Emerging Scientists in Analytical Sciences: Zhuoheng Zhou

IF 3 Q2 CHEMISTRY, ANALYTICAL Analytical science advances Pub Date : 2024-10-29 DOI:10.1002/ansa.202400057
Zhuoheng Zhou
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I was fascinated by the hidden complexity of mother nature, as much as the fact that it tends to leave us a backdoor to have a glimpse of the core. There was no such thing as attractive to a teenager as solving riddles with obscure hints scattered here and there. I later read about the biography of Antoine Lavoisier and his illumination on the essence of combustion [<span>1</span>]. The profound impact of his quantitative and analytical thinking on modern chemistry research made me realize that analytical science is <i>de facto</i> pro-science, the philosophies, methodologies and techniques it comprises are primers for any scientific research activities. To understand something, you have to “see” or “feel” it first.</p><p>I also owe my special thanks to Professor Bo Zhang at Xiamen University for hosting me as a research trainee during the summer of my second bachelor's year. This was the first time I had a chance to step on the floor of a research lab and to have a flavour of modern analytical instruments (mainly chromatographs, electrophoresis apparatus and spectrometers). Despite my “greenness”, I was still assigned to a small project on developing a fast reverse-phase chromatographic method as the second-separation dimension in a two-dimensional liquid chromatography (2D-LC) set-up, following a first-dimension strong cation-exchange separation, for intact protein profiling in velvet antler (a constituent of traditional Chinese medicines). The trainee program had a very generous deadline which permitted me to start with extensive exploratory scouting runs (a nicer way to say “trial-and-error”) and to get myself familiarized with various column chemistries and separation conditions (although in a rather primitive and intuitive way). I remember vividly the goose bumps that I had when I saw my first chromatogram on the screen with five distinctive peaks, only five minutes after I manually injected some sort of colourless, transparent, and homogeneous liquid that a senior prepared for me. The joy of knowing the supposedly unknown (more like finishing the last chapter of a detective fiction book) sparked my deep interest in the world of analytical science and particularly separation science up to this day.</p><p>My PhD thesis comprised of two major parts. The first part revolves around the instrumentation design and optimization for ultra-high-pressure LC (UHPLC). We demonstrated the influence of fluidic configurations on the dispersive and thermodynamic properties of chromatography eluents under extremely high-pressure conditions (1500 bar) [<span>2</span>]. To shed light on the trade-off between gain in overall efficiency and loss in column-accessible pressure when narrower tubing and smaller detector cells are used, we have borrowed the principle of the kinetic plot method—which has been mainly applied for column characterization—and extended its application to evaluate the kinetic performances of entire UHPLC system. Based on the insights we gained throughout this project, we have in collaboration with instrument vendors and government labs, successfully established an optimum UHPLC system configuration and applied it for ultra-high resolution profiling of multi-class residuals analysis in dairy products [<span>3</span>].</p><p>The second part focuses on advancing column technology for low-flow LC (LF-LC), as an indispensable chromatography technique coupled to mass spectrometry (MS) for bio-separations with significantly improved sensitivity. We have developed a protocol [<span>4</span>] describing the fabrication process of monolithic polymer stationary phases in capillary column formats, alongside guidelines on tuning the macropore structure targeting high kinetic performance. The versatility of the developed columns was demonstrated for high-throughput and high-resolution LF-LC bio-separations of intact proteins, peptides, and oligonucleotides, with verified robustness and repeatability. To further strengthen our understanding of the transport phenomena in polymer monoliths, tomographic reconstruction and stereological analysis were leveraged, to correlate the column bed morphology and kinetic performances for future column designing [<span>5</span>].</p><p>During my PhD, I was fortunate to have time to tidy up some of my research results and turn them into seven peer-reviewed publications and four international conference presentations. I was also privileged to be interviewed by LCGC as Rising Star of Separation Science in 2023, which offered me a great opportunity to share my work with the community. Besides external recognition, my proudest moments are actually dispersed in the daily research. In the project of a quantitative assessment of the 3D monolithic porous network, we went to great lengths to collaborate with chemical engineers, image analysts, and data scientists to find the proper morphological descriptors that can be used to chromatographically size a monolith, which has long been deemed challenging. We were among the first to explore the possibility of quantifying the Giddings’ trans-column velocity bias in a monolith, which surprisingly did not differ significantly from that of particulate columns. We believe these novel insights will be the cornerstones for future monolithic column design.