The Basic Principles Of high performance liquid chromatography
The Basic Principles Of high performance liquid chromatography
Blog Article
ディテクター（検出器）としては目的とする物質の性質に応じて光学的性質（吸光度、屈折率、蛍光等）、電気化学的性質、質量分析法などを利用する装置がある。
Rotating the inner valve (proven in purple) to the inject place directs the cellular section with the sample loop and on to the column.
Acid–foundation chemistry is not the only example of a secondary equilibrium reaction. Other examples incorporate ion-pairing, complexation, and also the conversation of solutes with micelles. We're going to think about the past of such in Chapter 12.7 once we focus on micellar electrokinetic capillary chromatography.
Non-polar molecules are slowed down on their own way through the column. They sort different degrees of attraction Using the hydrocarbon teams principally via van der Waals dispersion forces and hydrophobic interactions.
イオン交換クロマトグラフィーでは、無機イオンや高極性分子を電荷を利用して分離する。陽イオンタイプと陰イオンタイプの両方がある。イオン交換樹脂を利用する。
we figured out how to adjust the cellular period’s polarity by blending alongside one another two solvents. A polarity index, nevertheless, is simply a tutorial, and binary mobile period mixtures with identical polarity indices might not take care of equally a pair of solutes. Desk 12.5.2
In a very fuel chromatograph the pressure from the compressed gas cylinder is ample to push the cell section in the column. Pushing a liquid cellular period through a column, having said that, takes a fantastic offer much more hard work, creating pressures in excess of a number of hundred atmospheres.
前述した従来の順相タイプに対して、逆相クロマトグラフィーにおいては固定相に低極性のもの（例えばシリカゲルにアルキル基を共有結合させたもの）を、移動相に高極性のもの（例えば水や塩類の水溶液、アルコール、アセトニトリルなどの有機溶媒）を用いる。また珍しいケースではあるが、分離のための移動相pHをシリカゲルの使用範囲から外れたところに設定する必要がある場合、あるいはシリカゲル表面に残っている未反応シラノール基が分離に悪影響を及ぼし、かつそれが移動相の変更によっても解決できない場合には、固定相として樹脂を用いることがある。分析物はより極性の低いほどより強く固定相と相互作用して溶出が遅くなる。また極性の低い物質の割合が多い移動相ほど溶出が早くなる。
The concentration of caffeine in beverages is decided by a reversed-stage HPLC separation using a cell period of 20% acetonitrile and eighty% h2o, and using a nonpolar C8 column. Final results for any number of 10-μL injections of caffeine benchmarks are in the next table.
-hydroxybenzoic acid (PH) on the nonpolar C18 column subject matter to the maximum Evaluation time of 6 min. The shaded regions characterize locations in which a separation is not possible, Along with the unresolved solutes click here determined.
, which is the greater common kind of HPLC, the stationary stage is nonpolar as well as mobile section is polar. The most common nonpolar stationary phases use an organochlorosilane the place the R team is really an n
溶媒の組成に勾配を付けて（すなわち組成を連続的に変えて）溶出を行うことも多い。たとえば後述の逆相クロマトグラフィーにおいて水/メタノール勾配を使う場合、まずメタノールの少ない条件で極性の高い物質が溶出し、その後メタノールの割合を増加させてゆくに従ってより極性の低い物質が順次溶出する。これをグラジェント分析と呼ぶ。これに対し、一定組成の溶媒で分析物を溶出させる分析法をアイソクラテック分析と呼ぶ。
The elution order of solutes in HPLC is governed by polarity. For a standard-period separation, a solute of decreased polarity spends proportionally less time within the polar stationary period and elutes in advance of a solute which is extra polar. Presented a specific stationary period, retention moments in normal-section HPLC are managed by adjusting the cellular period’s properties. For example, if the resolution amongst two solutes is lousy, switching into a a lot less polar mobile phase retains the solutes around the column for an extended time and delivers extra opportunity for his or her separation.
Yet another useful detector can be a mass spectrometer. Determine twelve.5.13 shows a block diagram of a typical HPLC–MS instrument. The effluent from your column enters the mass spectrometer’s ion source applying an interface the removes many of the mobile stage, an read more essential will need because of the incompatibility among the liquid cellular stage plus the mass spectrometer’s high vacuum ecosystem.