Discrimination of Ganoderma lucidum according to geographical origin with near infrared diffuse reflectance spectroscopy and pattern recognition techniques

A rapid and nondestructive near infrared (NIR) method combined with chemometrics was used to discriminate Ganoderma lucidum according to cultivation area. Raw, first, and second derivative NIR spectra were compared to develop a robust classification rule. The chemical properties of G. lucidum samples were also investigated to find out the difference between samples from six varied origins. It could be found that the amount of polysaccharides and triterpenoid saponins in G. lucidum samples was considerably different based on cultivation area. These differences make NIR spectroscopic method viable. Principal component analysis (PCA), discriminant partial least-squares (DPLS) and discriminant analysis (DA) were applied to classify the geographical origins of those samples. The results showed that excellent classification could be obtained after optimizing spectral pre-treatment. For the discriminating of samples from three different provinces, DPLS provided 100% correct classifications. Moreover, for samples from six different locations, the correct classifications of the calibration as well as the validation data set were 96.6% using the DA method after the SNV first derivative spectral pre-treatment. Overall, NIR diffuse reflectance spectroscopy using pattern recognition was shown to have significant potential as a rapid and accurate method for the identification of herbal medicines.     

Metabolite profiling on apple volatile content based on solid phase microextraction and gas-chromatography time of flight mass spectrometry

A headspace SPME GC-TOF-MS method was developed for the acquisition of metabolite profiles of apple volatiles. As a first step, an experimental design was applied to find out the most appropriate conditions for the extraction of apple volatile compounds by SPME. The selected SPME method was applied in profiling of four different apple varieties by GC-EI-TOF-MS. Full scan GC-MS data were processed by MarkerLynx software for peak picking, normalisation, alignment and feature extraction. Advanced chemometric/statistical techniques (PCA and PLS-DA) were used to explore data and extract useful information. Characteristic markers of each variety were successively identified using the NIST library thus providing useful information for variety classification. The developed HS-SPME sampling method is fully automated and proved useful in obtaining the fingerprint of the volatile content of the fruit. The described analytical protocol can aid in further studies of the apple metabolome.     

Evaluation of traditional Chinese herbal medicine: Chaihu (Bupleuri Radix) by both high-performance liquid chromatographic and high-performance thin-layer chromatographic fingerprint and chemometric analysis

Chaihu (Bupleuri Radix), roots of Bupleurum chinense and B. scorzonerifolium, is an authentic Chinese Materia Medica in the Chinese Pharmacopoeia. Some other species such as the roots of B. falcatum, B.bicaule and B. marginatum var. stenophyllum similar to Chaihu can also be occasionally found in local raw herb markets. The quality of 33 lots of authenticated Chaihu samples vs. 31 lots of commercial samples was evaluated by both high-performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD) and high-performance thin-layer chromatography (HPTLC) analyses of its principal bioactive components (saikosaponins). The pre-treated data acquired from both HPLC fingerprints and HPTLC fluorescent images were processed by chemometrics for similarity and pattern recognition, including Artificial Neural Networks (ANNs), k-nearest neighbor (k-NN) and an expert’s panel. It was apparent that k-NN classifier exhibited good performance with sufficient flexibility for processing HPTLC fingerprint images which were otherwise not easily dealt with by other algorithms due to the shift of R_f values and varying hue/saturation of the band colours between different TLC plates. These two chromatographic fingerprint methods can be considered complementary measure of quality control. The roots of Chaihu from different species of the genus Bupleurum could readily be distinguished from each other so that commercial samples can easily be classified. Chaihu collected from several major herbal distribution centers was found to belong to B. chinense with great variation in the content of its major saikosaponins.     

Application of ionic liquids based microwave-assisted simultaneous extraction of carnosic acid, rosmarinic acid and essential oil from Rosmarinus officinalis

An ionic liquid based microwave-assisted simultaneous extraction and distillation (ILMSED) method has been developed for the effective extraction of carnosic acid (CA), rosmarinic acid (RA) and essential oil (EO) from Rosmarinus officinalis. A series of 1-alkyl-3-methylimidazolium ionic liquids differing in composition of anion and cation were evaluated for extraction yield in this work. The results obtained indicated that the anions and cations of ionic liquids had influences on the extraction of CA and RA, 1.0M 1-octyl-3-methylimidazolium bromide ([C8mim]Br) solution was selected as solvent. In addition, the ILMSED procedures for the three target ingredients were optimized and compared with other conventional extraction techniques. ILMSED gave the best result due to the highest extraction yield within the shortest extraction time for CA and RA. The novel process developed offered advantages in term of yield and selectivity of EO and shorter isolation time (20 min in comparison of 4h of hydrodistillation), and provides a more valuable EO (with high amount of oxygenated compounds). The microstructures and chemical structures of rosemary samples before and after extraction were also investigated. Moreover, the proposed method was validated by the stability, repeatability and recovery experiments. The results indicated that the developed ILMSED method provided a good alternative for the both extraction of non-volatile compounds (CA and RA) and EO from rosemary as well as other herbs.     

Ultra high performance liquid chromatography-time of flight mass spectrometry for analysis of avocado fruit metabolites: method evaluation and applicability to the analysis of ripening degrees

We have developed an analytical method using UHPLC-UV/ESI-TOF MS for the comprehensive profiling of the metabolites found in the methanolic extracts of 13 different varieties of avocado at two different ripening degrees. Both chromatographic and detection parameters were optimized in order to maximize the number of compounds detected and the sensitivity. After achieving the optimum conditions, we performed a complete analytical validation of the method with respect to its linearity, sensitivity, precision, accuracy and possible matrix effects. The LODs ranged from 1.64 to 730.54 ppb (in negative polarity) for benzoic acid and chrysin, respectively, whilst they were found within the range from 0.51 to 310.23 ppb in positive polarity. The RSDs for repeatability test did not exceed 7.01% and the accuracy ranged from 97.2% to 102.0%. Our method was then applied to the analysis of real avocado samples and advanced data processing and multivariate statistical analysis (PCA, PLS-DA) were carried out to discriminate/classify the examined avocado varieties. About 200 compounds belonging to various structural classes were tentatively identified; we are certain about the identity of around 60 compounds, 20 of which have been quantified in terms of their own commercially available standard.