Naslov
From the Analytical HPLC Level Upwards and Downwards

Autori
Šegudović, Nikola

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
6th Balaton symposium on high performance separation methods, Book of abstracts / Sz. Nyiredy - Siófok : Hungarian society for separation scince, 2005

Skup
6th Balaton symposium on high performance separation methods

Mjesto i datum
Siófok, Mađarska, 07.09.2005. - 09.09.2005

Vrsta sudjelovanja
Pozvano predavanje

Vrsta recenzije
Međunarodna recenzija

Ključne riječi
analytical HPLC; Upscale HPLC; Downscale HPLC

Sažetak
Over the past 30 years HPLC has become one of the most widely used technique in laboratories worldwide. This is due to the fact that HPLC is a dynamic and versatile technique. HPLC has evolved during this period in three primary areas : chemistry, detectors , and system/data management. The majority of HPLC applications are devoted to separation, identification, and quantitation of target compounds or group of compounds present in analytical matrix. Roughly speaking, common analytical HPLC equipment involves : 150-300 x 4-5 mm column filled by 5 &micro ; m particles, flow rates 1 or close to 1 mL/min, injection volume ≤ 25 &micro ; L, injected amount ≤ 1 mg, run time 15-30 min, and it is assumed that sample size is so small that it has no effect on the retention, efficiency and resolution of individual peaks within chromatogram. Besides analytical purposes HPLC is used for isolation and purification of target components for further analysis.. When only small amounts of pure product are required, an analytical HPLC can be used to recover nanogram to-sub milligram quantities. Larger amount can be obtained by repetitive injections. But much easier way for such task is application of semi-prep. Or prep. HPLC. Prep. HPLC is focusing on separation, isolation, and collection of larger quantity of more purified product. Prep. HPLC involves : 250mm or longer x 9.6mm or wider column filled with 10 &micro ; m or bigger particles, flow rates ≤ 10 mL, injection volume in high &micro ; L or mL level, injection amount in mg to g and run time close to the one obtained in analytical HPLC. In scaling up to preparative separation, it is recommended that initial development methods use an analytical-sized column. In this process some parameters are scaleable ( flow rates, injection volume, loop size, detector flow cell, , dead volume, solvent consumption, and throughput ) but some are not ( packing material, sample concentration, temperature, gradient slope, back pressure, run time, percent recovery ). With the linear scale-up factor prep. HPLC method could be optimized and normalized. On the opposite side of prep. HPLC are micro HPLC. The demand for greater sample throughput, low amount of sample, faster results, and lower mobile phase consumption has driven the creation of fast and ultra fast HPLC methods. While in conventional HPLC run time or cycle time take 15 or more minutes, in fast HPLC it is less than 5 min. And in ultra fast ( ultra performance ) the cycle time is less than 1 min. Thanks to new column technology ( monolitics or particles less than 2 &micro ; m ), corresponding development in LC equipment and automated, rapid and sophisticated data evaluation a thousand of samples in 24x7 mode could be analysed. For fast, analytical and semi-prep. mode of analysis the same equipment can be used but for prep. and ultra fast analysis the equipment has to be changed or modified.. All varieties of HPLC ( NP, RP, SEC, IC, isocratic and gradient ) can be scaled up or down from analytical level of analysis.

Izvorni jezik
Engleski

Znanstvena područja
Kemija

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