Anthroposophical drug manufacturers [21, 22] have been searched for preparations in dilutions ≤ D2 and free of alcoholic content. Higher dilutions have been excluded in order to guarantee that the preparations contained active substances. Alcoholic content was not accepted because it exhibits antimicrobial effects itself. Preparations with more than one plant have also been excluded in order to observe merely the effects of individual plants. Out of the remaining preparations those were chosen for proper experiments which were used for the treatment of infections in AM.

Bacterial strains used for antimicrobial assays

Bacillus subtilis (ATCC 6633), Escherichia coli (ATCC 25922), Methicillin-susceptible Staphylococcus aureus (MSSA, ATCC 29213), Pseudomonas aeruginosa (ATCC 27853), Methicillin-resistant Staphylococcus aureus (MRSA, ATCC 43300) and clinical isolates of MSSA (MSSA 1883, 2114, 2289) and MRSA (MRSA 4331) from the Hygiene and Molecular Laboratory of the Institute of Environmental Health Sciences and Hospital Infection control of the University Medical Center, Freiburg, Germany were chosen for own experiments.

Activity screening following the agar diffusion method

A bacterial suspension of one of the microbial strains listed above was produced in isotonic saline solution by visual comparison to a 0.5 McFarland turbidity standard (bioMérieux, Germany). This standard corresponds to a concentration of 108 colony forming units (CFU)/ml. This solution was used to prepare bacterial smears in three layers on Columbia Blood Agar plates (Thermo Scientific, Great Britain). The test substances were then brought onto the bacterial layer in drops of 10 μl at defined positions. The agar plates were incubated for 24 hours at 37 °C.

Growth Kinetics following the drop plate method

An overnight culture was prepared the day before by solving 1-3 colonies of a bacterial strain in 5 ml of Mueller-Hinton broth (MHB) (Merck KGaA, Germany) and by incubating this solution for 24 hours at 37 °C in a shaker (Certomat® ; B Braun, Germany) at an intensity of 100 t/min. The overnight culture was then set to a concentration of 108 CFU/ml in a Turbiditymeter (Micro Scan, US) and diluted to a concentration of 106 CFU/ml. Control and test samples were prepared by mixing 1000 μl of double-concentrated MHB and 1000 μl of distilled water (control) or 1000 μl of the test substance, respectively. 20 μl of bacterial suspension of 106 CFU/ml were finally added to control and test samples. Up to six samples could be handled simultaneously. All samples were distributed into airtight Eppendorf tubes by pipetting 200 μl of each sample into eight Eppendorf tubes (two, each for 0, 4, 8 and 24 hours incubation). The 4, 8- and 24-hour-Eppendorf tubes were incubated at 37 °C in a shaker (Certomat®; B Braun, Germany) at an intensity of 100 t/min. The 0-hour-Eppendorf tubes were serially diluted with Mueller-Hinton broth using a microtiter plate. 5 μl of each concentration was afterwards pipetted onto Mueller-Hinton Agar-Plates (Merck KGaA, Germany). Five to six dilution steps fit the agar plate properly. In the same way was dealt with the t4-, t8- and t24-eppendorf tubes after the particular incubation times. All Mueller-Hinton-Agar-Plates were incubated at 37 °C for 24 hours. After 24 hours single colonies of bacterial growth could be counted in the areas where the former 5 μl-drops have been put on the Mueller-Hinton-Agar-Plate. The counted colonies yielded a specific growth value for each sample and incubation time.

Determination of the Minimal Inhibitory Concentration (MIC)

The MIC was determined in CAMHB (Cation Adjusted Mueller-Hinton Broth) according to the guidelines of CLSI in macrodilution-method. For Berberis Decoctum D2 a serial dilution has been performed with a concentration of 5 mg/ml in the first and a final concentration of 9.8 μg/ml in the last tube. The initial concentration of bacteria was set to 106 CFU/ml and was diluted to 105 CFU/ml in the samples. The samples were incubated at 36 °C and the MIC was determined as the lowest concentration without visible bacterial growth after 24 hours of incubation.

Statistics

All experiments on bacterial strains which play a major role in human infections and showed noteworthy effects after 24 hours have been carried out at least twice on the same bacterial strain or different clinical isolates of one bacterial species. Because the growth of bacteria in a growth kinetic assay varies to a small extend between different experiments an inhibitory effect was defined as the difference of at least 1 logarithmic unit between test sample and control. Thus only marked differences of growth were included.

