Advertisment ACS-IndiaSymposium
 
Journal of Pharmaceutical Negative Results
  Print this page Email this page Small font sizeDefault font sizeIncrease font size 
Search Article 
  
Advanced search 
 Home | About us | Editorial board | Search | Ahead of print | Current issue | Archives | Submit article | Instructions | Subscribe | Contacts  
 


 
  Table of Contents  
ORIGINAL ARTICLE
Year : 2016  |  Volume : 7  |  Issue : 1  |  Page : 16-20  

Insignificant antifungal activity of plant extracts on Malassezia furfur


Department of Microbiology and Biotechnology, Jnanabharathi Campus, Bangalore University, Bengaluru, Karnataka, India

Date of Web Publication19-Feb-2016

Correspondence Address:
Sharmila Thirumale
Department of Microbiology and Biotechnology, Jnanabharathi Campus, Bangalore University, Bengaluru - 560 056, Karnataka
India
Login to access the Email id

Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0976-9234.177055

Rights and Permissions
   Abstract 

Objective: The aim of present study was to investigate the antimalassezial potential of the selected plant extracts known to possess medicinal values. Materials and Methods: The crude extracts of five botanicals were evaluated against Malassezia furfur. The dried and pulverized plant material was extracted with water and also successively extracted with hexane, chloroform, and methanol, using the Soxhlet apparatus. The antimicrobial activity assay was done by disk diffusion and 96-well plate microdilution method. Results: The results point toward the fact that all the tested extracts extracted in solvents of different polarities exhibited no antifungal activity. Conclusion: The tested extracts did not have, or had too little, antifungal activity to be used as a promising source of novel antimicrobial substances against M. furfur.

Keywords: Disk diffusion, Malassezia furfur, microdilution, plant extracts


How to cite this article:
Lalitha SG, Ramachandrappa LT, Krishnamurthy S, Basavaraju J, Thirumale S. Insignificant antifungal activity of plant extracts on Malassezia furfur. J Pharm Negative Results 2016;7:16-20

How to cite this URL:
Lalitha SG, Ramachandrappa LT, Krishnamurthy S, Basavaraju J, Thirumale S. Insignificant antifungal activity of plant extracts on Malassezia furfur. J Pharm Negative Results [serial online] 2016 [cited 2017 Jul 25];7:16-20. Available from: http://www.pnrjournal.com/text.asp?2016/7/1/16/177055


   Introduction Top


Pityriasis capitis, commonly known as dandruff, is a most common physiological condition causing desquamation of the skin surface due to the imbalance caused to the stratum corneum. [1] Seborrheic dermatitis is yet another condition that is closely related to dandruff, characterized by highly pruritic chronic inflammatory lesions on the head. [2] These pathologic conditions are considered to be related to three etiologic factors: Malassezia yeasts, secretions of the sebaceous glands, and susceptibility to infections in individuals. It has been generally accepted and through the literature inferred by studies, indicating a strong link to said conditions caused by Malassezia yeasts, especially Malassezia furfur and, established later, its two species forms Malassezia globosa and Malassezia restricta. [3] Other conditions associated with this species are pityriasis versicolor, folliculitis, atopic dermatitis, psoriasis, and confluent and reticulate papillomatosis. [4]

The hydrolytic enzyme lipase secreted by Malassezia species residing on the surface of the scalp and in the follicular infundibulum cleaves sebaceous triglycerides into free fatty acids and glycerol into the extracellular milieu. The Malassezia consumes necessary saturated fatty acids required for their proliferation and leave behind an increased quantity of irritating unsaturated fatty acids. The unsaturated fatty acids penetrate the stratum corneum, and in susceptible individuals, due to the nonuniform structure, breach the skin barrier function. This barrier breach induces an irritation response leading to scalp hyperproliferation, ultimately leading to pathological conditions such as dandruff and seborrheic dermatitis. [5]

Treatment strategies include synthetic drug-based formulations of ketoconazole, zinc pyrithione, selenium sulfide, ciclopirox, etc., which reduce visible symptoms of flaking and restore the normalcy of the skin. [6] In fact, most medicated formulations carry risks of side effects such as dryness of hair/skin, associated cytostasis, and eczema, and also frequent reoccurrence made therapy costlier. So, it is imperative to search for drugs that are safe, cost-effective, and eco-friendly. Thus, there was a call to return to nature and botanical remedies became a part of the green revolution.

