Abstract
Plant-based food ingredients such as garlic, Chinese leek, Chinese onion, green onion and onion are widely used in many cuisines around the world. However, these ingredients known as the “five forbidden vegetables” (FFVs) are not allowed in some vegetarian diets. In this study, a loop-mediated isothermal amplification (LAMP) assay was developed for the detection of FFVs using five respective LAMP primer sets. The specific primers targeted the ITS1-5.8S-ITS2 nuclear ribosomal DNA sequence regions among the five vegetables. The results demonstrated that the identification of FFVs using the newly developed LAMP assay is more sensitive than the traditional PCR method. Using pepper, basil, parsley, chili and ginger as references, established LAMP primer sets showed high specificity for the identification of the FFV species. Moreover, when FFVs were mixed with other plant ingredients at different ratios (100:0, 50:50, 20:80, 10:90, 5:95, 2:98, and 1:99), no cross-reactivity was evident using LAMP. Finally, genomic DNAs extracted from boiled and steamed FFVs in processed foods were used as templates; the performance of the LAMP reaction was not influenced using validated LAMP primers. Not only can FFV ingredients be identified but commercial foods containing FFVs can also be authenticated. This LAMP method will be useful for the authentication of FFVs in practical food markets in the future.
Highlights
In recent years, many people have become vegetarian due to rising health concerns[1]
With respect to the four other loop-mediated isothermal amplification (LAMP) primer sets for the identification of Chinese leek, Chinese onion, green onion, and onion, our results showed that LAMP products were detected when their target DNAs were present in the samples
When any one set of LAMP primers such as those designed to identify garlic was used, the LAMP reaction was not influenced by the genomic DNAs of garlic, Chinese leek, Chinese onion, green onion, or onion when mixed at different percentages (Fig. 1B)
Summary
Many people have become vegetarian due to rising health concerns[1]. In addition to the exclusion of animal ingredients including meat, milk and eggs, it is forbidden for an ASVM to use aromatic or spicy plants or vegetables and their related flavours. One previous official document from the Taiwan Food and Drug Administration (TFDA) reported that FFVs can successfully be detected using PCR16 Some disadvantages, such as the amount of time required, assay sensitivity and integrity of the template DNA, that may reduce the effectiveness of the PCR assay were mentioned. Isothermal DNA amplification was applied for the development of a LAMP assay for the rapid, sensitive and specific identification of FFVs. the cross-reactivity of LAMP primers, integrity of the template DNA and commercially processed food products containing FFVs were examined to assess the feasibility of the established LAMP method for the identification of FFVs. To the best of our knowledge, this is the first report to identify FFVs using isothermal DNA amplification
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