</p><p>Besides Professor Bo Zhang who I mentioned supervised my Bachelor final-year project, I owe my deepest gratitude to my two PhD promotors. Professor Sebastiaan Eeltink, as my main promotor, his dedicated commitment to chromatography, profound knowledge of polymer chemistry, unbelievably hard-working ethics, detail-oriented philosophy and admirable people skills have shed tremendous light on every step of my path as a scientist. As a PhD promotor, his generosity truly gives meaning to this title. He would seize every opportunity to introduce myself and my research works to the peer scientists, not only to promote but also to attract potential interests of collaborators and eventually ended up with many joyful and fruitful scientific adventures.</p><p>My heartfelt thanks extend to my co-promotor Professor Gert Desmet, the chromatography “guru”, the living textbook, and my go-to-dictionary only one floor away. Whether it was the explanation of various solvent-strength models on the first day we met or our recent discussion on Giddings’ geometric constants, each scientific exchange with him has been challenging to fully grasp. Yet, the aftermath that lingered in the days following has deepened my understanding of concepts and reshaped my perspective on models.</p><p>Currently, we have been exploring the potential of ion-mobility spectrometry (IMS) in combination with high-resolution chromatography and MS to further push forward the data quality and analytical throughput of proteomic profiling (Figure 1, also see our recently published review article [<span>6</span>]). IMS, as a gaseous phase separation technique, adds an additional resolving capability after chromatographic elution. This second separation dimension not only isolates the peptide features from matrixial contaminants and differentiates isobaric pairs subject to co-fragmentation, but also offers the structural elucidation capability to provide additional attributes to increase identification confidence. Yet, similar to two-dimensional chromatography, the addition of IMS between LC and MS also requires method optimization to prevent separation resolution loss, dilution, and under- or over-sampling.</p><p>One clear trend I see in current analytical instrumentation is high throughput, which is especially relevant in industrial chemical analysis where time and operational costs are the main constraints. High-throughput analysis not only emphasizes the fundamental transformation in analytical principles (e.g., UHPLC in chromatography, data-independent acquisition in MS) but also requires novel sampling strategies (e.g., compatibility between instrumental analysis with large assays and chemical libraries) and additional technical advancement on minimizing overhead times. What also intrigues me is the blasting data generation permitted by high-throughput analysis, which requires new perspectives on results interpretation, from traditionally “one-spectrum-at-a-time” to statistical mining with the assistance of newly advanced machine learning toolboxes.</p><p>Another topic I am particularly interested in is the everlasting pursuit of instrument hyphenation to provide complementary characterizations with multiple data dimensions. Analytical science has significantly benefited from the assorts of hyphenated instruments as trivial as meeting an ad-hoc measuring task and as significant as unlocking a transformative research field (e.g., LC-MS to proteomics, cytometry by time of flight to single-cell analysis). These combinations have significantly improved the separation, detection, and analysis of complex samples, making it possible to explore previously uncharted areas in biology, environmental science, and materials research. As these technologies continue to evolve, they are likely to become more integrated, automated, and accessible, contributing to the advancement of scientific discovery and innovation across diverse industries. However, challenges remain in terms of instrument compatibility, cost, and complexity, requiring ongoing research and innovation.</p><p>As a separation scientist, my research interests are rooted deeply in bridging fundamental and technological advancement with real-world separation challenges. One of my areas of interest is the emerging field of biopharmaceutical analysis. The newly introduced biomolecule-based drug modalities, for example, recombinant and fusion proteins, monoclonal antibodies (and their drug conjugates), oligonucleotides and mRNA vaccines, come often with high structural complexity, chemical heterogeneity, and low diffusivity, which challenges the conventional separation techniques regarding both selectivity and efficiency. In order to meet the analytical needs of industrial applications, novel separation techniques together with instrumentation engineering are both required to further improve R&amp;D productivity. Conducting research in this field is where I see myself in the next ten years and probably many years to come.</p><p>When I am not in the lab, I often see myself in museums or on historical sites, either trying to decipher a Latin inscription or struggling to date an unknown painting based on the brushwork and pigments. 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Abstract

