Paclitaxel , extraction

F.14 Paclitaxel, which is extracted from the Pacific yew tree Taxus brevifolia, has antitumor activity for ovarian and breast cancer. It is sold under the trade name Taxol. On analysis, its mass percentage composition is 66.11% C, 6.02% H, and 1.64% N, with the balance being oxygen. What is the empirical formula of paclitaxel ... 

Systems with different selectivity were nsed for the separation of 10-deacetyl-baccatin III (10 DAB 111) from yew extracts [69]. A silica column with stepwise gradient elution with aqneous methanolic mobile phases can be nsed for separation of the taxoid fraction from nonpolar materials, partial separation of the taxoid fraction into a polar one (containing 10-DAB 111), and for a medinm polarity taxoid fraction (containing paclitaxel and cephalomannine). Most polar material (tannins... 

Schellen A. et al., 2000. High throughput online solid phase extraction/tandem mass spectrometric determination of paclitaxel in human serum. Rapid Commun Mass Spectrom 14 230. 

The advantages of MAE are short extraction times (10 min), extraction of many samples at one time (up to 14, depending on the system), and less organic solvent consumed. In one recent study [29], MAE was used to extract paclitaxel from Iranian yew trees. The needles of the tree were air-dried and ground. The needles were covered with methanol-water and placed in the MAE apparatus. Extractions took 9-16 min. The extracts were filtered and analyzed by HPLC. Further optimization of the method resulted in less than 10% RSDs for precision and greater than 87% recovery. The overall benefits of the MAE method are reduced extraction times (15-20 min versus 17 h), minimal sample handling, and 75-80% reduction in solvent consumption [29]. 

Paclitaxel (Taxol, Bristol-Myers Squibb) is a chemotherapy drug for ovarian cancer, breast cancer, and certain lung cancers. It was discovered by the US National Cancer Institute in the 1960s. Originally, it was extracted from the bark of the North American yew tree (Taxus brevifo-lia). Clinical tests had necessitated the harvesting of the bark, and this method damaged the trees irreversibly. 

The NCI scientists discovered that extracts of the bark and needles of a yew tree, Taxus brevifolia of the Pacific Northwest, killed trunor cells. Fresh samples were obtained from the forests in the state of Washington in August 1962. Paclitaxel was first isolated from yew tree extract in 1967, and retested on cells in the laboratory. After it was found to be effective in tests on animals with tmnors, paclitaxel was studied in a large munber of human cancer patients and finally approved by the Food and Drug Administration (FDA) in 1992 for use in treating cancer in people. 

Paclitaxel was first manufactured by extracting the active drug from the bark of the Pacific yew tree, but that approach was imac-ceptable, both for the manufacturer, Bristol-Myers Squibb, and for... 

Historically, plants and other living creatures have been used as medical treatments, and early chemically produced drugs were based on or identical to compounds extracted from natural sources. Nature continues to provide modern drugs, and searches stiU begin with systematic laboratory testing or by following clues from traditional folk healers. Modern chemical methods are used to improve upon what is foimd in nature. Some of these drugs, such as paclitaxel, provide major improvements in medical care. 

A brilliant example for the industrial-scale application of plant cell fermentation is the new process for the production of the anticancer drug paclitaxel developed by Bristol-Myers Squibb (see Figure 15.1). It starts with clusters of paclitaxel producing cells from the needles of the Chinese yew, T. chinensis, and was introduced in 2002. The API is isolated from the fermentation broth and is purified by chromatography and crystallization. The new process substitutes the previously used semisynthetic route. It started with lO-deacetylbaccatin(III), a compound that contains most of the structural complexity of paclitaxel and can be extracted from leaves and twigs of the European yew, T. baccata. The chemical process to convert 10-deacetylbaccatin(III) to paclitaxel is complex. It includes 11 synthetic steps and has a modest yield. 

The taxanes are a group of polycyclic diterpenes produced by various species of trees (such as yew) and are most commonly known as the potent anticancer drug paclitaxel (Taxol). The extraction process is costly. 

