ABSTRACT
1,4-dihydropyridine (DHP) scaffold occupies a prominent position among all heterocyclic compounds owing to its versatile pharmacological properties, particularly its well-known calcium channel blocking activity. In the quest of developing new calcium channel blockers, fifty seven 5-oxo-hexahydroquinoline (HHQ) derivatives carrying DHP framework in a condensed ring system were recently synthesized as racemic mixtures. Due to their potential as drug candidates, enantiomers arising from the asymmetric center at the C-4 position of the HHQ ring were separated. Four modern columns packed with 2.7⯵m superficially porous particles bonded with a chiral selector were used. The chiral selectors were three macrocyclic glycopeptide selectors: vancomycin, teicoplanin, and a macrocyclic derivative called nico. The fourth bonded selector was the dinitrobenzamido-tetrahydrophenanthrenyl derivative called Whelko. The four chromatographic modes were assayed with the mobile phase compositions: reversed phase with acetonitrile/buffer 30/70â¯%v/v, normal phase with hexane/ethanol 80/20â¯%v/v, and subcritical fluid chromatography with CO2/methanol 80/20â¯%v/v at 25⯰C. The WhelkoShell column was the most effective in separating this set of 57 compounds. Several enantioresolution factors passed 20 with enantioselectivity ratios higher than 4. Molecular modeling showed that the compounds had a T-shape that fitted well the molecular structure of the WhelkoShell selector in the normal or subcritical modes. Additionally, seven compounds had a second chiral center. The NicoShell column was able to separate all four stereoisomers of these compounds in the reversed phase mode. The preparative production of pure enantiomers of these compounds would be straightforward using the WhelkoShell column in the subcritical mode.
ABSTRACT
Since the identified standard genetic code contains 61 triplet codons of three bases for the 20 L-proteinogenic amino acids (AAs), no D-AA should be found in natural products. This is not what is observed in the living world. D-AAs are found in numerous natural compounds produced by bacteria, algae, fungi, or marine animals, and even vertebrates. A review of the literature indicated the existence of at least 132 peptide natural compounds in which D-AAs are an essential part of their structure. All compounds are listed, numbered and described herein. The two biosynthetic routes leading to the presence of D-AA in natural products are: non-ribosomal peptide synthesis (NRPS), and ribosomally synthesized and post-translationally modified peptide (RiPP) synthesis which are described. The methods used to identify the AA chirality within naturally occurring peptides are briefly discussed. The biological activity of an all-L synthetic peptide is most often completely different from that of the D-containing natural compounds. Analyzing the selected natural compounds showed that D-Ala, D-Val, D-Leu and D-Ser are the most commonly encountered D-AAs closely followed by the non-proteinogenic D-allo-Thr. D-Lys and D-Met were the least prevalent D-AAs in naturally occurring compounds.
ABSTRACT
A loading and productivity study was done using three racemates on vancomycin and teicoplanin-bonded chiral stationary phases of different particle formats. Two columns were packed with 2.7 µm superficially porous particles and two columns were packed with identically bonded 5 µm fully porous particles. The last two columns were packed with specially synthesized 4.5 µm vancomycin and teicoplanin superficially porous particles. The loading of different chiral compounds showed that the columns filled with 2.7-µm chiral stationary phases were inappropriate for preparative separations due to their very low permeability which precluded high flow rates. However, columns containing 4.5 µm superficially porous (core-shell) particles were as effective for small-scale preparative chiral separations as columns filled with classical 5 µm fully porous particles. Comparing the 4.5 µm superficially porous particles and 5 µm fully porous particles teicoplanin columns, the observed respective productivities of 270 and 265 mg/g chiral phase/h for 5-methyl-5-phenyl hydantoin enantiomers were obtained. Particular attention was given to the peculiar case of the mianserin enantiomeric separation on vancomycin columns that gave observed productivities of 200 and 205 mg/g chiral phase/h on the 4.5 µm superficially porous particles and 5 µm fully porous particles, respectively.