Through a collection of editorials titled “Emerging Scientists in Analytical Sciences,” we aim to spotlight promising individuals who are actively engaged in the realm of analytical sciences. For this editorial, we invited Zhuoheng Zhou who recently submitted his PhD thesis at the Vrije Universiteit Brussel.

During my junior high school years, I was very grateful to have attended a scientific event named “knowing about colours”, where teachers demonstrated the light dispersion through a prism and the separation of chlorophyll green and carotenoid yellow by a piece of filter paper (until many years later that I realized it was the “Eureka moment” of chromatography). I was fascinated by the hidden complexity of mother nature, as much as the fact that it tends to leave us a backdoor to have a glimpse of the core. There was no such thing as attractive to a teenager as solving riddles with obscure hints scattered here and there. I later read about the biography of Antoine Lavoisier and his illumination on the essence of combustion [1]. The profound impact of his quantitative and analytical thinking on modern chemistry research made me realize that analytical science is de facto pro-science, the philosophies, methodologies and techniques it comprises are primers for any scientific research activities. To understand something, you have to “see” or “feel” it first.

I also owe my special thanks to Professor Bo Zhang at Xiamen University for hosting me as a research trainee during the summer of my second bachelor's year. This was the first time I had a chance to step on the floor of a research lab and to have a flavour of modern analytical instruments (mainly chromatographs, electrophoresis apparatus and spectrometers). Despite my “greenness”, I was still assigned to a small project on developing a fast reverse-phase chromatographic method as the second-separation dimension in a two-dimensional liquid chromatography (2D-LC) set-up, following a first-dimension strong cation-exchange separation, for intact protein profiling in velvet antler (a constituent of traditional Chinese medicines). The trainee program had a very generous deadline which permitted me to start with extensive exploratory scouting runs (a nicer way to say “trial-and-error”) and to get myself familiarized with various column chemistries and separation conditions (although in a rather primitive and intuitive way). I remember vividly the goose bumps that I had when I saw my first chromatogram on the screen with five distinctive peaks, only five minutes after I manually injected some sort of colourless, transparent, and homogeneous liquid that a senior prepared for me. The joy of knowing the supposedly unknown (more like finishing the last chapter of a detective fiction book) sparked my deep interest in the world of analytical science and particularly separation science up to this day.

My PhD thesis comprised of two major parts. The first part revolves around the instrumentation design and optimization for ultra-high-pressure LC (UHPLC). We demonstrated the influence of fluidic configurations on the dispersive and thermodynamic properties of chromatography eluents under extremely high-pressure conditions (1500 bar) [2]. To shed light on the trade-off between gain in overall efficiency and loss in column-accessible pressure when narrower tubing and smaller detector cells are used, we have borrowed the principle of the kinetic plot method—which has been mainly applied for column characterization—and extended its application to evaluate the kinetic performances of entire UHPLC system. Based on the insights we gained throughout this project, we have in collaboration with instrument vendors and government labs, successfully established an optimum UHPLC system configuration and applied it for ultra-high resolution profiling of multi-class residuals analysis in dairy products [3].

The second part focuses on advancing column technology for low-flow LC (LF-LC), as an indispensable chromatography technique coupled to mass spectrometry (MS) for bio-separations with significantly improved sensitivity. We have developed a protocol [4] describing the fabrication process of monolithic polymer stationary phases in capillary column formats, alongside guidelines on tuning the macropore structure targeting high kinetic performance. The versatility of the developed columns was demonstrated for high-throughput and high-resolution LF-LC bio-separations of intact proteins, peptides, and oligonucleotides, with verified robustness and repeatability. To further strengthen our understanding of the transport phenomena in polymer monoliths, tomographic reconstruction and stereological analysis were leveraged, to correlate the column bed morphology and kinetic performances for future column designing [5].

