Compendium of the most significant studies with reference to properties, intake, effects.

Te Welscher, Y. M., Ten Napel, H. H., Balagué, M. M., Souza, C. M., Riezman, H., De Kruijff, B., & Breukink, E. (2008). Natamycin blocks fungal growth by binding specifically to ergosterol without permeabilizing the membrane. Journal of Biological Chemistry, 283(10), 6393-6401.

Abstract. Natamycin is a polyene antibiotic that is commonly used as an antifungal agent because of its broad spectrum of activity and the lack of development of resistance. Other polyene antibiotics, like nystatin and filipin are known to interact with sterols, with some specificity for ergosterol thereby causing leakage of essential components and cell death. The mode of action of natamycin is unknown and is investigated in this study using different in vitro and in vivo approaches. Isothermal titration calorimetry and direct binding studies revealed that natamycin binds specifically to ergosterol present in model membranes. Yeast sterol biosynthetic mutants revealed the importance of the double bonds in the B-ring of ergosterol for the natamycin-ergosterol interaction and the consecutive block of fungal growth. Surprisingly, in strong contrast to nystatin and filipin, natamycin did not change the permeability of the yeast plasma membrane under conditions that growth was blocked. Also, in ergosterol containing model membranes, natamycin did not cause a change in bilayer permeability. This demonstrates that natamycin acts via a novel mode of action and blocks fungal growth by binding specifically to ergosterol.

Resa, C. P. O., Jagus, R. J., & Gerschenson, L. N. (2014). Natamycin efficiency for controlling yeast growth in models systems and on cheese surfaces. Food Control, 35(1), 101-108.

Abstract. Consumers demand for more natural ingredients in processed foods is a result of a requirement for more healthy and safe food with also a need for a balanced and adequate diet. Natamycin is a polyene macrolide antibiotic which is active against yeasts and moulds but not against bacteria, viruses and protozoa. In this study the effectiveness of natamycin delivered by different methods against Saccharomyces cerevisiae, Zygosaccharomyces rouxii and Yarrowia lipolytica using both food models and cheese with natural antimicrobials. It was observed that natamycin concentration and yeast type influenced whether the natamycin effect in tapioca starch films was cidal or inhibitory. This was also observed when the antimicrobial was applied directly to a liquid system for comparison purposes. Bioavailability was not compromised by the polymeric supporting matrix and natamycin efficiency against a S. cerevisiae contamination that preceded antimicrobial application was superior when film action was compared with spraying.

Pedersen, J. C. (1992). Natamycin as a fungicide in agar media. Applied and environmental microbiology, 58(3), 1064-1066.

Abstract. Fungal inhibition in four commonly used agar media was improved by substituting natamycin (pimaricin) for cycloheximide. The recovery of bacteria was not affected by natamycin, whereas fungal contamination from a variety of samples was significantly suppressed. Furthermore, natamycin lacks the occupational health hazards of cycloheximide. Medium-dependent natamycin degradation occurred during the preparation and refrigerated storage of agar plates, but the addition of natamycin at 21.6 μg/ml resulted in effective residual activity.

Meena, M., Prajapati, P., Ravichandran, C. et al. Natamycin: a natural preservative for food applications—a review. Food Sci Biotechnol 30, 1481–1496 (2021). https://doi.org/10.1007/s10068-021-00981-1

Abstract. Natamycin is a natural antimicrobial peptide produced by the strains of Streptomyces natalensis. It effectively acts as an antifungal preservative on various food products like yogurt, khoa, sausages, juices, wines, etc. Additionally, it has been used as a bio preservative and is listed as generally recognized as a safe ingredient for various food applications. In this review, natamycin properties, production methods, toxicity, and application as a natural preservative in different foods are emphasized. This review also focuses on optimal condition and process control required in natamycin production. The mode of action and inhibitory effect of natamycin on yeast and molds inhibition and its formulation and dosage to preserve various food products, coating, and hurdle applications are summarized. Understanding the scientific factors in natamycin's production process, its toxicity, and its efficiency as a preservative will open its practical application in various food products.