Results and Discussion

Eighty four preparations with a total of 54 different ingredients matched the inclusion criteria and have been selected out of the Anthroposophical drug registers. 33 of those were then chosen for proper experiments because they were either used for infection control in AM or because they were available in dilutions ≤ D2 (Table 1). It can be assumed that the majority of all available and eligible preparations have been screened.
Table 1

Preparations of AM with botanical name and dosage form which were chosen for proper experiments

Botanical name

listed preparation of AM

dosage form

Arnica montana

Arnica e planta tota (Arnica montana e planta tota ferm 33c) D2

preparation for injection

Astragalus exscapus

Astragalus exscapus D3

preparation for injection

Belladonna

Belladonna Rh D3

aqueous solution

Berberis vulgaris

Berberis, Decoctum D2

preparation for injection

 

Berberis, Fructus Rh D2

preparation for injection

Betula pendula

Betula, Cortex, Decoctum D2

preparation for injection

Bryonia cretica

Bryonia D3

preparation for injection

Bryophyllum

Bryophyllum 5 %

preparation for injection

Selenicereus grandiflorus

Cactus ex herba (Selenicereus grandiflorus ex herba ferm 33d) D2

preparation for injection

Cinnamomum camphora

Camphora (Camphora aquos.) D3

preparation for injection

Cetraria islandica

Cetraria praeparata (Cetraria islandica) 2 %

preparation for injection

Chamomilla recutita

Chamomilla, Radix, Decoctum D3

preparation for injection

Cichorium intybus

Cichorium Rh D3

aqueous solution

Cydonia oblonga

Cydonia e fructibus (Cydonia oblonga e fructibus ferm 33b) D2

preparation for injection

Echinacea angustifolia

Echinacea angustifolia Rh D3

preparation for injection

 

Echinacea e planta tota (E. pallida e planta tota ferm 33c) D2

preparation for injection

Gentiana lutea

Gentiana lutea Rh 5 %

aqueous solution

Geum urbanum

Geum urbanum e radice (WS: G.u. e radice ferm 33c) D3

preparation for injection

 

Geum urbanum e radice D1

mother tincture

Levico

Levico (Levico water) D3

preparation for injection

Levisticum

Levisticum Rh D3

aqueous solution

Mandragora officinarum

Mandragora officinarum e radice ferm 34d, D2

preparation for injection

Nicotiana tabacum

Nicotiana e foliis (Nicotiana tabacum e foliis ferm 33b) D2

preparation for injection

Oxalis acetosella

Oxalis, Folium Rh D3

preparation for injection

Prunus spinosa

Prunus spinosa, Fructus Rh D3

preparation for injection

 

Prunus spinosa, Summitates Rh D3

preparation for injection

Rosmarinus officinalis

Rosmarinus, Infusum 5 %

preparation for injection

Urginea maritima

Scilla e bulbo (Urginea maritima var. Rubra e bulbo ferm 33b) D2

preparation for injection

Solidago virgaurea

Solidago virgaurea ex herba (S. virgaurea ex herba ferm 33c) D3

preparation for injection

 

Solidago virgaurea ex herba D1

mother tincture

Symphytum officinale

Symphytum e radice (S. officinale e radice ferm 34c) D2

preparation for injection

Taraxacum officinale

Taraxacum e planta tota (T. officinale e planta tota ferm 34c) D3

preparation for injection

 

Taraxacum e radice (autumnale) (T.off. e radice ferm 34c) D3

preparation for injection

Out of the 33 preparations screened with the agar diffusion method, four (Berberis Decoctum D2, Betula Cortex Decoctum D2, Solidago virgaurea Mother Tincture and Geum urbanum Mother Tincture) showed antimicrobial effects. The antimicrobial activity of the mother tinctures Solidago virgaurea and Geum urbanum could be related to their pH-values (3.5 and 5.0, respectively). In a buffered assay with a neutral pH (7) the antimicrobial activity vanished. The two tinctures were therefore excluded from further experiments. It is advisable to determine the pH of plant-based preparations routinely before performing antimicrobial assays [6], not only because acidity itself has effects on bacterial growth behavior but also because it influences the impacts of active ingredients in preparations [23].