The term "botanicals" refers to the preparations derived from herbs, spices, roots, stems, and other materials of botanical origin. [7] Plants that are rich in a wide array of antifungal secondary metabolites are gaining importance as a primary source of commercial medicines and drug leads, and have been used for the treatment of skin disorders for centuries. [8] Herbal drug technology has picked up strongly in terms of the extraction and characterization of active compounds and also the processing of herbs into medicine, considering the fact that modern medicine is not capable of providing a "cure all" solution for the pathologies mentioned above. Herbal therapy is considered as a therapeutic alternative, as a safer choice than synthetic medicines, or sometimes even as the only successful therapeutic way left to treat these disorders. [9] It is important to document their uses and their distinct modes of action because such information can help in obtaining maximum benefits from these resources, and this increases the possibility of their safe and efficient use in the future. Traditionally, many botanicals tend to possess anti-dandruff activity, but scientific validation is needed to justify their role in this activity. [10]

In this present scenario, a developing country such as India must focus on documentation of medicinal plants and development of herbal formulations to combat skin infections. The plants used in the present investigation, such as Punica granatum L. (Lythraceae), Illicium verum Hook (Schisandraceae), Nyctanthes arbor-tristis L. (Oleaceae), Thespesia populnea L. (Malvaceae), and Piper betle L. (Piperaceae) were used to treat cutaneous infections such as psoriasis, tinea infection, and eczema. [11],[12],[13],[14],[15] Thus, the present study was conducted with particular emphasis on aforementioned medicinal plants used traditionally against the dermatologically prevalent yeast, M. furfur.


   Materials And Methods Top


Collection and maintenance of test microorganism

Malassezia furfur (strain no. 1374) was obtained from the Microbial Type Culture Collection and Genebank (MTCC), Chandigarh, India. The culture was maintained in Leeming-Notman (LN) agar, modified Emmons agar medium, and Sabouraud's dextrose agar (SDA) supplemented with milk. LN agar medium was used for carrying out antimicrobial studies. [16]

Plant materials

Plant materials used were selected based on their topical therapeutic properties according to the Ayurvedic system of medicine. [17] Authenticated plants were collected from Gandhi Krishi Vignan Kendra, Bengaluru, Karnataka, India. The voucher specimens of all the plants were maintained at the Department of Microbiology and Biotechnology, Bangalore University, Bengaluru.

Preparation of plant extracts

Fresh plant materials were blot-dried and macerated with sterile, distilled water in a blender (1:3 w/v) for 10 min. The macerate was first filtered through double-layered muslin cloth and centrifuged at 8000 rpm for 10 min. The supernatant was filtered through Whatman no. 1 filter paper and sterilized at 120°C for 30 min, and this served as the mother extract. All the extracts were concentrated in water bath and stored at 4°C for further use. [18]

For solvent extraction, 60 g of shade-dried plant material was pulverized, placed to fill the thimble of a Soxhlet extractor, and extracted with 250 mL of different solvents (sequentially using hexane, chloroform, and methanol). The process of extraction continued until the solvent in the siphon of the extractor become colorless. The extracts were concentrated using rotary evaporator and the dried extract was kept in the refrigerator at 4°C until further use. [19],[20]

Antimicrobial susceptibility test

As an initial step toward confirming preliminary antifungal activity, the disk diffusion assay was done to screen the antifungal activity of all the extracts. Inoculum suspension previously adjusted to 0.5 McFarland standard was swabbed evenly over the sterile  LN agar medium plates set for the disk diffusion assay. Sterile 6 mm diameter disks were placed equidistantly round the margin of the  Petri dish More Details. Of the test sample drug, 10 μL (25 mg mL -1 ) was dispensed on the disks. The inoculated plates were incubated at 32 ± 2°C for 48 h. Each test was performed in triplicate. After the incubation period, the antimicrobial activity was recorded as the width (diameter of inhibition zone plus diameter of the disk) of the inhibition zone. Minimum inhibitory concentrations (MICs) were determined in liquid culture by a 96-well microtiter plate assay recommended by the National Committee for Clinical Laboratory Standards (NCCLS) with minor modification. [21] Further, minimal yeast-cidal concentration (MYC) has been evaluated by streaking all treated samples on agar medium and assessing the concentration that inhibits complete yeast growth after incubation. [22]

Statistical analysis

Statistical analysis was performed using  SPSS software version 16.0 (Armonk, NY, IBM Corporation). The results were expressed as mean ± (standard deviation) SD (n = 3). Univariate analysis followed by Tukey's honestly significant difference (HSD) post hoc test was applied for statistical analysis, with the level of significance set at P < 0.05. Results with P < 0.05 were considered to be statistically significant.