Wang D, Shen W, Yuan J, Sun J, Wang M. Advances in the biosynthesis of natamycin and its regulatory mechanisms. Sheng Wu Gong Cheng Xue Bao. 2021 Apr 25;37(4):1107-1119. Chinese. doi: 10.13345/j.cjb.200394. 

Abstract. Natamycin is a polyene macrolide antibiotics with strong and broad spectrum antifungal activity. It not only effectively inhibits the growth and reproduction of fungi, but also prevents the formation of some mycotoxins. Consequently, it has been approved for use as an antifungal food preservative in most countries, and is also widely used in agriculture and healthcare. Streptomyces natalensis and Streptomyces chatanoogensis are the main producers of natamycin. This review summarizes the biosynthesis and regulatory mechanism of natamycin, as well as the strategies for improving natamycin production. Moreover, the future perspectives on natamycin research are discussed.

Burkin MA, Moshcheva AG, Galvidis IA. Immunoassay for Natamycin Trace Screening: Bread, Wine and Other Edibles Analysis. Biosensors (Basel). 2022 Jul 6;12(7):493. doi: 10.3390/bios12070493. 

Abstract. The antifungal drug natamycin (NAT) is widely used in medicine and in the food industry as preservative E235 for a wide variety of foods. The risk of the development of resistance to NAT and its spread in relation to other polyene antibiotics is fraught with the emergence of incurable infections. This work is devoted to the development of an immunoassay to investigate the prevalence of NAT use for food preservation. Two immunogen designs based on tetanus toxoid, conjugated to NAT through different sites of hapten molecules, were compared in antibody generation. Assay formats using heterologous coating antigens were superior for both antibodies. The ELISA variant demonstrated the highest sensitivity (IC50 = 0.12 ng/mL), and a limit of detection of 0.02 ng/mL was selected for NAT determination. The optimized extraction procedure provided a recovery rate of 72–106% for various food matrixes with variations below 12%. Cyclodextrins, as well as NAT–cyclodextrin complex formulations, showed no interference with the quantification of NAT. One hundred and six food product brands, including baked goods, wines, beers, drinks, sauces, and yogurts, were tested to assess the prevalence of the undeclared use of NAT as a preservative. The screening examination revealed three positive yogurts with an undeclared NAT incorporation of 1.1–9.3 mg/kg.

Chen D, Förster H, Nguyen K, Adaskaveg JE. Organic Acid Sanitizers for Natamycin and Other Fungicides in Recirculating Application Systems for Citrus Postharvest Decay Management. Plant Dis. 2021 Oct;105(10):2907-2913. doi: 10.1094/PDIS-01-21-0227-RE. Epub 2021 Nov 1. 

Abstract. Natamycin is a new postharvest biofungicide for citrus and some other fruit crops in the United States that can be effectively used in recycling drench or flooder treatments. These applications necessitate sanitation of the fungicide solution to ensure that it remains free from contamination by bacteria that are potentially human pathogens. During in vitro experiments, heated (48°C) citric acid (1,100 or 2,200 μg/ml) amended with sodium dodecylbenzenesulfonate (SDBS) (60 or 120 μg/ml, respectively) significantly reduced the viability of a nonpathogenic strain of Escherichia coli in natamycin solutions by >5 log10 compared with the control. During laboratory studies with Penicillium digitatum-inoculated lemon fruit, 1,000 μg/ml of natamycin mixed with 1,000 μg/ml of lactic acid or citric acid and with or without SDBS (55 μg/ml) effectively and significantly reduced green mold. Natamycin mixed with lactic acid at ≥2,000 μg/ml, however, caused fruit injury, resulting in browning and rind pitting. Natamycin was incompatible with peroxyacetic acid, resulting in reduced efficacy against green mold. Sodium hypochlorite mixed with natamycin lost its toxicity to E. coli; however, the performance of natamycin was not affected. With heated (average 49°C) drench treatments on an experimental packing line, natamycin (1,000 μg/ml), fludioxonil (300 μg/ml), or azoxystrobin (300 μg/ml) mixed with citric acid (1,000 μg/ml) and SDBS (55 μg/ml) were effective against green mold without fruit injury. At a pH between 3.6 and 3.8, citric acid-SDBS significantly reduced the viability of E. coli by approximately 4 log10 in mixtures with fludioxonil or azoxystrobin, but not with natamycin. However, natamycin at 1,000 μg/ml mixed with 2,000 μg/ml of citric acid and SDBS (55 μg/ml) significantly reduced E. coli counts by >4 log10 within 4 min when the pH was maintained between 3.0 and 3.3, and the efficacy of the fungicide was retained. The use of citric acid with a surfactant can be a viable alternative sanitation method for natamycin in citrus packinghouses utilizing heated recirculating fungicide systems.