During my PhD, I was fortunate to have time to tidy up some of my research results and turn them into seven peer-reviewed publications and four international conference presentations. I was also privileged to be interviewed by LCGC as Rising Star of Separation Science in 2023, which offered me a great opportunity to share my work with the community. Besides external recognition, my proudest moments are actually dispersed in the daily research. In the project of a quantitative assessment of the 3D monolithic porous network, we went to great lengths to collaborate with chemical engineers, image analysts, and data scientists to find the proper morphological descriptors that can be used to chromatographically size a monolith, which has long been deemed challenging. We were among the first to explore the possibility of quantifying the Giddings’ trans-column velocity bias in a monolith, which surprisingly did not differ significantly from that of particulate columns. We believe these novel insights will be the cornerstones for future monolithic column design.

Besides Professor Bo Zhang who I mentioned supervised my Bachelor final-year project, I owe my deepest gratitude to my two PhD promotors. Professor Sebastiaan Eeltink, as my main promotor, his dedicated commitment to chromatography, profound knowledge of polymer chemistry, unbelievably hard-working ethics, detail-oriented philosophy and admirable people skills have shed tremendous light on every step of my path as a scientist. As a PhD promotor, his generosity truly gives meaning to this title. He would seize every opportunity to introduce myself and my research works to the peer scientists, not only to promote but also to attract potential interests of collaborators and eventually ended up with many joyful and fruitful scientific adventures.

My heartfelt thanks extend to my co-promotor Professor Gert Desmet, the chromatography “guru”, the living textbook, and my go-to-dictionary only one floor away. Whether it was the explanation of various solvent-strength models on the first day we met or our recent discussion on Giddings’ geometric constants, each scientific exchange with him has been challenging to fully grasp. Yet, the aftermath that lingered in the days following has deepened my understanding of concepts and reshaped my perspective on models.

Currently, we have been exploring the potential of ion-mobility spectrometry (IMS) in combination with high-resolution chromatography and MS to further push forward the data quality and analytical throughput of proteomic profiling (Figure 1, also see our recently published review article [6]). IMS, as a gaseous phase separation technique, adds an additional resolving capability after chromatographic elution. This second separation dimension not only isolates the peptide features from matrixial contaminants and differentiates isobaric pairs subject to co-fragmentation, but also offers the structural elucidation capability to provide additional attributes to increase identification confidence. Yet, similar to two-dimensional chromatography, the addition of IMS between LC and MS also requires method optimization to prevent separation resolution loss, dilution, and under- or over-sampling.

One clear trend I see in current analytical instrumentation is high throughput, which is especially relevant in industrial chemical analysis where time and operational costs are the main constraints. High-throughput analysis not only emphasizes the fundamental transformation in analytical principles (e.g., UHPLC in chromatography, data-independent acquisition in MS) but also requires novel sampling strategies (e.g., compatibility between instrumental analysis with large assays and chemical libraries) and additional technical advancement on minimizing overhead times. What also intrigues me is the blasting data generation permitted by high-throughput analysis, which requires new perspectives on results interpretation, from traditionally “one-spectrum-at-a-time” to statistical mining with the assistance of newly advanced machine learning toolboxes.

Another topic I am particularly interested in is the everlasting pursuit of instrument hyphenation to provide complementary characterizations with multiple data dimensions. Analytical science has significantly benefited from the assorts of hyphenated instruments as trivial as meeting an ad-hoc measuring task and as significant as unlocking a transformative research field (e.g., LC-MS to proteomics, cytometry by time of flight to single-cell analysis). These combinations have significantly improved the separation, detection, and analysis of complex samples, making it possible to explore previously uncharted areas in biology, environmental science, and materials research. As these technologies continue to evolve, they are likely to become more integrated, automated, and accessible, contributing to the advancement of scientific discovery and innovation across diverse industries. However, challenges remain in terms of instrument compatibility, cost, and complexity, requiring ongoing research and innovation.