Allwood, M. C, and Martin, H. (1996),The extraction of diethylhexylphthalate (DEHP) from polyvinyl chloride components of intravenous infusion containers and administration sets by paclitaxel injection, Int. J. Pharm., 127, 65-71. 

The crude paclitaxel is recovered from the rich aqueous fermentation broth by liquid/liquid extraction with a mixture of isobutyl acetate (IBA) and isopropanol (IPA), both class 3 solvents. The waste aqueous phase is stripped to remove residual organic solvents (IBA/IPA), treated with sodium hydroxide to deactivate any paclitaxel residues, and processed through a standard wastewater treatment facility. The amount of solid waste biomass generated in the process is negligible. 

The crude paclitaxel is dissolved in dichloromethane, filtered, and solvent-exchanged into a mixture of dimethylformamide (DMF) and formamide. This solution is loaded onto a chromatography column and eluted with acetonitrile/ water as the mobile phase. The acceptably pure fractions are combined and concentrated to remove acetonitrile, and the paclitaxel is extracted into dichloromethane. Finally, the dichloromethane is replaced with IPA (distillative exchange) and paclitaxel API is isolated by crystallization. 

The more recently discovered Taxol (paclitaxel) was extracted from the bark of the Pacific yew.f It stabilizes microtubules, inhibiting their disassembly.s Taxol also blocks mitosis and causes the cells which fail to complete mitosis to die. Taxol has been syn-thesizedf and is a promising drug that is being used... 

In order to extract paclitaxel and docetaxel (the I.S.) selectively from cell lysates, while simultaneously reducing the quantity of matrix components that were also derived from the samples, the SPE wash and elution conditions were optimized in a manner similar as these described in the paradigm 1. Detailed experimental conditions can be found in a previous publication [5],... 

Fig. 8 Optimization of solvent composition for maximizing absolute recovery of paclitaxel after SPE extraction. Solvents used in the elution step were composed of 0.1 % formic acid in the indicated solvent composition, (a) Optimization of the strong-wash solvent triplicate SPE cartridges were washed with a range of 55-95 % methanol drug was eluted from all cartridges with 100%meth-anol and (b) optimization of the elution solvent triplicate SPE cartridges were washed with 5 % methanol and eluted with methanol concentrations ranging from 60 to 100 % (Reproduced with permission from Elsevier)...

Fig. 9 Optimization of sample injection volume. Various injection volumes (VL) of spiked cell lysate samples were investigated to optimize VL. Lysates were spiked with 250 pg/mL of paclitaxel and extracted by selective SPE. The VL was (a) 0.1 pL (b) 0.5 pL (c) 2.0 pL (d) 8.0 pL. A 15 cm x 0.5 mm I.D. capillary column was used for separation the manufacturer-recommended Vnj for the column was 0.1-0.2 pL (Reproduced with permission from Elsevier)...

Elicitation is effective in enhancing metabolite synthesis in some cases, such as in production of paclitaxel by Taxus cell suspension cultures [2] and tropane alkaloid production by suspension cultures of Datura stramonium [3]. Increasing the activity of metabolic pathways by elicitation, in conjunction with end-product removal and accumulation in an extractive phase, has proven to be a very successful strategy for increasing metabolite productivity [4]. For example, two-phase operation with elicitation-enhanced alkaloid production in cell suspension cultures of Escherichia californica [5,6]. 

The tubulin-binding properties of (-)-rhazinilam were discovered through screening of a number of Malaysian plant extracts [60]. Natural (-)-rhazinilam induces tubulin spiralization, inhibiting tubulin assembly in the same way as vinblastine-like alkaloids, and protects microtubules from cold disassembly such as with paclitaxel [67]. This effect has never been observed with other microtubule poisons. For this reason, and despite the in vivo inactivity of (-)-rhazinilam [67], a number of analogues have been prepared by semi-synthesis and total synthesis (see Sections 3.1.3. and 3.2.3.) in order to improve the pharmacological properties of this molecule. 

---