ABSTRACT
Superficially porous silica bonded with macrocyclic glycopeptides can separate enantiomers in various chromatographic formats, including normal phase liquid chromatography (NPLC). The conventional wisdom in NPLC is to avoid intentionally adding water in the eluents. Herein we examine the effects of small quantities of water as an additive on chiral separations in NPLC with the n-hexane-ethanol system. A phase diagram (n-hexane-ethanol-water) is used to analyze the physicochemical properties of the mobile phase. The relative polarity change of solvents upon adding water was determined by using bathochromic shifts of dissolved Nile Red dye. The effectiveness of chiral NPLC with water traces is demonstrated for various pharmaceutically relevant enantiomeric compounds. It is postulated that water molecules weaken stationary phase-solute interactions, resulting in lower retention times for both enantiomers in addition to significantly higher efficiencies. Gibbs free energy changes provided an understanding of the different enantioselectivity shifts caused by water addition. Some interesting kinetic effects also were observed. Classical van Deemter curves are not observed on macrocyclic glycopeptide stationary phases due to slow mass transfer kinetics and thermal effects at high flow rates. The most significant advantage of adding water in NPLC is reducing mass transfer kinetics and altering the mass overloading properties which is highly beneficial on macrocyclic glycopeptide phases. By overloading a 10 × 0.46-cm column with up to 0.6 mg alprenolol, it was found that the relative adsorption isotherm of the first eluting enantiomer was switched from Langmuir to anti-Langmuir type by water addition. The peak shape tuning effect demonstrated the strong influence of water on specific interaction sites of the chiral stationary phases. Water addition effects were most beneficial for enantiomeric and preparative separations in NP mode.
Subject(s)
Glycopeptides , Water , Chromatography, High Pressure Liquid/methods , Chromatography, Liquid , Glycopeptides/chemistry , Solvents , StereoisomerismABSTRACT
Eleven racemic ethanolamine derivatives were prepared, and their enantiomers were separated using liquid chromatography with various chiral columns. These derivatives included chiral vicinal amino alcohols, ß-hydroxy ureas, ß-hydroxy thioureas, and ß-hydroxy guanidines, all of which are present in many active pharmaceutical ingredients. The screening study was performed with six chiral stationary phase containing columns, including four recently introduced superficially porous particles bonded with two macrocyclic glycopeptides, a cyclodextrin derivative and a cyclofructan derivative. The two remaining columns contained chiral stationary phases, based on either a cellulose derivative or derivatized amylose, both bonded to fully porous particles. The cyclodextrin and cellulose-based chiral stationary phases proved to be the most broadly effective selectors and were able to separate 8 and 7 of the 11 tested compounds, respectively. With respect to analyte structural features, marked differences in enantiorecognition were observed between compounds containing phenyl and cyclohexyl groups adjacent to the stereogenic center. Additionally, replacing a small electronegative oxygen atom by a larger and less electronegative sulfur atom induced a significant difference in chiral recognition by the cellulose derivative as well as by the vancomycin-based chiral selectors.
Subject(s)
Ethanolamine , Glycopeptides , Chromatography, High Pressure Liquid/methods , Chromatography, Liquid , Glycopeptides/chemistry , StereoisomerismABSTRACT
Many steps are needed in the synthesis of a new active pharmaceutical ingredient (API). In a practical case proposed by a French pharmaceutical company, an intermediate synthesis step, needed to protect 8 hydroxyl groups before oxidation, could produce a mixture of neutral compounds containing up to 652 structures being positional isomers of 18 molecular formulas. Some mixtures allowed obtaining the desired API, others did not. An efficient analytical method was needed to characterize these neutral positional isomers and identify the mixtures to reject. Two samples were provided by the pharmaceutical company: Sample A was conform, Sample B was not. 8 RPLC columns were used with different gradients to screen Sample A. Next, the best RPLC separation was used as the second dimension fast analysis in a comprehensive 2D-RPLC systems. Two columns were used as first dimension: a fluorinated one and a zirconium based one. An order of magnitude was gained in peak capacity, but a better sample characterization was still needed. An off-line RPLC x SFC x Q-TOF/MS analysis was performed collecting 96 RPLC fractions and analyzing them by SFC with Q-TOF/MS detection. A home-made software associated the 96 SFC MS chromatograms to produce either base peak (BPC) or extract ion (EIC) contour plots that allowed for a satisfying characterization of the samples. Subtracting the EIC of expected m/z compounds from the Sample B BPC contour plot produced a unique new contour plot clearly pointing out unexpected compounds explaining the failure of the synthesis and possibly allowing improving the synthesis process.