As a separation scientist, my research interests are rooted deeply in bridging fundamental and technological advancement with real-world separation challenges. One of my areas of interest is the emerging field of biopharmaceutical analysis. The newly introduced biomolecule-based drug modalities, for example, recombinant and fusion proteins, monoclonal antibodies (and their drug conjugates), oligonucleotides and mRNA vaccines, come often with high structural complexity, chemical heterogeneity, and low diffusivity, which challenges the conventional separation techniques regarding both selectivity and efficiency. In order to meet the analytical needs of industrial applications, novel separation techniques together with instrumentation engineering are both required to further improve R&D productivity. Conducting research in this field is where I see myself in the next ten years and probably many years to come.

When I am not in the lab, I often see myself in museums or on historical sites, either trying to decipher a Latin inscription or struggling to date an unknown painting based on the brushwork and pigments. Solving puzzles that have been architected by human civilization is as much fun as that orchestrated by nature (Figure 2).

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分析科学领域的新锐科学家:周卓恒
我们希望通过题为 "分析科学领域的新锐科学家 "的社论集,聚焦活跃在分析科学领域的有为人士。在这篇社论中,我们邀请了最近在布鲁塞尔自由大学(Vrije Universiteit Brussel)递交了博士论文的周卓恒。我被大自然隐藏的复杂性所吸引,就像它往往会给我们留一个后门来一窥其核心一样。对于一个十几岁的孩子来说,没有什么比解开散落着晦涩提示的谜语更有吸引力的了。后来,我读到了安托万-拉瓦锡的传记以及他对燃烧本质的启示[1]。他的定量分析思想对现代化学研究的深远影响让我意识到,分析科学事实上是亲科学的,它所包含的哲学、方法论和技术是任何科学研究活动的入门读物。我还要特别感谢厦门大学的张波教授,他在我本科二年级的暑假接待了我,让我成为了一名研究实习生。这是我第一次有机会踏进研究实验室的地板,领略现代分析仪器(主要是色谱仪、电泳仪和光谱仪)的魅力。尽管 "青涩",我还是被分配到了一个小项目,开发一种快速反相色谱法,作为二维液相色谱(2D-LC)装置中的第二分离维度,继第一维度强阳离子交换分离之后,用于鹿茸(一种中药成分)中完整蛋白质的分析。见习项目的期限非常宽松,允许我从大量的探索性运行("试错 "的好听说法)开始,熟悉各种色谱柱化学成分和分离条件(尽管是以相当原始和直观的方式)。我清楚地记得,当我手动注入前辈为我准备的某种无色、透明、均质的液体仅仅五分钟后,就在屏幕上看到了我的第一张色谱图,上面有五个明显的色谱峰时,我的鸡皮疙瘩都起来了。