All further experiments have been performed using the drop plate method. Berberis Decoctum D2 exhibited bactericidal effects on B. subtilis and MSSA (ATCC 29213) as shown in Fig. 1. Betula Cortex Decoctum D2 did not show any noteworthy effects and was therefore excluded from further experiments.
Fig. 1

Berberis Decoctum D2 – effects on bacterial growth after 0, 4, 8 and 24 hours on logarithmic scale. Control sample: distilled water (DW); test sample: Berberis Decoctum D2. Control sample and test sample were added in a ratio of 1:2 to the culture medium. Effects are visible for B. subtilis (ATCC 6633) (n = 1) and MSSA (ATCC 29213) (n = 3) after eight and after 24 hours. Effects can be considered as bactericidal. No relevant effects after 24 hours could be found for E. coli (n = 1) and P. aeruginosa (n = 1)

The bactericidal impact of Berberis Decoctum D2 could be confirmed by demonstrating the same effects on clinical isolates of MSSA (abscess-associated 1883, catheter-associated 2114, wound swab 2289) and on one clinical isolate of a multi-drug resistant strain MRSA (wound swab 4331). Because B. subtilis plays no major role in human infections further experiments were focused on S. aureus strains.

Figure 2 shows the same outcomes for all tested S. aureus strains including one multi-drug resistant strain. Berberis Decoctum D2 is manufactured from the bark and roots of Berberis vulgaris and contains the alkaloid berberine which has been described as antimicrobial active in several publications [24]. According to the manufacturers information one ampoule of Berberis Decoctum D2 contains 10 mg of the dried drug of Berberis cortex and the bark used for manufacture of the injectable contains at least 2 % of alkaloids which can be mainly considered as berberine. During growth kinetics Berberis Decoctum D2 was used in a concentration of 1:2 to the culture medium, which equals a dosage of 5 mg/ml of the dried drug in the experiment. For MSSA (ATCC 29213) and MRSA (ATCC 43300) the MIC could be determined as 5 mg/ml. A lower dosage wasn’t able to exhibit bactericidal effects. The MIC values of pure berberine for MSSA (ATCC 29213) and MRSA (ATCC 43300) were 64 μg/ml and 256 μg/ml, respectively. These values equal approximately the proclaimed concentration of berberine in the injectable Berberis Decoctum D2 (100 μg/ml). It can, therefore, be assumed that the effects of Berberis Decoctum D2 are due to the content of berberine.
Fig. 2

Effects on bacterial growth of Staphylococcus aureus strains (clinical isolates, each n = 1) after 0, 4, 8 and 24 hours on logarithmic scale. Control sample distilled water (DW) and test sample Berberis Decoctum D2 were added in a ratio of 1:2 to the culture medium. Bactericidal effects are visible for all S. aureus strains

Compared to common antibiotics, the concentrations of Berberis Decoctum D2 needed for bactericidal effects are considerably higher. Interesting for systemic usability would be a concentration <100 μg/ml for plant extracts or <10 μg/ml for isolated compounds [6]. In an assay revealing the effects of Berberis Decoctum D2 on human lymphocytes we found that Berberis Decoctum D2 is inducing apoptosis in about 70 % of lymphocytes if applied in a concentration of 5 mg/ml (MIC results not shown). Systemic application in a bactericidal dosage of 5 mg/ml would therefore be toxic. In AM, however, the preparation is injected subcutaneously, close to the infected areas, e.g. around the paranasal sinuses to treat sinusitis [20]. This application might indeed induce brief antibacterial concentrations in the subcutaneous tissues. It remains an open question, whether this is relevant for the treatment with this medication and weather it improves healing properties according to the concepts of AM.

The antimicrobial effects of Berberis Decoctum D2 were selective. It worked on strains of S. aureus and B. subtilis, but spared the gram-negative strains of E. coli and P. aeruginosa. It would therefore be interesting to examine the effects of Berberis Decoctum D2 on strains of the human gut flora and emphasize on selective effects in terms of a destruction of harmful germs and simultaneous sparing of protective species. For berberine-chloride, obtained from the roots of Coptis japonica, such a positive selectivity on germs of the intestinal flora has already been reported [25]. Furthermore it would be interesting to examine the effects of a topical application of Berberis Decoctum D2, especially in case of colonization with multi-resistant S. aureus strains. For this purpose Berberis Decoctum D2 could be applied in concentrations higher than the determined MIC of 5 mg/ml. Synergistic effects of Berberis Decoctum D2 to antibiotics have not yet been investigated. Such synergistic effects have been reported repeatedly for different plant derived substances [7, 26, 27]. Regarding the fact that Berberis Decoctum D2 has, in contrast to berberine, the status of an approved drug in Germany, investigations using this plant extract would be worthwhile.

Conclusions

Our investigations revealed that Berberis Decoctum D2 has bactericidal effects on Staphylococcus aureus, including methicillin resistant strains, which might be clinically useful in local application.