   Results Top


The study examined the antimalassezial activity of selected extracts on the dermatologically prevalent yeast M. furfur using the in vitro technique, which is reliable, sensitive, and widely used. The hexane, chloroform, methanol, and water extracts of the selected plants yielded 0.48-18.15% of their dry weight. The yield was not constant with any particular solvent or plant part. Antimicrobial activity assessed in terms of zone of inhibition in mm of the ten extracts of five plants with different solvents and tested against M. furfur was recorded and can be seen in [Table 1]. In the current study, a total of ten extracts of selected plants were tested for their antifungal activity, of which all the extracts showed no activity or very little activity against M. furfur at the tested concentration of 25 mg mL -1 . Nevertheless, the highest antimicrobial activities were recorded for N. arbor-tristis in methanol and hexane, with zones of inhibition of 8.33 ± 0.57 mm and 7.73 ± 0.46 mm, respectively. In case of the hexane extract of I. verum, the zone of inhibition observed was 7.66 ± 0.57 mm and the methanol extract of P. granatum showed the inhibition zone of 7.33 ± 0.57 mm, thus manifesting limited antifungal potential against M. furfur. The extracts that showed activity in disk diffusion assay were evaluated for their MIC and MYC values by the 96-well broth dilution method and are tabulated in [Table 2]. The MIC value of the extracts that showed confined activity was 25 mg mL -1 , while the MYC value was 50 mg/mL -1 . The remaining extracts exhibited no activity.
Table 1: Antifungal activity of different plant extracts of Malassezia furfur


Click here to view
Table 2: MIC and MYC of crude extracts


Click here to view



   Discussion Top


Antimicrobials of plant origin are in great demand due to their widespread biological activities providing a source for the discovery of many types of effective bioactive compounds. Of these, very few successful drugs are now available for the treatment of fungal infections, especially for superficial mycoses. [23] Ever since the importance of the distribution of pharmacologically active principles in higher plants was understood and acknowledged, the importance of such plant-derived medicines in modern therapeutic practice has paved the way for the development of new drug leads that are safe, cost-effective, and eco-friendly. Taking this into account, the present study was conducted to assess the antifungal potential of medicinal plants, particularly to target the dandruff-causing yeast M. furfur. The present findings were in accordance to the reports of Dikshit et al., Lee et al., and Hayouni et al. [24],[25],[26] However, the studied extracts had relatively high MIC and MYC values and thus cannot be considered a good antifungal agent.


   Conclusion Top


To conclude, it is imperative to emphasize the significance of more comprehensive studies that will detect new lead molecules underlying the action of crude extracts that allow for new discoveries. Essentially, the present study, which establishes the lack of significant antimalassezial activity, removes the need for any repetitive study with these extracts in the future.

Acknowledgment

The authors would like to extend their sincere thanks and appreciation to the Department of Microbiology and Biotechnology, Bangalore University for providing adequate lab facilities and the Himalaya Drug Co, Nelamangala, Bangalore for their magnanimous gesture in supporting research and providing the required plant extracts needed for the study.

Financial support and sponsorship

UGC - BSR meritorious fellowship for Sreelatha G.L.

Conflicts of interest

There are no conflicts of interest.

 
   References Top

1.
Turner GA, Hoptroff M, Harding CR. Stratum corneum dysfunction in dandruff. Int J Cosmet Sci 2012;34:298-306.  Back to cited text no. 1
    
2.
Hay RJ. Malassezia, dandruff and seborrhoeic dermatitis: An overview. Br J Dermatol 2011;165(Suppl 2):2-8.  Back to cited text no. 2
    
3.
DeAngelis YM, Gemmer CM, Kaczvinsky JR, Kenneally DC, Schwartz JR, Dawson TL Jr. Three etiologic facets of dandruff and seborrheic dermatitis: Malassezia fungi, sebaceous lipids, and individual sensitivity. J Investig Dermatol Symp Proc 2005;10:295-7.  Back to cited text no. 3
    
4.
Gupta AK, Batra R, Bluhm R, Boekhout T, Dawson TL Jr. Skin diseases associated with Malassezia species. J Am Acad Dermatol 2004;51:785-98.  Back to cited text no. 4
    
5.
Dawson TL Jr. Malassezia globosa and restricta: Breakthrough understanding of the etiology and treatment of dandruff and seborrheic dermatitis through whole-genome analysis. J Investig Dermatol Symp Proc 2007;12:15-9.  Back to cited text no. 5
    
6.
Pierard GE, Arrese JE, Pierard-Franchimont C, DE Doncker P. Prolonged effects of antidandruff shampoos - time to recurrence of Malassezia ovalis colonization of skin. Int J Cosmetic Sci 1997;19:111-7.  Back to cited text no. 6
    
7.
Reuter J, Merfort I, Schempp CM. Botanicals in dermatology: An evidence-based review. Am J Clin Dermatol 2010;11:247-67.  Back to cited text no. 7
    
8.
Ali-Shtayeh MS, Abu Ghdeib SI. Antifungal activity of plant extracts against dermatophytes. Mycoses 1999;42:665-72.  Back to cited text no. 8
    
9.
Joshi K, Chavan P, Warude D, Patwardhan B. Molecular markers in herbal drug technology. Curr Sci 2004;87:159-65.  Back to cited text no. 9
    