Subject(s)
Chromatography, Reverse-Phase , Pharmaceutical Preparations , IsomerismABSTRACT
Twelve new azole compounds were synthesized through an ene reaction involving methylidene heterocycles and phenylmaleimide, producing four oxazoles, five thiazoles, and one pyridine derivative, and ethyl glyoxal for an oxazole and a thiazole compound. The twelve azoles have a stereogenic center in their structure. Hence, a method to separate the enantiomeric pairs, must be provided if any further study of chemical and pharmacological importance of these compounds is to be accomplished. Six chiral stationary phases were assayed: four were based on macrocyclic glycopeptide selectors and two on linear carbohydrates, i.e., derivatized maltodextrin and amylose. The enantiomers of the entire set of new chiral azole compounds were separated using the three different mobile phase elution modes: normal phase, polar organic, and reversed phase. The most effective chiral stationary phase was the MaltoShell column, which was able to separate ten of the twelve compounds in one elution mode or another. Structural similarities in the newly synthesized oxazoles provided some insights into possible chiral recognition mechanisms.
Subject(s)
Amylose/chemistry , Azoles/chemistry , Azoles/isolation & purification , Glycopeptides/chemistry , Polysaccharides/chemistry , Molecular Structure , StereoisomerismABSTRACT
N-methyl-D-aspartate (NMDA) receptors, which are widely present in the central nervous system, have also been found to be up-regulated in a variety of cancer cells and tumors and they can play active roles in cancer cell growth regulation. NMDA receptor antagonists have been found to affect cancer cell viability and interfere with tumor growth. Moreover, cancer cells also have been shown to have elevated levels of some D-amino acids. Two human skin cell lines: Hs 895.T skin cancer and Hs 895.Sk skin normal cells were investigated. They were derived from the same patient to provide tumor and normal counterparts for comparative studies. The expression of specific NMDA receptors was confirmed for the first time in both skin cell lines. Dizocilpine (MK-801) and memantine, NMDA receptor channel blockers, were found to inhibit the growth of human skin cells by reducing or stopping NMDA receptor activity. Addition of D-Ser, D-Ala, or D-Asp, however, significantly reversed the antiproliferative effect on the human skin cells triggered by MK-801 or memantine. Even more interesting was the finding that the specific intracellular composition of a few relatively uncommon amino acids was selectively elevated in skin cancer cells when exposed to MK-801. It appears that a few specific and upregulated D-amino acids can reverse the drug-induced antiproliferative effect in skin cancer cells via the reactivation of NMDA receptors. This study provides a possible innovative anticancer therapy by acting on the D-amino acid pathway in cancer cells either blocking or activating their regulatory enzymes.
Subject(s)
Amino Acids/metabolism , Cell Proliferation/drug effects , Excitatory Amino Acid Antagonists/pharmacology , Neuroprotective Agents/pharmacology , Receptors, N-Methyl-D-Aspartate/metabolism , Skin Neoplasms/metabolism , Alanine/metabolism , Alanine/pharmacology , Amino Acids/pharmacology , Asparagine/metabolism , Asparagine/pharmacology , Cell Line, Tumor , Cell Survival/drug effects , Dizocilpine Maleate/pharmacology , Female , Humans , Memantine/pharmacology , Middle Aged , Receptors, N-Methyl-D-Aspartate/antagonists & inhibitors , Receptors, N-Methyl-D-Aspartate/genetics , Serine/metabolism , Serine/pharmacologyABSTRACT
Power functions, p n ( x ) = [ f ( x ) ] n , are embedded in some modern chromatography detectors and software which not only alter the linear dynamic range of such detectors but also improve the cosmetic aspects of the chromatograms. These aspects include a reduction in baseline noise, improved peak symmetry, and better resolution. However, after raising the electronic output of a detector to a selected power (n > 1), the original peak area information is lost as are very small peaks. Recent advances in the peak processing protocol allow us to recover peak areas from overlapping peak areas, even in noisy environments using power functions. One of the primary protocol requirements of this approach was to have resolution factors ≥0.9. An increasing positive bias in the recovered area was observed as the resolution factors decreased below 0.9. In this work, we extend the capabilities of power function to lower resolution values. This bias is addressed by considering the increasing contributions in peak height coming from adjacent peaks. It is shown that the power function can now be used as long as the peak maxima are visible, making R s = 0.5, the lower treatable resolution factor for two peaks of similar areas.