Abbreviations

(CA)MHB: 

(Cation adjusted) Mueller-Hinton broth

AM: 

Anthroposophical Medicine

ATCC: 

American Type Culture Collection

B.: 

Bacillus (as in B. subtilis)

CFU: 

Colony Forming Unit

CLSI: 

Clinical & Laboratory Standards Institute

DW: 

Distilled water

E.: 

Escherichia (as in E. coli)

log: 

logarithmic

MHB: 

Mueller-Hinton broth

MIC: 

Minimal Inhibitory Concentration

MRSA: 

Methicillin-resistant Staphylococcus aureus

MSSA: 

Methicillin-susceptible Staphylococcus aureus

P.: 

Pseudomonas (as in P. aeruginosa)

S.: 

Staphylococcus (as in S. aureus)

Declarations

Acknowledgements

The authors thank Prof. Dr. med. Daniel Jonas and Marion Buck from the Hygiene and Molecular Laboratory of the Institute of Environmental Health Sciences and Hospital Infection control of the University Medical Center, Freiburg for methodological and laboratory support.

Funding

Co-author Carsten Gründemann is financed from Software AG foundation and DAMUS-DONATA e.V. This funding did not have any influence on design, analysis or reporting of the study.

Availability of data and materials

Original data are available on the server of University Medical Center Freiburg. Availability of materials (bacterial strains and culture media) is mentioned in the methods and materials section of the manuscript.

Authors’ contributions

ER performed the experiments with bacterial strains and wrote a draft of the manuscript. CG performed the experiments with human lymphocytes and critically reviewed the manuscript. IE supported the laboratory work and critically reviewed the manuscript. RH initiated the experiments, supervised the work and critically reviewed the manuscript. All authors read and approved the final manuscript.

Competing interests

The authors declare that they have no conflict of interests concerning this work.

Consent for publication

All authors agree to publish the manuscript in its present form.

Ethics approval and consent to participate

Patients gave their written consent to give blood for research purposes. All experiments conducted on human material were approved by the Local Ethics Committee of the University of Freiburg (482/14).

Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

Authors’ Affiliations

(1)
Center for Complementary Medicine
(2)
Hygiene and Molecular Laboratory, Environmental Health Sciences and Hospital Infection Control, University Medical Center Freiburg
(3)
Center for Complementary Medicine, Department of Environmental Health Sciences, University Medical Center of Freiburg

References

  1. Fauci AS, NIAID Tuberculosis Working Group. Multidrug-resistant and extensively drug-resistant tuberculosis: the National Institute of Allergy and Infectious Diseases Research agenda and recommendations for priority research. J Infect Dis. 2008;197:1493–8.View ArticlePubMedGoogle Scholar
  2. Lowy FD. Staphylococcus aureus infections. N Engl J Med. 1998;339:μ20–32.View ArticleGoogle Scholar
  3. Gold HS, Moellering RC. Antimicrobial-drug resistance. N Engl J Med. 1996;335:1445–53.View ArticlePubMedGoogle Scholar
  4. Spellberg B, Guidos R, Gilbert D, Bradley J, Boucher HW, Scheld WM, Bartlett JG, Edwards J. and Infectious Diseases Society of America. The epidemic of antibiotic-resistant infections: a call to action for the medical community from the Infectious Diseases Society of America. Clin Infect Dis Off Publ Infect Dis Soc Am. 2008;46:155–64.View ArticleGoogle Scholar
  5. Abreu AC, McBain AJ, Simões M. Plants as sources of new antimicrobials and resistance-modifying agents. Nat Prod Rep. 2012;29:1007–21.View ArticlePubMedGoogle Scholar
  6. Ríos JL, Recio MC. Medicinal plants and antimicrobial activity. J Ethnopharmacol. 2005;100:80–4.View ArticlePubMedGoogle Scholar
  7. Han Y, Lee J-H. Berberine synergy with amphotericin B against disseminated candidiasis in mice. Biol Pharm Bull. 2005;28:541–44.View ArticlePubMedGoogle Scholar
  8. Jayaraman P, Sakharkar MK, Lim CS, Tang TH, Sakharkar KR. Activity and interactions of antibiotic and phytochemical combinations against Pseudomonas aeruginosa in vitro. Int J Biol Sci. 2010;6:556–68.View ArticlePubMedPubMed CentralGoogle Scholar
  9. Oluwatuyi M, Kaatz GW, Gibbons S. Antibacterial and resistance modifying activity of Rosmarinus officinalis. Phytochemistry. 2004;65:3249–54.View ArticlePubMedGoogle Scholar
  10. Timmer A, Günther J, Motschall E, Rücker G, Antes G, Kern WV. Pelargonium sidoides extract for treating acute respiratory tract infections. Cochrane Database Syst Rev. 2013;10:CD006323.Google Scholar
  11. Albrecht U, Goos KH, Schneider B. A randomised, double-blind, placebo-controlled trial of a herbal medicinal product containing Tropaeoli majoris herba (Nasturtium) and Armoraciae rusticanae radix (Horseradish) for the prophylactic treatment of patients with chronically recurrent lower urinary tract infections. Curr Med Res Opin. 2007;23:2415–22.View ArticlePubMedGoogle Scholar
  12. Conrad A, Biehler D, Nobis T, Richter H, Engels I, Biehler K, Frank U. Broad spectrum antibacterial activity of a mixture of isothiocyanates from nasturtium (Tropaeoli majoris herba) and horseradish (Armoraciae rusticanae radix). Drug Res. 2013;63:65–8.View ArticleGoogle Scholar
  13. Fintelmann V, Albrecht U, Schmitz G, Schnitker J. Efficacy and safety of a combination herbal medicinal product containing Tropaeoli majoris herba and Armoraciae rusticanae radix for the prophylactic treatment of patients with respiratory tract diseases: a randomised, prospective, double-blind, placebo-controlled phase III trial. Curr Med Res Opin. 2012;28:1799–807.View ArticlePubMedGoogle Scholar
  14. Maranduba CM, De Castro SBR, de Souza GT, Rossato C, da Guia FC, Valente MAS, et al. Intestinal microbiota as modulators of the immune system and neuroimmune system: impact on the host health and homeostasis. J Immunol Res. 2015;2015:931574.View ArticlePubMedPubMed CentralGoogle Scholar
  15. Tilg H. Obesity, metabolic syndrome, and microbiota: multiple interactions. J Clin Gastroenterol. 2010;44 Suppl 1:S16–8.View ArticlePubMedGoogle Scholar
  16. Wilson MT, Hamilos DL. The Nasal and Sinus Microbiome in Health and Disease. Curr Allergy Asthma Rep. 2014;14.Google Scholar
  17. Bosca-Watts MM, Tosca J, Anton R, Mora M, Minguez M, Mora F. Pathogenesis of Crohn’s disease: Bug or no bug. World J Gastrointest Pathophysiol. 2015;6:1–12.View ArticlePubMedPubMed CentralGoogle Scholar
  18. Redinbo MR. The Microbiota, Chemical Symbiosis, and Human Disease. J Mol Biol. 2014;426:3877–91.View ArticlePubMedPubMed CentralGoogle Scholar
  19. Zwiauer J. Anthroposophisch erweiterte Medizin. Ch.Möllmann, Jear: 2013.Google Scholar
  20. Schramm H. Heilmittel der anthroposophischen Medizin Grundlagen Arzneimittelporträts Anwendung. München: Urban&Fischer; 2009.Google Scholar
  21. WALA Arzneimittelverzeichnis (drug register). WALA Heilmittel GmbH Bad Boll, Germany, 34th edition 2013/2014.Google Scholar
  22. Weleda Arzneimittelverzeichnis (drug register), Weleda AG Schwäbisch Gmünd, Germany, 27th edition 2015.Google Scholar
  23. Janssen AM, Scheffer JJ, Baerheim-Svendsen A. Antimicrobial activity of essential oils: a 1976-1986 literature review. Aspects of the test methods. Planta Med. 1986;53:395–98.View ArticleGoogle Scholar
  24. Imanshahidi M, Hosseinzadeh H. Pharmacological and therapeutic effects of Berberis vulgaris and its active constituent, berberine. Phytother Res. 2008;22:999–1012.View ArticlePubMedGoogle Scholar
  25. Ahn YJ, Park SJ, Lee SG, Shin SC, Choi DH. Cordycepin: selective growth inhibitor derived from liquid culture of Cordyceps militaris against Clostridium spp. J Agric Food Chem. 2000;48:2744–48.View ArticlePubMedGoogle Scholar
  26. Diarra MS, Block G, Rempel H, Oomah BD, Harrison J, McCallum J, Boulanger S, Brouillette É, Gattuso M, Malouin F. In vitro and in vivo antibacterial activities of cranberry press cake extracts alone or in combination with β-lactams against Staphylococcus aureus. BMC Complement Altern Med. 2013;13:90.View ArticlePubMedPubMed CentralGoogle Scholar
  27. Freitas E, Aires A, de Santos Rosa EA, Saavedra MJ. Antibacterial activity and synergistic effect between watercress extracts, 2-phenylethyl isothiocyanate and antibiotics against 11 isolates of Escherichia coli from clinical and animal source. Lett Appl Microbiol. 2013;57:266–73.PubMedGoogle Scholar

Copyright

© The Author(s). 2016

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