Wheat-Dasypyrum villosum translocated chromosomes T6V#2S•6AL and T6V#4S•6DL are known to confer excellent resistance to wheat powdery mildew (PM). However, it is difficult to distinguish the two sources of PM resistance genes through multi-pathotype testing because to date no virulence for them has been found. To reveal the relationship between the PM resistance genes from the two translocations, the sequence of the Stpk-V gene, a key member of powdery mildew resistance locus Pm21, was used as a reference to isolate homologous genes from a D. villosum accession No.1026 and its derivatives 6V#4(6D) disomic substitution (DS) line RW15 and T6V#4S•6DL translocation line Pm97033. Two genes Stpk-V2 and Stpk-V3 were cloned from No.1026. Sequence alignment showed that Stpk-V2 and Stpk-V3 shared 98.2 % and 96.2 % of their DNA and 99.3 % and 100 % of their amino acids in identity with Stpk-V. Compared with Stpk-V, a 22-bp direct sequence repeat and a miniature inverted-repeat transposable element (MITE) were found in the intron 4 of Stpk-V2 and Stpk-V3, respectively. However, Stpk-V2 was not present in DS line RW15 and translocation line Pm97033 based on the PCR result, indicating that Stpk-V2 did not contribute to the PM resistance of RW15 and Pm97033. In the promoter region, a 78-bp insertion was found not only in Stpk-V2 and Stpk-V3, but also in its orthologous gene Stpk-A of wheat. In addition, there was a 17 bp/8 bp deletion/insertion in the putative promoter of Stpk-V3 in comparison with that of Stpk-V/Stpk-V2. Real-time quantitative RT-PCR analysis indicated that the expression levels of Stpk-V and Stpk-V3 genes in the translocation lines were induced by the pathogen, but Stpk-V had a higher expression level than Stpk-V3 at 12 h after inoculation with Bgt. The diversity of Stpk-V gene will help to explore new resistance genes to PM in D. villosum for wheat breeding.

Summary. An improved ion-pairing reversed-phase high-performance liquid chromatography method coupled with evaporative light scattering detection (HPLC-ELSD) was developed to determine spectinomycin and its related substances in commercial samples. The method was validated in accordance with International Conference on Harmonization (ICH) guidelines. The specificity of the HPLC-ELSD method was similar to that of the European Pharmacopoeia (Ph. Eur.) method, and repeatability and robustness were markedly improved relative to other reported methods due to our empirical evaluation of separation columns. Indeed, it is a more specific assay of spectinomycin than traditional microbiological techniques. The HPLC-ELSD method was used to evaluate the impurity profiles of eight compounds in seven spectinomycin batches from five different companies. Liquid chromatography coupled with electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) was employed to characterize the structures of these compounds. Though the HPLC-ELSD method was not as sensitive as the Ph. Eur. method, its limit of quantitation (LOQ) (0.16%) was lower than the disregard limit (0.3%) described by the Ph. Eur. 7.0. This suggests that the HPLC-ELSD method is appropriate for routine analysis of spectinomycin and its related substances.

The functional food ingredients of apple juice can significantly change during processing, transportation, and storage, thus affecting the quality of the product. A simple and derivation‐free analytical method based on ultra‐high‐performance liquid chromatography–tandem mass spectrometry (UPLC‐MS/MS) was developed and optimized for the simultaneous determination of functional food ingredients in apple juice bought in the market. Cleanup steps and chromatographic conditions were optimized to remove interference and decrease the matrix effect. The nine target analytes were separated on an Acquity UPLC system equipped with a BEH C18 column and detected by electrospray ionization source (ESI) operating in positive subsection acquisition mode under multiple reaction monitoring (MRM) conditions. The results showed that p‐hydroxybenzoic acid, protocatechuate, caffeic acid, chlorogenic acid, epicatechin, phloridzin, hyperoside, procyanidin B2, and rutin could be sufficiently separated for content determination within 6 min. In the concentration range of 20 μg/L–50 mg/L, nine standard samples exhibited a good linear fit with correlation coefficients above .985.