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Broad-Spectrum Anti-Fungal Activity of Pyrithione Zinc and its Effect on the Causes of Dandruff and Associated Itch

Yvonne DeAngelis, CVT, Erin MacDonald, BS, Kathy Kramp, BS, Rob Bacon, BS, David J. Kaufman, BS, James R. Schwartz, PhD, and Thomas L. Dawson, Jr, PhD

The Procter & Gamble Company, Cincinnati, Ohio


Application of new molecular and biochemical tools has increased our understanding of the organisms, mechanisms, and treatments of dandruff and seborrheic dermatitis.

Malassezia has long been a suspected cause, implicated by its presence and lipophylic

nature1. However, lack of correlation between numbers of Malassezia and dandruff has been perplexing2.

We have previously shown M. globosa and M. restricta as the causal organisms.

Here we show:

  • Increased numbers of M. restricta in dandruff
  • in vivo reduction of M. restricta, M. globosa and multiple fungi during treatment
  • in vitro potency of monographed anti-dandruff actives against M. globosa and
  • correlate this reduction with reduction of scalp flaking and itch.


  • Quantify the causal microorganism(s) in dandruff and seborrheic dermatitis.
  • Test the in vitro potency of monographed anti-dandruff actives against the causal


  • Quantify the in vivo efficacy of Pyrithione Zinc shampoo against the causal species.
  • Correlate reduction in Malassezia to reduction in dandruff and seborrheic dermatitis

  flaking and associated itch.


qPCR (quantitative Polymerase Chain Reaction)

Quantitative PCR was performed on samples prepared as for tFLP3. Panfungal primers were targeted to the 5.8s and 28s ribosomal genes4. Species specific primers were designed in house. Standard curves were constructed against cloned ribosomal gene sequences. Standard

curves: M. restricta from M. restricta 7877; M. globosa from a combination of 7966, 7874, and 7990; Panfungal from a combination of M. globosa 7966 and M. restricta 7877. All standard strains were obtained from the CBS, The Netherlands.


Malassezia MIC testing.

Minimum Inhibitory Concentrations were calculated by standard plate dilution methodology, in mLNA growth media. Actives were tested at 2x dilutions. Growth conditions for M. restricta do not permit this type of analysis.


Assessment of Scalp Itch.

Itch severity was measured by subject self-perception. The self-perception scale was 0-9 with 9 as severe itch. Data presented are least squares means as difference from baseline.


Assessment of Flaking.

Dandruff was assessed with the adherent scalp flaking scale (ASFS)5. A numerical grade was assigned (0 to 10, with 10 the most severe) at 8 sites, yielding a composite score ranging from 0 to 80. Panelists used a non-dandruff shampoo for three weeks prior to baseline. At baseline,

those with ASFS of ≥24 were randomly assigned to either a commercial small-particle platelet PtZ or a placebo shampoo.


Malassezia Numbers

dandruff versus non-dandruff


In vitro MIC Values for Malassezia


In vivo Malassezia

Reduction post-treatment with PtZ


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Figure 1. All fungi and Malassezia restricta are found at higher numbers in dandruff subjects.


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Figure 2. Pyrithione zinc is a potent antifungal monographed antidandruff active as measured by MIC (experiment cannot be conducted with M. restricta).

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Figure 3. Treatment with a 2.5 platelet Pyrithione zinc shampoo efficiently reduces M. restricta, M. globosa, and all fungi.

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Figure 4. Treatment with 2.5 platelet Pyrithione zinc shampoo visually improves dandruff flaking.

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Figure 5. Treatment with Pyrithione zinc shampoo efficiently removes dandruff flakes.

(Insert Itch Severity graph here)
Figure 6. Treatment with 2.5platelet Pyrithione zinc shampoo relieves the itch associated with dandruff flaking.


Malassezia restricta and/or Malassezia globosa are the most likely causal micro-organisms in dandruff and seborrheic dermatitis. The number of M. restricta and all fungi are elevated in dandruff populations. Treatment with a 2.5 platelet-size Pyrithione zinc shampoo effectively removes M. restricta, M. globosa, and general fungi. Pyrithione zinc is the most potent monographed anti-dandruff active against the microbial species which cause dandruff. Removal of the microbial species and thereby the irritating byproducts which

cause dandruff reduces both dandruff flaking and associated itch.



1. S. Shuster. The aetiology of dandruff and the mode of action of therapeutic agents, Brit. J. Dermatol. 1984; 111:235-42.
2. GE Pierard, JE Arrese, C. Pierard-Franchimont. Prolonged effects of antidandruff shampoos - time to recurrence of Malassezia ovalis colonization of skin, Intl. J. Cosm. Sci. 1997; 19:111-7.
3. C. Gemmer, , Y. DeAngelis, B. Theelen, T. Boekhout, and T.L. Dawson, Jr A fast, non-invasive method for identification and speciation of Malassezia, and it application to dandruff biology, J. Clin. Micro. 2002, 40 (9): 3350-3357.
4. T White, S Lee, J Taylor. 1990. Amplification and direct sequencing of fungal ribosomal rna genes for phylogenetics p314-322., In M. Innus, D.H Gelfand, J.J. Snisky, and T.J. White (eds) PCR Protocols, A guide to methods and applications, Academic Press, In., San Diego, Ca.
5. Van Abbe, N.J. The investigation of dandruff, J. Soc. Cosmetic
Chem., 1964. 15: p. 609-630.

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