**1. Introduction**

Honey bees (*Apis mellifera* L.) are primary pollinators with an important role in ecosystem conservation [1], offering many services and products, such as honey. Honey through the centuries has always been a vital food for humans, with many health properties [2,3]. The Mediterranean region, specifically Greece pronounces a set of several common and rare monofloral honeys in international markets [4]. Additionally, nowadays few rare honeys, like heather, have become increasingly well-known for their special characteristics and have received several awards in national and international food quality or taste competitions [5]. The term "heather" is used for plant species belonging to *Erica* and *Calluna* genera. However, this term is used to describe the honey produced from *Calluna vulgaris* (L.) Hull and not from other Ericaceae botanical sources [6]. In relation to honey from common species, including *Erica arborea* L., *Erica carnea* L., and *Erica cinerea* L., the given names are "Tree heath", "Spring heather", and "Bell heather", respectively [6].

**Citation:** Xagoraris, M.;

Chrysoulaki, F.; Revelou, P.-K.; Alissandrakis, E.; Tarantilis, P.A.; Pappas, C.S. Unifloral Autumn Heather Honey from Indigenous Greek *Erica manipuliflora* Salisb.: SPME/GC-MS Characterization of the Volatile Fraction and Optimization of the Isolation Parameters. *Foods* **2021**, *10*, 2487. https://doi.org/10.3390/ foods10102487

Academic Editors: Olga Escuredo and M. Carmen Seijo

Received: 28 September 2021 Accepted: 15 October 2021 Published: 17 October 2021

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**Copyright:** © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

Greek flora includes four Ericaceae nectar-secretion bee plants. Two of them are spring flowering species including *Erica arborea* L., and *Rhododendron* sp. while the other two (*Erica manipuliflora* Salisb. and *Arbutus unedo* L.) bloom in autumn. *Erica manipuliflora* is indigenous in Greece and is known as "autumn heather", while the traditional term used is "sousoura". However, honey from *E. manipuliflora* should not be confused with other heather honeys produced during autumn, including from *C. vulgaris*, and *Erica multiflora* L. Monofloral autumn heather honey can be quite easily produced [7], as its collection period does not coincide with the blooming of other bee plants, with the exception of *A. unedo* honey, which blooms in late autumn and its blooming period follows that of *E. manipuliflora*.

Greek autumn heather honey is well-known for its extraordinary aroma profile, characterized by perfume reminiscent "caramel" notes, which is worth studying since data for this honey variety are scarce. In the last twenty years, just two studies [8,9] have dealt with the volatile fraction of *E. manipuliflora* honey. However, there are numerous studies concerning heather honey [7,10–17]. As shown in a review study [18], the above studies refer to different botanical species, geographical origin, number of samples, isolation, and analysis procedures.

The volatile isolation method is usually followed by gas chromatography-mass spectrometry (GC-MS), and plays a significant role in the qualitative and quantitative determination of volatiles. Solid-phase microextraction (SPME) as a volatile fraction extraction methodology constitutes a simple procedure with no pre-treatment of samples and environmentally friendly solvents [19]. The main factors, including temperature, equilibration time, extraction time, sample volume, water-honey ratio, and magnetic stirring velocities contribute simultaneously to the isolation of volatiles, sometimes synergistically [20]. For this reason, it is necessary to study all-factors-at-a-time, in terms of their effectiveness of volatiles isolation. This may be possible by using multivariate statistic techniques, like response surface methodology (RSM) [20,21].

The aim of the present study was the identification and semi-quantification of the volatile fraction of indigenous monofloral Greek autumn heather honey from *E. manipuliflora*. The main SPME factors were simultaneously examined for their potential to isolate the dominant volatile fraction and each molecule separately using RSM.

### **2. Materials and Methods**
