No abstract

Current clinical strategy for staging and prognostication of colorectal cancer (CRC) relies mainly upon the TNM or Duke system. This clinicopathological stage is a crude prognostic guide because it reflects in part the delay in diagnosis in the case of an advanced cancer and gives little insight into the biological characteristics of the tumor. We hypothesized that global metabolic profiling (metabonomics/metabolomics) of colon mucosae would define metabolic signatures that not only discriminate malignant from normal mucosae, but also could distinguish the anatomical and clinicopathological characteristics of CRC. We applied both high-resolution magic angle spinning nuclear magnetic resonance (HR-MAS NMR) and gas chromatography mass spectrometry (GC/MS) to analyze metabolites in biopsied colorectal tumors and their matched normal mucosae obtained from 31 CRC patients. Orthogonal partial least-squares discriminant analysis (OPLS-DA) models generated from metabolic profiles obtained by both analytical approaches could robustly discriminate normal from malignant samples (Q(2) > 0.50, Receiver Operator Characteristic (ROC) AUC >0.95, using 7-fold cross validation). A total of 31 marker metabolites were identified using the two analytical platforms. The majority of these metabolites were associated with expected metabolic perturbations in CRC including elevated tissue hypoxia, glycolysis, nucleotide biosynthesis, lipid metabolism, inflammation and steroid metabolism. OPLS-DA models showed that the metabolite profiles obtained via HR-MAS NMR could further differentiate colon from rectal cancers (Q(2)> 0.60, ROC AUC = 1.00, using 7-fold cross validation). These data suggest that metabolic profiling of CRC mucosae could provide new phenotypic biomarkers for CRC management.

The lipotoxicity hypothesis posits that obesity predisposes individuals to metabolic diseases because the oversupply of lipids to tissues not suited for fat storage leads to the accumulation of fat-derived molecules that impair tissue function. Means of combating this have been to stimulate anabolic processes to promote lipid storage or to promote catabolic ones to drive fat degradation. Herein, we demonstrate that ablating dihydroceramide desaturase 1 (Des1), an enzyme that produces ceramides, leads to the simultaneous activation of both anabolic and catabolic signaling pathways. In cells lacking Des1, the most common sphingolipids were replaced with dihydro forms lacking the double bond inserted by Des1. These cells exhibited a remarkably strong activation of the antiapoptotic and anabolic signaling pathway regulated by Akt/protein kinase B (PKB), were resistant to apoptosis, and were considerably larger than their wild-type counterparts. Paradoxically, Des1؊/؊ cells exhibited high levels of autophagy. Mechanistic studies revealed that this resulted from impaired ATP synthesis due in part to decreased expression and activity of several complexes of the electron transport chain, particularly complex IV, leading to activation of AMP-activated protein kinase and its induction of the autophagosome. Thus, Des1 ablation enhanced starvation responses but dissociated them from the anabolic, prosurvival, and antiautophagic Akt/PKB pathways.

The in vitro acetylcholinesterase (AChE) inhibitory potential of the hydroalcoholic extract and of the essential oil from Acorus calamus (AC) rhizomes and that of its major constituents were evaluated based on the Ellman's method. GC/MS analysis of the oil revealed that the major constituents were beta-asarone (79.54%) and alpha-asarone (8.47%). The IC50 values were obtained for the hydroalcoholic extract, the essential oil, beta-asarone and alpha-asarone and were 182.31+/-16.78 microg/mL, 10.67+/-0.81 microg/mL, 3.33+/-0.02 microM and 46.38+/-2.69 microM, respectively. Physostigmine was used as standard inhibitor with an IC50 value of 0.28+/-0.015 microM. The experimental observations revealed that the AC essential oil and its constituents have significant AChE inhibitory potential. beta-Asarone, the major phytoconstituent present in the essential oil, showed the maximum inhibitory potential.

Acorus calamus Linn. (Araceae), commonly known as ''sweet flag'' or ''calamus'', is a semiaquatic, perennial, aromatic herb with creeping rhizomes. The plant is found in the northern temperate and subtropical regions of Asia, North America, and Europe. The plant exhibits polyploidy. Many ethnomedicinal and ethnobotanical uses have been ascribed to the rhizomes of the plant. A. calamus Linn. (AC) has been used as traditional Chinese and Indian prescriptions for its beneficial effects on memory disorder, learning performance, lipid peroxide content, and anti-aging and anticholinergic activity. Moreover, pharmacological studies have revealed that Acorus rhizome and its constituents, particularly a-and b-asarone, possess a wide range of pharmacological activities such as sedative, CNS depressant, behavior modifying, anticonvulsant, acetylcholinesterase inhibitory, memory enhancing, anti-inflammatory, antioxidant, antispasmodic, cardiovascular, hypolipidemic, immunosuppressive, cytoprotective, antidiarrheal, antimicrobial, anthelmintic, insecticidal, adulticidal, diuretic, antioxidant, genotoxic, and mutagenic activities. This review is an effort to explore the different phytoconstituents and pharmacological activities of Acorus calamus.