ABSTRACT
Affinity monolith columns of 375 nL (effective length 8.5 cm, internal diameter 75 µm) were developed for protein-ligand affinity investigations needing only 3 µg of human serum albumin (HSA). To promote specific interactions and avoid non-specific ones, different combinations of monolithic supports and bio-functionalization pathways were evaluated. Silica and glycidylmethacrylate based monoliths were in-situ synthesized and grafted with HSA. Two direct grafting methods epoxy-amine and Schiff Base plus the streptavidin-biotin method were compared. The columns were evaluated by frontal analysis with ligands of known affinity for HSA. It is shown that a classical capillary electrophoresis instrument equipped with an external pressure device can be used to do weak affinity chromatography at low pressure (less than 1.2 MPa) in a fully automated way and with very low reagent consumption. The grafting pathways were compared in terms of (i) total and active amounts of immobilized protein, (ii) non-specific interactions, (iii) protein denaturation. According to these criteria, the organic monoliths combined with the streptavidin-biotin approach provided the best results. This immobilization pathway led to the highest active protein content (40 pmol of HSA per 8.5-cm column) with less than 10% non-specific interactions and 84% protein activity. The target grafting step lasts only 10 min and is UV-monitored, the UV breakthrough curve giving the exact amount of bound protein. This novel approach was validated by Kd measurements of 3 known ligands of HSA. Streptavidin generic monolith columns could be stored at 4 °C for 3 months maintaining activity. µg of a biotin modified sensitive protein could be attached to a stable streptavidin monolith for immediate interaction studies avoiding stability problems. This development was subsequently extended to another protein of higher pharmaceutical interest: the N-terminal domain of HSP90. Affinity was measured for two known ligands and determined Kd values were in accordance with the literature, proving that our technique is applicable to other proteins.
Subject(s)
Biotin/chemistry , Chromatography, Affinity/methods , Protein Binding , Streptavidin/chemistry , Electrophoresis, Capillary , Epoxy Compounds/chemistry , Humans , Methacrylates/chemistry , Serum Albumin, Human/chemistry , Silicon Dioxide/chemistryABSTRACT
Dicationic ionic liquids (DILs) are more and more accepted as a new class of high temperature and polar stationary phases for gas chromatography (GC). This study deals with the effect of seven different fluorosulfonyl derivatized anions associated with two dications: 1,3-di(3-methylimidazolium)-2-methylpropane [2mC3(mim)2], and 1,3-di(3-methylimidazolium)-isobutene [i-eneC4(mim)2]. Thermophysical properties of the 14 synthesized DILs were evaluated in terms of melting point, viscosity, and thermal stability. The optimal physicochemical properties of 13 DILs allowed preparing 13â¯GC capillary columns. Accordingly, the polarity and selectivity of the DILs were evaluated by determining the Rohrschneider-McReynolds constants and the equivalent chain lengths of C18 fatty acid methyl esters (FAMEs). The symmetrical fluoroalkylsulfonyl and the trifluorosulfonate anions seem to produce the most polar DILs. Compared to classical polyethyleneglycol phases, the DILs showed substantially decreased retention of apolar compounds and a much stronger retention of the polar ones. Unique selectivities were observed with unsaturated FAMEs, polyaromatic hydrocarbons, and bacterial specific FAMEs. The two applications presented included a biodiesel and bacterial FAME analyses.
Subject(s)
Chromatography, Gas/methods , Esters/analysis , Fatty Acids/analysis , Ionic Liquids/chemistry , Polycyclic Aromatic Hydrocarbons/analysis , Anions/chemistry , Cations/chemistry , Molecular Structure , TemperatureABSTRACT
Current trends in chiral analysis of pharmaceutical drugs are focused on faster separations and higher separation efficiencies. Core-shell or superficially porous particles (SPP) based chiral stationary phases (CSPs) provide reduced analysis times while maintaining high column efficiencies and sensitivity. In this study, mobile phase conditions suitable for chiral analyses with electrospray ionization LC-MS were systematically investigated using vancomycin as a representative CSP. The performance of a 2.7⯵m SPP based vancomycin CSP (SPP-V) 10â¯cmâ¯×â¯0.21â¯cm column was compared to that of a corresponding 5⯵m fully porous particles based analogue column. The results demonstrated that the SPP-V column provides higher efficiencies, 2-5â¯time greater sensitivity and shorter analysis time for a set of 22 basic pharmaceutical drugs. The SPP-V was successfully applied for the analysis of the degradation products of racemic citalopram whose enantiomers could be selectively identified by MS.
ABSTRACT
Three phosphonium-based dicationic ionic liquids were synthesized as bis(trifluoromethylsulfonyl) imide salts. The three dications had a nonyl spacer between two identical phosphonium-substituted groups. The three phosphonium moieties were dipropyl(phenyl), diphenyl(propyl), and diphenyl(toluyl). The physicochemical properties of the dicationic ionic liquids were appropriate to prepare 30 m capillary columns that were tested in gas chromatography. A unique selectivity compared to different polysiloxane or polar cyanosiloxane commercial columns was observed for selected mixes of phthalates, polyaromatic hydrocarbons, polychlorobiphenyls, and dioxins. Only minor selectivity variations were observed with the three dicationic ionic liquids. The different number of aromatic rings on the positively charged phosphonium group did not influence the dicationic ionic liquid selectivity significantly.
ABSTRACT
A modified macrocyclic glycopeptide-based chiral stationary phase (CSP), prepared via Edman degradation of vancomycin, was evaluated as a chiral selector for the first time. Its applicability was compared with other macrocyclic glycopeptide-based CSPs: TeicoShell and VancoShell. In addition, another modified macrocyclic glycopeptide-based CSP, NicoShell, was further examined. Initial evaluation was focused on the complementary behavior with these glycopeptides. A screening procedure was used based on previous work for the enantiomeric separation of 50 chiral compounds including amino acids, pesticides, stimulants, and a variety of pharmaceuticals. Fast and efficient chiral separations resulted by using superficially porous (core-shell) particle supports. Overall, the vancomycin Edman degradation product (EDP) resembled TeicoShell with high enantioselectivity for acidic compounds in the polar ionic mode. The simultaneous enantiomeric separation of 5 racemic profens using liquid chromatography-mass spectrometry with EDP was performed in approximately 3 minutes. Other highlights include simultaneous liquid chromatography separations of rac-amphetamine and rac-methamphetamine with VancoShell, rac-pseudoephedrine and rac-ephedrine with NicoShell, and rac-dichlorprop and rac-haloxyfop with TeicoShell.
Subject(s)
Anti-Bacterial Agents/chemistry , Chromatography, High Pressure Liquid/instrumentation , Vancomycin/chemistry , Chromatography, High Pressure Liquid/methods , Mass Spectrometry/methods , Spectrophotometry, Ultraviolet , StereoisomerismABSTRACT
The authors would like to call the reader's attention to the fact that the original publication included some corrections needed to be addressed.
ABSTRACT
The rapid expansion in the study and use of ionic liquids (ILs) is a result of their unique properties including negligible volatility, high thermal stability, and ability to dissolve disparate compounds. However, because ILs have infinitely variable structures (often referred to as "tunability"), these properties can differ considerably. Herein, we focus on the thermal stability of 15 bis-/dicationic ionic liquids. Specifically, their thermal breakdown products are examined to determine the structural linkages, bonds, or atoms most susceptible to thermally induced changes and whether such changes occur before possible volatilization. In most cases, the heteroatom-carbon single bonds were susceptible to thermolytic decomposition. Graphical abstract Capture of dicationic ionic liquid thermal decomposition products for subsequent identification.
ABSTRACT
Twelve bis- or dicationic ionic liquids (ILs) including eight based on imidazolium, a single one based on phosphonium, and three based on pyrrolidinium cationic units were prepared with the bis(trifluoromethyl sulfonyl) imide anion. The two identical cationic moieties were attached by different alkyl spacers having three or five carbons and differing alkyl substituents attached to the spacer. The SLB-IL111 column, as the most polar commercial stationary phase known, was included in the study for comparison. Isothermal separations of a rapeseed oil fatty acid methyl ester (FAME) sample were used to study and compare the 12 IL-based column performances and selectivities. The retention times of the most retained methyl esters of lignoceric (C24:0) and erucic (C22:1) acids were used to estimate the IL polarity. The phosphonium dicationic IL column was, by far, the least polar. Imidazolium-based dicationic IL columns were the most polar. Polarity and selectivity for the FAME separation were somewhat related. The separation of a 37-FAME standard mixture allowed the investigation of selectivity variations observed on the 12 IL-based columns under temperature gradients up to 230 °C. The remarkable selectivity of the IL-based columns is demonstrated by the detailed analysis of the cis/trans C18:1 isomers of a partially hydrogenated vegetable oil sample on 30-m columns, separations competing with that done following an "official method" performed on a 100-m column. Graphical abstract Separation of fatty acid methyl esters on a 30-m 3m2C5(mpy)2. 2NTf2 branched-chain dicationic IL-based column. Branched chain dicationic ILs show great selectivity for separation of cis/trans, ω-3/ω-6, and detailed analysis of cis/trans fats.
