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Background: Colorants derived from natural sources look set to overtake synthetic colorants in market value as manufacturers continue to meet the rising demand for clean label ingredients -particularly in food applications. Many ascomycetous fungi naturally synthesize and secrete pigments and thus provide readily available additional and/or alternative sources of natural colorants that are independent of agro-climatic conditions. With an appropriately selected fungus; using in particular chemotaxonomy as a guide, the fungal natural colorants could be produced in high yields by using the optimized cultivation technology. This approach could secure efficient production of pigments avoiding use of genetic manipulation.

Exogenous pigments produced by ascomycetous filamentous fungi belonging to the genera Penicillium, Epicoccum, and Monascus, preselected based on chemotaxonomic knowledge, have been extracted and characterized by quantitative colorimetry. The color characteristics of the fungal extracts were compared to water soluble natural colorants derived from sources currently in use. The tested fungal extracts also included some commercially available Monascus colorants. The a values for the fungal extracts were found to be both positive and negative, the b values were found to be positive, while the hue angles of the fungal color extracts ranged from 40 to 110 indicating the color distribution of fungal extracts over the red-orange-yellow region of the CIELAB color space. The fungal extracts exhibited additional color hues in the red spectrum and similar hues in the yellow spectrum as compared to the reference natural colorants. They were also found to be similar or brighter in terms of chroma to some of the reference natural colorants. Principal component analysis was performed to group and distinguish different colors based on the a and b values. The fungal color extracts could be grouped in accordance with the similarity or difference in the color to those of the existing natural colorants. The diversity of colors was not only found among different fungal genera and/or species but also within the same species on changing the media. There was a marked change in the color composition of the extracts resulting in relatively different hues. Our results, thus, indicate that there exists pigment-producing genera of ascomycetous fungi other than Monascus that produce color shades in the red and the yellow spectra in addition or similar to reference colorants. These color shades could add to the color palette of the natural colorants currently in use. In addition, the multivariate approach in distinguishing and classifying the colorants was shown to be a very useful tool in colorimetric comparison of colorants.

Monascus pigments have been used as natural food colorants in Asia for centuries. They are not authorized for use in the European Union and the United States mainly due to the risk of coproduction of the mycotoxin citrinin by Monascus spp. In the present study, we screened for novel producers of Monascus-like pigments from ascomycetous filamentous fungi belonging to Penicillium subgenus Biverticillium that are not reported to produce citrinin or any other known mycotoxins. The screening was carried out using the X-hitting algorithm as a tool to quickly screen through chromatographic sample data files of 22 different Penicillium extracts with 12 Monascus pigment extracts as controls. The algorithm searched for the most similar UV-vis spectra of the metabolites (cross hits) present in the pigment extracts to those of the selected reference metabolites viz. monascin, rubropunctatin, rubropunctamine, and citrinin. The cross hits were then manually identified on the basis of their UV-vis and mass spectra. X-hitting was found to be a good tool in the rapid screening of crude pigment extracts. Monascus pigments were discovered in the extracts of two closely related species of Penicillium that were only distantly related to the genus Monascus. Monascorubrin, xanthomonasin A, and threonine derivatives of rubropunctatin were identified in the extract of Penicillium aculeatum IBT 14263, and monascorubrin was identified in the extract of Penicillium pinophilum IBT 13104. None of the tested Penicillium extracts showed the presence of citrinin. Thus, the present study brought out two novel promising sources of yellow, orange, and purple-red Monascus-like food pigments in the species of Penicillia that do not produce citrinin and opened the door to look for several more new promising sources of natural food colorants in the species of Penicillia.