Colorectal cancer (CRC) is the fourth most common cause of death from cancer in the world. The limitations of the currently available methods and biomarkers for CRC management highlight the necessity of finding novel markers. Metabonomics can be used to search for potential markers that can provide molecular insight into human CRC. The emergence of two-dimensional gas chromatography time of flight mass spectrometry (GC × GC/TOFMS) has comprehensively enhanced the metabolic space coverage of conventional GC/MS. In this study, a GC × GC/TOFMS was developed for the tissue-based global metabonomic profiling of CRC. A Pegasus GC × GC/TOFMS (Leco Corp., St. Joseph, MI, USA) system comprising an Agilent 7890 GC and Pegasus IV TOFMS was used for this purpose. An Agilent DB-1 (30 m × 250 μm × 0.25 μm) fused silica capillary column and a Restek Rxi®-17 (1 m × 100 μm × 0.10 μm) fused silica capillary column were used as the primary and secondary columns, respectively. The method was applied for global metabonomic profiling of matched CRC and normal tissues (n = 63) obtained from 31 CRC patients during surgery. An attempt was also made to compare GC × GC/TOFMS with GC/MS and NMR in similar application. The results showed that the metabotype associated with CRC is distinct from that of normal tissue and led to the identification of chemically diverse marker metabolites. Metabolic pathway mapping suggested deregulation of various biochemical processes such as glycolysis, Krebs cycle, osmoregulation, steroid biosynthesis, eicosanoid biosynthesis, bile acid biosynthesis, lipid, amino acid and nucleotide metabolism.

A sensitive and accurate High-Performance TLC (HPTLC) method has been developed to determine the quantity of 6-gingerol in rhizomes of Zingiber officinale (family: Zingiberaceae), commonly known as ginger. Methanol extracts of rhizomes from three different sources were used for HPTLC, n-hexane, and diethyl ether (40:60 v/v) as the mobile phase. The Rf of 6-gingerol was found to be 0.40. The calibration plot was linear in the range of 250-1200 ng of 6-gingerol and the correlation coefficient of 0.9997 was indicative of good linear dependence of peak area on concentration. The mean quantity of 6-gingerol was found to be 60.44+/-2.53 mg/g of ginger extract. The method permits reliable quantification of 6-gingerol and good resolution and separation of 6-gingerol from other constituents of ginger. To study the accuracy and precision of the method, recovery studies were performed by the method of standard addition. Recovery values from 99.79 to 99.84% showed the excellent reliability and reproducibility of the method. The proposed HPTLC method for quantitative monitoring of 6-gingerol in ginger can be used for routine quality testing of ginger extracts.

In this study, a gas chromatography/mass spectrometry (GC/MS) method was developed and validated for the metabolic profiling of human colon tissue. Each colon tissue sample (20 mg) was ultra-sonicated with 1 mL of a mixture of chloroform/methanol/water in the ratio of 20:50:20 (v/v/v), followed by centrifugation, collection of supernatant, drying, removal of moisture using anhydrous toluene and finally derivatization using N-methyl-N-trifluoroacetamide (MSTFA) with 1% trimethylchlorosilane (TMCS). A volume of 1 microL of the derivatized mixture was injected into the GC/MS system. A total of 53 endogenous metabolites were separated and identified in the GC/MS chromatogram, all of which were selected to evaluate the sample stability and precision of the method. Of the identified endogenous metabolites 19 belonging to diverse chemical classes and covering a wide range of the GC retention times (Rt) were selected to investigate the quantitative linearity of the method. The developed GC/MS method demonstrated good reproducibility with intra- and inter-day precision within relative standard deviation (RSD) of +/-15%. The metabolic profiles of the intact tissue were determined to be stable (100 +/- 15%) for up to 90 days at -80 degrees C. Satisfactory results were also obtained in the case of other stability-indicating studies such as freeze/thaw cycle stability, bench-top stability and autosampler stability. The developed method showed a good linear response for each of the 19 analytes tested (r(2) > 0.99). Our GC/MS metabolic profiling method was successfully applied to discriminate biopsied colorectal cancer (CRC) tissue from their matched normal tissue obtained from six CRC patients using orthogonal partial least-squares discriminant analysis [two latent variables, R(2)Y = 0.977 and Q(2) (cumulative) = 0.877].