ABSTRACT
Current trends in chiral analysis of pharmaceutical drugs are focused on faster separations and higher separation efficiencies. Core-shell or superficially porous particles (SPP) based chiral stationary phases (CSPs) provide reduced analysis times while maintaining high column efficiencies and sensitivity. In this study, mobile phase conditions suitable for chiral analyses with electrospray ionization LC-MS were systematically investigated using vancomycin as a representative CSP. The performance of a 2.7 μm SPP based vancomycin CSP (SPP-V) 10 cm × 0.21 cm column was compared to that of a corresponding 5 μm fully porous particles based analogue column. The results demonstrated that the SPP-V column provides higher efficiencies, 2–5 time greater sensitivity and shorter analysis time for a set of 22 basic pharma-ceutical drugs. The SPP-V was successfully applied for the analysis of the degradation products of racemic citalopram whose enantiomers could be selectively identified by MS.
ABSTRACT
Centrifugal partition chromatography (CPC) is the branch of countercurrent chromatography (CCC) that works with single axis hydrostatic columns with rotary seals. The hydrodynamic of the liquid stationary phase-liquid mobile phase equilibrium in the CPC chambers has been studied theoretically and with specially designed CPC columns. In this work, we selected a simple analytical separation (no loading study) of three test solutes, coccine red, coumarin and carvone, with a commonly used heptane/ethyl acetate/methanol/water 1:1:1:1v/v biphasic liquid system and two different rotors: a commercially available 30-mL CPC instrument and a 80-mL prototype rotor designed for productivity. We fully studied this separation in many possible practical operating conditions of the two rotors, aiming at a generic column characterization. The rotor rotation was varied between 1000 and 2800rpm, the aqueous mobile phase flow rate was varied between 1 and 22mL/min with the 30-mL rotor and 10 and 55mL/min with the 80-mL rotor, the upper limits being mechanical constraints and some liquid stationary phase remaining in the rotor. The variations of Sf, the volume ratio of stationary phase in the rotor, were studied versus mobile phase flow rate and rotor rotation speed. A maximum mobile phase linear velocity was found to depend on the centrifugal field for the 30-mL rotor. This maximum velocity was not observed with the 80-mL rotor. Studying the changes in coumarin and carvone peak efficiencies, it is established that the number of cells required to make one theoretical plate, i.e. one chromatographic exchange, is minimized at maximal rotation speed and, to a lesser extent, at high mobile phase flow rate (or linear velocity). Considering the throughput, there is evidence of an optimal flow rate depending on the rotor rotation that is not necessarily the highest possible.
Subject(s)
Countercurrent Distribution/methods , Countercurrent Distribution/standards , Algorithms , Centrifugation , Coumarins/chemistry , Countercurrent Distribution/instrumentation , Cyclohexane Monoterpenes , Monoterpenes/chemistry , Reproducibility of Results , SolventsABSTRACT
This paper illustrates the application of a recently proposed protocol allowing the scale-up prediction on hydrostatic countercurrent chromatography columns (centrifugal partition chromatographs or CPC). A commercial extract of rosemary (Rosmarinus officinalis L.) was used as the starting material containing 0.48% of carnosol, an active pharmaceutical ingredient with great potential. After a rapid method development on a small-scale 35-mL CPC instrument that allowed for the determination of the solvent system and maximum sample concentration and volume, the purification was transferred on two larger instruments using the "free space between peaks" method. The method takes into account the technical limitations of the larger instruments, such as pressure and/or maximum centrifugal field, and allows, by simply running an analytical-sized injection on the large scale rotor, to give an accurate prediction of the maximum sample load and best throughput. The 0.27g of rosemary extract maximum load on a 35-mL CPC was transferred as a 1.9g load on a 254-mL medium size CPC and 9g load on a 812-mL CPC. The maximum process efficiency of 3.1mg of carnosol per hour obtained on the small 35-mL column was transferred on the 254-mL CPC giving 8.3mg/h and, on the larger 812-mL column 49.4mg of carnosol could be obtained per hour. If the scaling-up in CPC instruments is not directly homothetic, it can be highly predictable through a few simple experiments.