Vasquez JL, Lai Y, Annamalai T, Jiang Z, Zhang M, Lei R, Zhang Z, Liu Y, Tse-Dinh YC, Agoulnik IU. Inhibition of base excision repair by natamycin suppresses prostate cancer cell proliferation. Biochimie. 2020 Jan;168:241-250. doi: 10.1016/j.biochi.2019.11.008. 

Abstract. Prostate cancer (PCa) progression is characterized by increased expression and transcriptional activity of the androgen receptor (AR). In the advanced stages of prostate cancer, AR significantly upregulates the expression of genes involved in DNA repair. Upregulation of expression for base excision repair (BER) related genes is associated with poor patient survival. Thus, inhibition of the BER pathway may prove to be an effective therapy for prostate cancer. Using a high throughput BER capacity screening assay, we sought to identify BER inhibitors that can synergize with castration therapy. An FDA-approved drug library was screened to identify inhibitors of BER using a fluorescence-based assay suitable for HTS. A gel-based secondary assay confirmed the reduction of BER capacity by compounds identified in the primary screen. Five compounds were then selected for further testing in the independently derived, androgen-dependent prostate cancer cell lines, LNCaP and LAPC4, and in the nonmalignant prostate derived cell lines PNT1A and RWPE1. Further analysis led to the identification of a lead compound, natamycin, as an effective inhibitor of key BER enzymes DNA polymerase β (pol β) and DNA Ligase I (LIG I). Natamycin significantly inhibited proliferation of PCa cells in an androgen depleted environment at 1 μM concentration, however, growth inhibition did not occur with nonmalignant prostate cell lines, suggesting that BER inhibition may improve efficacy of the castration therapies.

Koontz JL, Marcy JE, Barbeau WE, Duncan SE. Stability of natamycin and its cyclodextrin inclusion complexes in aqueous solution. J Agric Food Chem. 2003 Nov 19;51(24):7111-4. doi: 10.1021/jf030333q. 

Abstract. Aqueous solutions of natamycin and its beta-cyclodextrin (beta-CD), hydroxypropyl beta-cyclodextrin, and gamma-cyclodextrin (gamma-CD) inclusion complexes were completely degraded after 24 h of exposure to 1000 lx fluorescent lighting at 4 degrees C. After 14 days of storage in darkness at 4 degrees C, 92.2% of natamycin remained in active form. The natamycin:beta-CD complex and natamycin:gamma-CD complex were significantly more stable (p < 0.05) than natamycin in its free state in aqueous solutions stored in darkness at 4 degrees C. Clear poly(ethylene terephthalate) packaging with a UV light absorber allowed 85.0% of natamycin to remain after 14 days of storage under 1000 lx fluorescent lighting at 4 degrees C. Natamycin:cyclodextrin complexes can be dissociated for analysis in methanol/water/acetic acid, 60:40:5, v/v/v. Natamycin and its complexes in dissociated form were quantified by reverse phase HPLC with detection by photodiode array at 304 nm.