Pharmanza Herbal Pvt. Ltd, Plot # 214, Borsad-Tarapur Road, Nr. Vadadla Patiya, At&PO: Kaniya, Petlad, Anand, Gujarat 388430, India.
J AOAC Int. 2024 Jan 4;107(1):129-139. doi: 10.1093/jaoacint/qsad110.
The sympatric occurrence of the species that often resulted in different gatherings of plant material, ambiguous history on traditional use, and taxonomic flux due to similarities within the Tinospora (Menispermaceae) taxa are some of the reasons that triggered the necessity to develop robust analytical methods for efficient QC, especially to recognize dry and powder forms.
To develop novel HPTLC-based fingerprinting of two closely resembling Tinospora species followed by HPTLC-MS analysis and identification of compounds differentiating Tinospora crispa (TCP) and Tinospora cordifolia (TCR) and a rapid and quantitative assessment by HPLC with a photodiode array detector (HPLC-PDA) with MS/MS characterization of specific TCP and TCR analytical markers.
An HPTLC-based method was developed using chloroform-toluene-methanol-formic acid (7 + 4 + 2 + 0.2, by volume). The TCP compounds could be distinguished and isolated using successive column chromatography with complete characterization. Further these used in the reverse phase (RP)-HPLC-PDA coupled with LC-ESI (electrospray ionization)-MS/MS to quantify and confirmation in TCP and TCR.
The fingerprinting showed distinct bands in TCP stems, confirmed as clerodane- furanoditerpenoids with indirect profiling by the HPTLC-MS technique. Systematic isolation confirmed these compounds as borapetosides B and E. Thus, the RP-HPLC-PDA method was developed for these borapetosides B and E, with tinosporide to differentiate these two species. The quantitation method was well validated with good linearity (r2 >0.99) with sensitive LOD (0.49-3.71 mcg/mL) and LOQ (1.48-11.23 mcg/mL) with recoveries of 92.34-96.19%.
A novel, validated HPLC-PDA method showed good resolution and reliability (up to 1% adulteration) in quantification for targeted major analytical markers from TCP to differentiate TCR. Thus, HPTLC and HPLC-PDA-based techniques are helpful with MS/MS-based characterization to identify and quantify these analytical markers from TCP (borapetoside B and E) and TCR (tinosporide) in dry and powder form.
This article reports on the systemic use of HPTLC-MS for separating and identifying analytical markers in Tinospora species, distinguishing TCR and TCP with quantitative HPLC-PDA and MS/MS assessment.
同域发生的物种经常导致植物材料的不同聚集，传统用途的历史不明确，以及由于三叶藤属（防己科）分类群内的相似性导致的分类学通量，这些都是需要开发强大的分析方法进行有效质量控制的原因，特别是用于识别干品和粉末形式。
建立两种相似三叶藤属植物的新型 HPTLC 指纹图谱，并进行 HPTLC-MS 分析，鉴定区分三叶藤（TCR）和三叶藤（TCP）的化合物，并通过带有光电二极管阵列检测器（HPLC-PDA）的 HPLC 进行快速定量评估，采用 MS/MS 对特定 TCP 和 TCR 分析标记物进行特征描述。
采用氯仿-甲苯-甲醇-甲酸（7+4+2+0.2，体积比）建立基于 HPTLC 的方法。使用连续柱层析可以分离和鉴定 TCP 化合物，并进行完全表征。进一步将这些化合物用于反相（RP）-HPLC-PDA 与 LC-ESI（电喷雾电离）-MS/MS 联用，以定量和确认 TCP 和 TCR 中的分析标记物。
指纹图谱显示 TCP 茎部有明显的条带，通过 HPTLC-MS 技术间接分析证实为 clerodane-furanoditerpenoids。系统分离证实这些化合物为 borapetosides B 和 E。因此，建立了用于这些 borapetosides B 和 E 的 RP-HPLC-PDA 方法，并使用 tinosporide 来区分这两个物种。定量方法经过良好验证，具有良好的线性（r2>0.99），具有较低的检测限（0.49-3.71 mcg/mL）和定量限（1.48-11.23 mcg/mL），回收率为 92.34-96.19%。
一种新型的、经过验证的 HPLC-PDA 方法在定量分析中显示出良好的分辨率和可靠性（高达 1%的掺假），用于从 TCP 中分离 TCR 的靶向主要分析标记物。因此，基于 HPTLC 和 HPLC-PDA 的技术有助于结合 MS/MS 进行特征描述，从 TCP（borapetoside B 和 E）和 TCR（tinosporide）的干品和粉末形式中鉴定和定量这些分析标记物。
本文报道了系统使用 HPTLC-MS 分离和鉴定三叶藤属植物中的分析标记物，通过定量 HPLC-PDA 和 MS/MS 评估来区分 TCR 和 TCP。