我的博士论文由两大部分组成。第一部分围绕超高压液相色谱(UHPLC)的仪器设计和优化展开。我们证明了流体配置对超高压条件(1500 巴)下色谱洗脱液的分散性和热力学特性的影响[2]。为了弄清在使用更窄的管路和更小的检测器单元时,整体效率的提高与色谱柱可利用压力的降低之间的权衡,我们借鉴了动力学图法的原理(该方法主要用于色谱柱表征),并将其应用扩展到整个超高效液相色谱系统的动力学性能评估。基于在整个项目中获得的洞察力,我们与仪器供应商和政府实验室合作,成功建立了最佳超高效液相色谱系统配置,并将其应用于乳制品中多类残留物的超高分辨率谱分析[3]。第二部分的重点是推进低流量液相色谱(LF-LC)的色谱柱技术,这是一种与质谱(MS)联用的不可或缺的色谱技术,可显著提高生物分离的灵敏度。我们已经制定了一套方案[4],描述了毛细管色谱柱格式的整体聚合物固定相的制造过程,以及以高动力学性能为目标的大孔结构调整指南。所开发色谱柱的多功能性已在完整蛋白质、肽和寡核苷酸的高通量和高分辨率 LF-LC 生物分离中得到证实,其稳健性和可重复性也得到了验证。为了进一步加强我们对聚合物单体中传输现象的理解,我们利用了层析成像重建和立体分析技术,将柱床形态和动力学性能联系起来,用于未来的柱设计[5]。 我还有幸作为2023年分离科学新星接受了LCGC的采访,这为我提供了一个与社会分享我的工作的绝佳机会。除了外界的认可,我最自豪的时刻其实也分散在日常的科研工作中。在三维整体多孔网络定量评估项目中,我们不遗余力地与化学工程师、图像分析师和数据科学家合作,寻找适当的形态描述符,用于对整体进行色谱定量,而这一直被认为是一项挑战。我们是首批探索量化整体柱中吉丁斯跨柱速度偏差的研究人员之一,令人惊讶的是,整体柱的速度偏差与颗粒柱的速度偏差并无明显差异。我们相信,这些新见解将成为未来整体柱设计的基石。除了我提到的指导我本科毕业项目的张波教授外,我还要向我的两位博士生导师致以最深切的谢意。Sebastiaan Eeltink 教授是我的主要导师,他对色谱学的执着追求、对高分子化学的渊博知识、令人难以置信的勤奋道德、注重细节的理念以及令人钦佩的人际交往能力,为我科学家道路上的每一步指明了方向。作为一名博士生导师,他的慷慨确实赋予了这个称号以意义。他抓住一切机会向同行科学家介绍我和我的研究工作,不仅是为了宣传,也是为了吸引潜在的合作者的兴趣,最终成就了许多快乐而富有成果的科学探险。我衷心感谢我的合作导师 Gert Desmet 教授,他是色谱 "大师",是活生生的教科书,也是离我只有一层楼的我的必备字典。无论是第一天见面时对各种溶剂强度模型的解释,还是最近对吉丁斯几何常数的讨论,与他的每一次科学交流都具有挑战性,让人难以完全掌握。目前,我们一直在探索离子迁移谱法(IMS)与高分辨率色谱法和质谱法相结合的潜力,以进一步提高蛋白质组分析的数据质量和分析通量(图 1,另见我们最近发表的综述文章 [6])。IMS 作为一种气相分离技术,可在色谱洗脱后增加额外的分辨能力。这第二个分离维度不仅能将肽特征从基质污染物中分离出来,还能区分可能发生共破碎的等位肽对,同时还能提供结构阐释能力,为提高鉴定可信度提供更多属性。然而,与二维色谱法类似,在 LC 和 MS 之间添加 IMS 也需要对方法进行优化,以防止分离分辨率损失、稀释以及进样不足或进样过多。我在当前的分析仪器中看到的一个明显趋势是高通量,这在时间和操作成本是主要限制因素的工业化学分析中尤为重要。高通量分析不仅强调分析原理的根本转变(如色谱法中的超高效液相色谱、质谱法中的数据独立采集),而且还需要新颖的取样策略(如仪器分析与大型检测和化学库之间的兼容性),以及在最大限度缩短开销时间方面的更多技术进步。同样令我感兴趣的是高通量分析所允许的爆炸性数据生成,这需要新的结果解读视角,从传统的 "一次一谱 "到借助新的先进机器学习工具箱进行统计挖掘。我特别感兴趣的另一个话题是对仪器联用的不懈追求,以提供多数据维度的互补性表征。分析科学从联用仪器的组合中获益匪浅,小到满足一项临时测量任务,大到开启一个变革性的研究领域(例如,从液相色谱-质谱联用到蛋白质组学,从飞行时间细胞测量到单细胞分析)。这些组合极大地改进了复杂样品的分离、检测和分析,使人们有可能探索生物学、环境科学和材料研究领域以前未知的领域。随着这些技术的不断发展,它们可能会变得更加集成化、自动化和便捷化,从而推动各行各业的科学发现和创新。然而,在仪器兼容性、成本和复杂性方面仍然存在挑战,需要不断进行研究和创新。
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