Novel milk-clotting enzyme from sour orange flowers (Citrus aurantium L.) as a coagulant in Iranian white cheese

  • Elham Nasiri
  • Javad HesariEmail author
  • Seyed Shahram Shekarforoush
  • Sodeif Azadmard Damirchi
  • Sabrina Gensberger-Reigl
  • Monika Pischetsrieder
Original Paper


The aim of this work was to evaluate the partially purified protease of sour orange flower extract (OFR) by determining the protein profile and milk-clotting activity as well as chemical and sensory characteristics of cheese throughout 45 days of ripening. Three rennets were used to produce cheeses, (1) the sour orange flower rennet at a level of 0.2 g/L milk (OFR), (2) standard calf rennet at a level of 0.02 g/L milk (CR), and (3) OFR (0.1 g/L)/CR (0.01 g/L). OFR cheese acidity was higher compared to that of the calf rennet (CR) and OFR/CR (50:50) samples. Significantly (p < 0.05) higher proteolytic activity was observed in OFR cheese through measuring the soluble nitrogen (SN), non-protein nitrogen (NPN), and total free amino acids (FAA) content. Protein profiles of cheeses determined with SDS-PAGE also confirmed higher levels of casein hydrolysis in OFR cheese. Reversed-phase high-performance liquid chromatography revealed the noticeable qualitative and quantitative differences between peptide profiles of all treatments. Higher casein degradation and production of peptide plus amino acids in OFR and OFR/CR cheeses suggested faster and more intensive ripening. The flavor and texture acceptance scores were reduced on the last day of storage in OFR cheese, while OFR/CR cheese received the highest score for flavor. In conclusion, the blend of OFR/CR may be regarded as an alternative for calf rennet in white brined cheese production which accelerates ripening process and causes development of a unique flavor in the final product.


Sour orange flowers Protease White brined cheese Ripening 



We thank Dr. Amir Fattahi ( for statistical analysis and his academic advices. Some of the data included are part of the doctoral thesis of Elham Nasiri. The authors received no financial support for the research, authorship, and/or publication of this article.


The authors received no financial support for the research, authorship, and/or publication of this article.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Compliance with ethics requirements

This article does not contain any studies with human and animal subjects.


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Elham Nasiri
    • 1
    • 3
  • Javad Hesari
    • 1
    Email author
  • Seyed Shahram Shekarforoush
    • 2
  • Sodeif Azadmard Damirchi
    • 1
  • Sabrina Gensberger-Reigl
    • 3
  • Monika Pischetsrieder
    • 3
  1. 1.Department of Food Science and Technology, Faculty of AgricultureTabriz UniversityTabrizIran
  2. 2.Department of Food Hygiene and Public Health, School of Veterinary MedicineShiraz UniversityShirazIran
  3. 3.Department of Chemistry and Pharmacy, Food ChemistryFriedrich-Alexander Universität Erlangen-Nürnberg (FAU)ErlangenGermany

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