Novel Assays For Quality Evaluation Of Xue Bi Jing: Quality Variability Of A Chinese Herbal Injection Used In Sepsis Management

Xue Bi Jing is an intravenous five-herb injection licensed in China for sepsis.Which is prepared from a five-herb combination,comprising Carthamus tinctorius flowers,Paeonia lactiflora roots,Ligusticum chuanxiong rhizomes,Angelica sinensis roots and Salvia miltiorrhiza roots.The efficacy reproducibility of the injection depends on the injection’s quality consistency,this study was designed to develop new assays for quality evaluation and control of Xue Bi Jing.Objective:The aim of this investigation was to to evaluate the injection’s batch-to-batch variability by developing therapeutically relevant assays for quality evaluation and control of Xue Bi Jing.Method:In this study,selecting marker constituent is the first step for developing new assays,based on comprehensive understanding of chemical composition of the injection and the compounds exhibiting significant systemic exposure and desired pharmacokinetics/disposition,and potent bioactivities related to the injection’s antiseptic action.The second step is based the selected marker constituents for developing Liquid chromatography/tandem mass spectrometry(LC/MS-MS)-based and liquid chromatography/ultraviolet(LC/UV)-based multi-analyte assays using one-point calibration,which should meet the conditions: similarity of response to analyte in calibrator and in unknown samples and linearity through zero(i.e.,linear concentration-response function and near-zero intercept).The reliability of the assays was validated by overcoming interference from non-target constituents and excipients of Xue Bi Jing,and designing a linearity study.Measured using the validated assays,33 samples of Xue Bi Jing manufactured over a 3-year period were evaluated batch-to-batch variability,and the content levels measured by LC/MS-MS were compare with respective levels by LC/UV.Results:Hydroxysafflor yellow A,paeoniflorin,oxypaeoniflorin,albiflorin,senkyunolide I,senkyunolide G,ferulic acid,tanshinol,and protocatechuic aldehyde were selected as marker constituents,based on comprehensive understanding of the chemical composition of Xue Bi Jing and on the desired pharmacokinetics/disposition and known antisepsis-related bioactivities of the selected compounds.After overcoming interference from non-target constituents and excipients co-eluting with the marker constituents for LC/MS-MS and LC/UV measurement,a one-point calibrator was established,containing the nine selected compounds and Xue Bi Jing excipients,based on the linear ranges and near-zero intercepts of concentration-response function.Xue Bi Jing samples,as well as the calibrator,were diluted differently for different assays to guarantee compound measurement in the linear ranges.Measured using the validated assays,33 samples of Xue Bi Jing manufactured over a 3-year period exhibited small batch-to-batch variability in content levels of the marker constituents(4.1%–14.7%),except for senkyunolide I(26.5%).The content levels measured by LC/MS-MS were in good agreement with the respective levels by LC/UV,as indicated by the correlation coefficient of 0.9995(n = 297)with a slope of 1.009.The newly developed assays are reliable and can be routinely used in quality evaluation and control of Xue Bi Jing.Conclusion: The newly developed assays for quality evaluation and control of Xue Bi Jing,with the attributes: being therapeutically relevant,reflecting the wholeness of the Xue Bi Jing formula,and being reliable and practical are reliable and can be routinely used in quality evaluation and control of Xue Bi Jing.Measured using the validated assays,33 samples of Xue Bi Jing manufactured over a 3-year period exhibited small batch-to-batch variability in content levels of the marker constituents and showed good quality consistency.