Laboratory of Food Chemistry and Technology, School of Chemistry, Aristotle University of Thessaloniki, GR-54124 Thessaloniki, Greece; Institute of Applied Biosciences, CERTH, Thermi, GR-570 01 Thessaloniki, Greece.
Laboratory of Food Chemistry and Technology, School of Chemistry, Aristotle University of Thessaloniki, GR-54124 Thessaloniki, Greece.
Food Res Int. 2017 Oct;100(Pt 1):899-907. doi: 10.1016/j.foodres.2017.08.001. Epub 2017 Aug 3.
Saffron, the spice produced from the red stigmas of the flower of Crocus sativus L. is a frequent target of fraud and mislabeling practices that cannot be fully traced using the ISO 3632 trade standard specifications and test methods. A molecular approach is proposed herein as a promising branding strategy for the authentication of highly esteemed saffron brands such as the Greek Protected Designation of Origin (PDO) "Krokos Kozanis". Specific ISSR (inter-simple sequence repeat) markers were used to assess for the first time, the within species variability of several populations of C. sativus L. from the cultivation area of "Krokos Kozanis" as well as the potential differences with the band pattern produced by other Crocus species. Then, species-specific markers were developed taking advantage of an advanced molecular technique such as the HRM analysis coupled with universal DNA barcoding regions (trnL) (Bar-HRM) and applied to saffron admixtures with some of the most common plant adulterants (Calendula officinalis, Carthamus tinctorius, Gardenia jasminoides, Zea mays and Curcuma longa). The sensitivity of the procedure was tested for turmeric as a case study whereas HPLC-fluorescence determination of secondary metabolites was also employed for comparison. The overall results indicated that the Bar-HRM approach is quite effective in terms of specificity and sensitivity. Its effectiveness regarding the detection of turmeric was comparable to that of a conventional HPLC method (0.5% vs 1.0%, w/w). Yet, the proposed DNA-based method is much faster, cost-effective and can be used even by non-geneticists, in any laboratory having access to an HRM-capable real-time PCR instrumentation. It can be, thus, regarded as a strong analytical tool in saffron authentication studies.
藏红花，由番红花的红色柱头制成的香料，是经常受到欺诈和贴错标签做法的目标，这些做法无法完全通过 ISO 3632 贸易标准规范和测试方法来追踪。本文提出了一种分子方法，作为一种有前途的品牌策略，用于认证备受推崇的藏红花品牌，如希腊受保护的原产地名称（PDO）“Krokos Kozanis”。特定的 ISSR（简单序列重复间）标记首次被用于评估来自“Krokos Kozanis”种植区的几个番红花属植物种群的种内变异性，以及与其他番红花属物种产生的条带模式的潜在差异。然后，利用 HRM 分析与通用 DNA 条形码区域（trnL）（Bar-HRM）相结合的先进分子技术，开发了物种特异性标记，并将其应用于藏红花混合物与一些最常见的植物掺杂物（金盏花、红花、栀子、玉米和姜黄）。以姜黄为例，测试了该程序的灵敏度，同时还采用 HPLC-荧光法测定次生代谢物进行比较。总体结果表明，Bar-HRM 方法在特异性和灵敏度方面非常有效。其对姜黄的检测效果与传统的 HPLC 方法相当（0.5%对 1.0%，w/w）。然而，与传统的 HPLC 方法相比，这种基于 DNA 的方法速度更快、更具成本效益，即使是非遗传学家也可以在任何有能力进行 HRM 的实时 PCR 仪器的实验室中使用。因此，它可以被视为藏红花鉴定研究中的一种强大分析工具。