10.
Cavero RY, Akerreta S, Calvo MI. Medicinal plants used for dermatological affections in Navarra and their pharmacological validation. J Ethnopharmacol 2013;149:533-42.  Back to cited text no. 10
    
11.
Jurenka JS. Therapeutic applications of pomegranate (Punica granatum L.): A review. Altern Med Rev 2008;13:128-44.  Back to cited text no. 11
    
12.
Matan N, Matan N. Antifungal activities of anise oil, lime oil, and tangerine oil against molds on rubberwood (Hevea brasiliensis). Int Biodeter Biodegr 2008;62:75-8.  Back to cited text no. 12
    
13.
Rani C, Chawla S, Mangal M, Mangal AK, Khagla S, Dhawan AK. Nyctanthes arbor-tristis Linn. (Night Jasmine): A sacred ornamental plant with immense medicinal potentials. Indian J Tradit Knowl 2012;11:427-35.  Back to cited text no. 13
    
14.
Kumar VP, Chauhan NS, Padh H, Rajani M. Search for antibacterial and antifungal agents from selected Indian medicinal plants. J Ethnopharmacol 2006;107:182-8.  Back to cited text no. 14
    
15.
Trakranrungsie N, Chatchawanchonteera A, Khunkitti W. Ethnoveterinary study for antidermatophytic activity of Piper betle, Alpinia galanga and Allium ascalonicum extracts in vitro. Res Vet Sci 2008;84:80-4.  Back to cited text no. 15
    
16.
Leeming JP, Notman FH. Improved methods for isolation and enumeration of Malassezia furfur from human skin. J Clin Microbiol 1987;25:2017-9.  Back to cited text no. 16
    
17.
Khare CP. Indian Medicinal Plants: An Illustrated Dictionary.  India: Springer; 2007. p. 325-659.  Back to cited text no. 17
    
18.
Mohana DC, Raveesha KA. Anti-fungal evaluation of some plant extracts against some plant pathogenic field and storage fungi. J Agr Sci Tech 2007;4:119-37.  Back to cited text no. 18
    
19.
Sreelatha GL, Babu UV, Sharath Kumar LM, Soumya K, Sharmila T. Investigation on biochemical characterisation and in vitro antifungal efficacy of plant extracts on Malassezia furfur. Int J Pharm Bio Sci 2015;6:1027-41.  Back to cited text no. 19
    
20.
Yadav RN, Agarwala M. Phytochemical analysis of some medicinal plants. J Phytol 2011;3:10-4.  Back to cited text no. 20
    
21.
National Committee for Clinical Laboratory Standards. Reference Method for Broth Dilution Antifungal Susceptibility Testing of Yeasts. Approved Standard. Document M-27A. National Committee for Clinical Laboratory Standards. Wayne, Pennsylvania: National Committee for Clinical Laboratory Standards; 1997.   Back to cited text no. 21
    
22.
Lakshmeesha TR, Sateesh MK, Prasad BD, Sharma SC, Kavyashree D, Chandrasekhar M, et al. Reactivity of crystalline ZnO superstructures against fungi and bacterial pathogens: Synthesized using Nerium oleander leaf extract. Cryst Growth Des 2014;14:4068-79.  Back to cited text no. 22
    
23.
Afolayan AJ, Grierson DS, Mbeng WO. Ethnobotanical survey of medicinal plants used in the management of skin disorders among the Xhosa communities of the Amathole District, Eastern Cape, South Africa. J Ethnopharmacol 2014;153:220-32.  Back to cited text no. 23
    
24.
Dikshit A, Tiwari AK, Mishra RK, Kamran A, Pandey A, Kumar A, et al. Botanicals for the management of dandruff. Med Plants 2012;4:55-64.  Back to cited text no. 24
    
25.
Lee JH, Lee JS. Chemical composition and antifungal activity of plant essential oils against Malassezia furfur. Kor J Microbiol Biotechnol 2010;38:315-21.  Back to cited text no. 25
    
26.
Hayouni EA, Miled K, Boubaker S, Bellasfar Z, Abedrabba M, Iwaski H, et al. Hydroalcoholic extract based-ointment from Punica granatum L. peels with enhanced in vivo healing potential on dermal wounds. Phytomedicine 2011;18:976-84.  Back to cited text no. 26
    



 
 
    Tables

  [Table 1], [Table 2]



 

Top
  
 
  Search
 
    Similar in PUBMED
   Search Pubmed for
   Search in Google Scholar for
 Related articles
    Access Statistics
    Email Alert *
    Add to My List *
* Registration required (free)  

 
  In this article
    Abstract
   Introduction
    Materials And Me...
   Results
   Discussion
   Conclusion
    References
    Article Tables

 Article Access Statistics
    Viewed2337    
    Printed22    
    Emailed0    
    PDF Downloaded240    
    Comments [Add]    

Recommend this journal