SPE-通过固相萃取进行样品富集和纯化-waters

SPE——通过固相萃取进行样品富集和纯化

为何使用固相萃取（SPE）技术

1. 您需要从样品中去除特定干扰物，以免它们在目标分析物的检测和定量过程中影响实验结果。在此处所示的示例中，不适当的样品制备方案未能去除干扰物，导致提取物呈现出残留的黄色干扰物，色谱图中目标分析物与多个干扰峰发生了重叠。

2. 您需要提高初始样品中目标分析物的浓度，以便所用的分析技术能够更轻松地对其进行检测和准确定量。如果目标分析物可被较强地保留，那么可能需要在SPE色谱柱上加载较大的样品量，随后仅以极小体积的洗脱液将此分析物洗脱下来，由此提高样品中分析物的浓度。

3. 您需要去除样品中的干扰物（即使不可见），这些干扰物会在质谱检测中抑制目标分析物的信号。在此处的示例中，蛋白沉淀法无法去除血浆提取物中的磷脂，从而造成严重的离子抑制。优化的复合模式SPE方案可获取最纯净的提取物，并可在最大程度上降低离子抑制效应。

What is Solid-Phase Extraction (SPE)?

Don't be confused by the term solid-phase extraction [SPE]. A typical SPE device has 50 times more separation power than a simple, single liquid-liquid extraction. SPE is actually column liquid-solid chromatography. Since SPE is liquid chromatography [LC], its practice is

governed by LC principles. A sample is introduced into a column or a cartridge device containing a bed of appropriate particles, or other form, of a chromatographic packing material [stationary phase]. Solvent [mobile phase] flows through the bed. By choosing an appropriate combination of mobile and stationary phases, sample components may pass directly through the column bed, or they may be selectively retained.

Individual compounds in the sample each typically appear to travel at different speeds through the device. Using a weaker solvent causes them to move slowly and/or be strongly retained. A stronger solvent speeds up their passage through the bed and elutes the analyte(s) in a more concentrated volume. Elution from an SPE device is usually done by increasing the strength of the mobile phase in a series of discrete, rather than continuous, steps during which selected analytes or interferences are either fully retained or rapidly eluted-this variation of gradient elution called a step gradient.

Most commonly, SPE is practiced using miniature column or cartridge devices. An example is shown here. A mixture of three dyes is loaded onto the cartridge in a weak solvent, causing strong sample retention in a narrow band that appears black at the column inlet. Subsequent gradient steps, each with a successively stronger solvent, are used to elute the dyes individually [yellow, red, then blue].

Typical SPE cartridges are low-pressure devices-constructed of solvent-resistant plastic or glass-filled with particles ≥30 µm in diameter. Suitable flow rates may be achieved by gravity or with the assistance of vacuum or low positive pressure. [The latter requires putting a cap on the open inlet of a column or using a sealed device with inlet and outlet fittings.]

Importance of Sample Preparation

In the last two decades, dramatic advances in analytical instrumentation and laboratory information management systems shifted the analyst's predominant tasks from assay

measurements to sample preparation and data processing. As the stringency of requirements for higher sensitivity, selectivity, accuracy, precision, and number of samples to be processed has escalated, the corresponding increases in speed and sophistication of analysis and data collection have outpaced improvements in the many traditional techniques of sample

collection and preparation. By some estimates, 75 to 80% of the work activity and operating cost in a contemporary analytical lab is spent processing and preparing samples for

introduction or injection into an analytical separation and/or measurement device. Clearly, efforts directed and products designed to streamline sample preparation protocols are essential to future progress in analytical science.

Goals of Sample Preparation

Successful sample preparation for most analytical techniques [HPLC, GC, spectrophotometry, RIA, etc.] has a threefold objective: namely, to provide the sample component of interest

▪in solution

▪free from interfering matrix elements

▪at a concentration appropriate for detection or measurement.

To accomplish these goals, a sample, or a representative portion thereof [not always easy to obtain], is prepared via traditional methods of dissolution, homogenization, extraction

[liquid- or solid-phase], filtration, concentration, evaporation, separation, chemical

derivatization, standardization [internal or external], etc.

Usually such methods are used in combinations of multiple steps, which form a sample prep protocol. The fewer steps and methods used in any given protocol, the simpler, more

convenient, cost effective, and less time consuming it is. Simpler protocols lend themselves more readily to automation and also lead to increased accuracy, reliability, reproducibility, and safety.

Innovation in Sample Preparation Methods

There are many ways to combine standard tools and techniques to accomplish the goals of sample prep. However, it is best to seek innovative means to streamline sample prep

protocols:

▪to combine the functions of several steps, if possible, into one operation;

▪to eliminate needless sample transfers and manipulations;

▪to reduce the scale as much as practicable [gaining economies of time, labor, and cost];

▪to use new tools in creative ways.

Benefits of Solid-Phase Extraction [SPE] Cartridges

When compared to other sample preparation processes, solid-phase extraction using SPE cartridges offers: