A sensitivity enhanced high-performance liquid chromatography fluorescence method for the detection of nephrotoxic and carcinogenic aristolochic acid in herbal medicines

A new, sensitive and selective HPLC method with fluorescence detector (HPLC-FLD) for the determination of nephrotoxic and carcinogenic aristolochic acid (AA) in herbal medicines by using pre-column derivatization with zinc powder in acetic acid is presented. Variables governing the derivatization reaction, such as the amount of zinc powder and acetic acid, as well as the derivatization time were studied and optimized. An extended linear dynamic range over three orders of magnitude was observed for AA-I and AA-II (R(2)>0.9998). Method accuracy at low, medium and high spiked AA levels determined by the percentage mean deviation was below 4.4% and 7.2% for AA-I and AA-II, respectively. The detection limits of 0.39 ng/mL (AA-I) and 0.52 ng/mL (AA-II) were 2 orders of magnitude lower than those obtained from HPLC-MS or CE-ECD analyses, 3-4 orders of magnitude lower than those from HPLC-UV or CE-UV methods. The developed method has been applied for the determination of AA in herbal medicines. Among the tested samples, Guanmutong had the highest AA concentration (2607.0 microg/g AA-I, 711.2 microg/g AA-II). Comparison studies between HPLC-FLD and HPLC-MS/MS demonstrated that the two methods gave similar quantitative results for the selected herb samples.     

Determination of aristolochic acids in medicinal plant and herbal product by liquid chromatography-electrospray-ion trap mass spectrometry

This paper describes a liquid chromatography-electrospray-ion trap mass spectrometry (LC-ES-ITMS) method for the determination of aristolochic acid I and II (AA-I and AA-II) in medicinal plants and Chinese herbal remedies. A reversed phase C₁₈ column with gradient elution was utilized. The effects of mobile phase additives, acetic acid and ammonium acetate, on LC separation and ES ionization were investigated. For both AA-I and AA-II, the [M+NH₄]⁺ ion was found to be the precursor ion for target MS/MS analysis. The MS/MS product ion, [M+H−44]⁺, was used for the quantitative measurement of AA-I and AA-II. The linearity was good from 0.03 to 5 μg ml⁻¹ and good correlation (r²=0.999) over the range examined was determined for both AA. The detection limit based on a signal-to-noise ratio of three was 0.012 and 0.015 μg ml⁻¹ for AA-I and AA-II, respectively. Various Chinese herbal remedies obtained from renal failure patients and medicinal plants were examined by this newly developed method.     

Differentiation of herbs linked to "Chinese herb nephropathy" from the liquid chromatographic determination of aristolochic acids

A HPLC method was developed and applied to analyze aristolochic acids (AA-I and AA-II) in Chinese medicinal herbs. The herb samples were extracted by using ultrasonication with the extraction efficiency of better than 82%. Extracts were then filtered and injected onto a C18 column eluting under a gradient program using methanol and water-containing 0.5% acetic acid. The method with the detection limits of 1.33 ng for AA-I and 7.29 ng for AA-II per injection was successfully applied for the analysis of traditional Chinese medicine (TCM) and related products and differentiation of Chinese medicinal herbs that have previously been misused and caused toxicological effects. The developed protocol provided an example that analysis of selected component markers could serve for health security and quality control of TCM consumption.     

Study on separation of aristolochic acid I and II by micellar electrokinetic capillary chromatography and competition mechanism between SDS and beta-cyclodextrin

In this study, a rapid MEKC method using 40 mM sodium borate buffer containing 50 mM SDS as surfactant was developed for the analysis of aristolochic acid (AA) in Aristolochia plants. Baseline separation of AA-I and AA-II was achieved within 3 min with high separation efficiency, satisfactory sensitivity, repeatability, and recovery. Resolution between AA-I and AA-II is above 5 and great performance with higher than 200,000 theoretical plate numbers was obtained. The detection limits (based on 3 S/N) were both 1.0 microg/mL. Two kinds of AA in 35 herbal samples of Aristolochia plants were successfully determined. The competition mechanism between beta-CD and SDS was also investigated by changing the content ratio of beta-CD and SDS.     

Determination of aristolochic acid in Chinese herbal medicine by capillary electrophoresis with laser-induced fluorescence detection

We have demonstrated the analysis of aristolochic acids (AAs) that are naturally occurring nephrotoxin and carcinogen by capillary electrophoresis in conjunction with laser-induced fluorescence detection (CE-LIF). Owing to lack of intrinsic fluorescence characteristics of oxidized AAs (OAAs), reduction of the analytes by iron powder in 10.0 mM HCl is required prior to CE analysis. The reduced AAs (RAAs) exhibit fluorescence at 477 nm when excited at 405 nm using a solid-state blue laser. By using 50.0 mM sodium tetraborate (pH 9.0) containing 10.0 mM SDS, the determination of AA-I and AA-II by CE-LIF has been achieved within 12 min. The CE-LIF provides the LODs of 8.2 and 5.4 nM for AA-I and AA-II, respectively. The simple CE-LIF method has been validated by the analysis of 61 Chinese herbal samples. Prior to CE analysis, OAAs were extracted by using 5.0 mL MeOH, and then the extracts were subjected to centrifugation at 3,000 rpm for 5 min. After reduction, extraction, and centrifugation, the supernatants were collected and subjected to CE analysis. Of the 61 samples, 14 samples contain AA-I and AA-II, as well as 10 samples contain either AAI or AAII. The relative standard deviation (RSD) values of the migration times for AA-I and AA-II are less than 2.5% and 2.1% for three consecutive measurements of each sample. The RSD values for the peak heights corresponding to AA-I and AA-II in most samples are about 8.0% and 10.0%, respectively. The result shows that the present CE-LIF approach is sensitive, simple, efficient, and accurate for the determination of AAs in real samples.     

Online concentration of aristolochic acid I and II in Chinese medicine preparations by micellar electrokinetic chromatography

In this study, an online concentration method in micellar electrokinetic chromatography (MEKC) applying field-enhanced sample injection (FESI) mode was developed for the detection of aristolochic acids (AAs) in Chinese medicine preparations. AA-I and AA-II were baseline separated with high separation efficiency, and 100-fold enhancement of the detection sensitivity was achieved compared with those obtained from normal capillary zone electrophoresis (CZE) or simple MEKC method. The proposed method was successfully applied for the determination of AAs in Chinese medicine preparations.     

Rapid determination of aristolochic acid I and II in medicinal plants with high sensitivity by cucurbit[7]uril-modifier capillary zone electrophoresis

Aristolochic acids (AAs) are commonly found in medicinal plants such as Radix aristolochiae and have been reported to cause acute hepatitis and end-stage renal failure. Hence, quantitative analysis and quality control for the plants containing AAs is of great importance. In this study, a novel macrocylcic molecule, cucurbit[7]uril (CB[7]) was employed as a modifier in capillary zone electrophoresis (CZE) for rapid determination of aristolochic acid I and II in medicinal plants. In similarity to other macrocyclic molecules, such as cyclodextrins (CDs), CB[7] can be used to manipulate selectivity in CE because it can form inclusion complexes with a variety of guest molecules. During the running process, CB[7] bears a positive charge in the pH range of 2.5-7.5 and can be adsorbed onto the inner wall of a fused-capillary, leading to a reversal of the electroosmotic flow (EOF). By applying a negative polarity, a rapid separation of AA-I and AA-II was achieved within 7min using 100mM phosphate buffer (pH 7.5) containing 3mM CB[7] and 10% acetonitrile (v/v) as modifiers, due to the same directions of the EOF and the electrophoretic mobilities of the analytes. By applying electrokinetic injection with field-enhanced sample stacking, two kinds of aristolochic acids in four medicinal plants were successfully determined with high sensitivity, high separation efficiency, repeatability and recovery. The proposed method was also used to determine AA-I and AA-II in two slimming pills with complex matrix.     

Analysis of nephroloxic and carcinogenic aristolochic acids in Aristolochia plants by capillary electrophoresis with electrochemical detection at a carbon fiber microdisk electrode

Aristolochic acids (AAs) are the main bioactive ingredients in the most of Aristolochia plants, which are used to make dietary supplements, slimming pills and Traditional Chinese Medicines (TCMs). Excessive ingestion of AAs can lead to serious nephropathy. Therefore, quantitative analysis and quality control for the plants containing AAs is of great importance. In this paper, capillary electrophoresis (CE) with electrochemical detection (ED) at a 33 microm carbon fiber microdisk electrode (CFE) has been applied to detect AA-I and AA-II in Aristolochia plants. Under the optimum conditions: detection potential at 1.20 V, 2.0 x 10(-2) mol L(-1) phosphate buffer solution (PBS) (pH 10.0), injection time 25 s at a height of 17 cm and separation voltage at 12.5 kV, the AA-I and AA-II were baseline separated within 5 min. Low detection limits for AA-I and AA-II were 4.0 x 10(-8) mol L(-1) and 1.0 x 10(-7) mol L(-1), respectively. Wide linear ranges were from 4.0 x 10(-8) mol L(-1) to 1.9 x 10(-5) mol L(-1) and 1.0 x 10(-7) mol L(-1) to 5.0 x 10(-5) mol L(-1) for AA-I and AA-II, respectively. The proposed method has been successfully applied to analyze AAs contents in plant extracts. The results indicated that the contents of AAs in each part of Aristolochia debilis Sieb. Et Zucc. plant were different. Meanwhile, the CE-ED method was utilized for fingerprint analysis of medicine herbs. Six herbs (Radix aristolochiae, Fructus aristolochiae, Herba aristolochiae, Caulis aristolochiae manshuriensis, Caulis clematidis armandii, Caulis akebiae) were well distinguished by comparing their electropherograms obtained by CE-ED method.     

Kinetics of heterogeneous reactions of HO2 radical at ambient concentration levels with (NH4)2SO4 and NaCl aerosol particles

The HO2 uptake coefficient (gamma) for inorganic submicrometer wet and dry aerosol particles ((NH4)2SO4 and NaCl) under ambient conditions (760 Torr and 296 +/- 2 K) was measured using an aerosol flow tube (AFT) coupled with a chemical conversion/laser-induced fluorescence (CC/LIF) technique. The CC/LIF technique enabled experiments to be performed at almost the same HO2 radical concentration as that in the atmosphere. HO2 radicals were injected into the AFT through a vertically movable Pyrex tube. Injector position-dependent profiles of LIF intensity were measured as a function of aerosol concentration. Measured gamma values for dry aerosols of (NH4)2SO4 were 0.04 +/- 0.02 and 0.05 +/- 0.02 at 20% and 45% relative humidity (RH), respectively, while those of NaCl were <0.01 and 0.02 +/- 0.01 at 20% and 53% RH, respectively. For wet (NH4)2SO4 aerosols, measured gamma values were 0.11 +/- 0.03, 0.15 +/- 0.03, 0.17 +/- 0.04, and 0.19 +/- 0.04, at 45%, 55%, 65%, and 75% RH, respectively, whereas for wet NaCl aerosols the values were 0.11 +/- 0.03, 0.09 +/- 0.02, and 0.10 +/- 0.02 for 53%, 63%, and 75% RH, respectively. Wet (NH4)2SO4 and NaCl aerosols doped with CuSO4 showed gamma values of 0.53 +/- 0.12 and 0.65 +/- 0.17, respectively. These results suggest that compositions, RH, and phase for aerosol particles are significant to HO2 uptake. Potential HO2 loss processes and their atmospheric contributions are discussed.     

Aristolochic Acids as a Defensive Substance for the Aristolochiaceous Plant‐Feeding Swallowtail Butterfly,Pachliopta aristolochiae interpositus

The chemical constituents isolated from every stage of the Taiwanese swallowtail butterfly, Pachliopta aristolochiae interpositus, were studied and compared with the constituents of the insect feeding plant, A. cucurbitifolia. Two aristolochic acid derivatives ( 1 and 4 ) and three aristolactam analogues ( 5, 6 , and 7 ) were isolated from the larval osmeterial fluid and larvae of the insect. In addition, four purines ( 2, 3, 13 , and 14 ), one indole alkaloid ( 11 ), one steroid ( 10 ), two benzenoids ( 8 and 9 ) and allantoin ( 12 ) were also isolated from the insect. Among these compounds, aristolochic acid ‐I (AA‐I, 1 ) was detected in all life stages of the insect, especially in the larval osmeterial fluid. The studies confirm that the toxic AAs are used as larval feeding stimulants and deterrent allomones against birds.     

Rapid Determination of Aristolochic Acids I and II in Some Medicinal Plants by High-Performance Liquid Chromatography

Aristolochic acids (AA) are toxic components of Aristolochia plants which result in diseases of the kidney such as urothelial cancer. It is, therefore, essential to monitor the amount of aristolochic acid in herbal medicines. In this study a reversed-phase high-performance liquid-chromatographic (HPLC) method has been developed for rapid determination of aristolochic acids I and II. Baseline separation was achieved within five minutes by use of an ODS C₁₈ column with methanol–water, 60:40, as mobile phase. Two kinds of aristolochic acid were successfully determined in 31 herbal samples of Aristolochia fangchi Wu and Caulis Aristolochiae Manshuriensis. The results indicated that in most samples the aristolochic acid I content is much higher than that of aristolochic acid II. The two kinds of aristolochic acid were not detected in Aristolochia fangchi from the Guangdong region, so Aristolochia fangchi from this region is recommended for use in herbal remedies.     

SUBSTITUENT CHEMICAL SHIFT (SCS) AND THE SEQUENCE STRUCTURE OF ETHYLENE-VINYL ALCOHOL COPOLYMERS

Three ethylene-vinyl alcohol copolymers were studied by means of the substituent chemical shift (SCS) method. The SCS parameters of hydroxy (-OH) in two different solvents were obtained: in deuterium oxide/phenol (20/80 W/W ) the parameters are S_1 = 42.77±0.08ppm, S_2 = 7.15±0.06 ppm, S_3 (s)= -4.08±0.02ppm, S_3 (t) =-3.09±0.20ppm,S_4 = 0.48±0.03ppm, S_5 = 0.26±0.05ppm. In o-dichloro-benzen-d_4 S_1(s)=44.79±0.61ppm, S_2=7.40±0.00ppm, S_3(s)=-4.51±0.17ppm, S_3(t)=-3.13 ±0.00 ppm, S_4 =0.63±0.04ppm, S_5=0.36±0.00ppm.Simultaneously the ~(13)CNMR spectra of EVA copolymers were assigned by using the SCS parameters obtained.     

Simultaneous determination of nine aristolochic acid analogues in medicinal plants and preparations by high-performance liquid chromatography

By optimizing the extraction, separation and analytical conditions, a simple, reliable and effective high-performance liquid chromatography method coupled with photodiode array detector (HPLC-DAD) is presented for simultaneous determination of nine aristolochic acid (AA) analogues, i.e., AA I, AA II, AA C, AA D, 7-OH AA I, aristolic acid, AL II, AL III and AL IV, in twelve medicinal herbs and two preparations. The separation was completed on a C18 column with aqueous methanol containing 0.2% (V/V) acetic acid as mobile phase. Linearities of around two orders of magnitude were obtained with correlation coefficients exceeding 0.9950. Satisfactory intra-day and inter-day precisions were achieved with R.S.D.s less than 4.35%, and the average recovery factors obtained were in the range of 88.4-98.8%. The proposed method appears to be suitable for use as a tool for safety assurance and quality control for commercially available suspect samples containing aristolochic acid analogues.     

Enhancement of photocatalytic activity and regeneration of Fe-doped TiO2 (Ti1−xFexO2) nanocrystalline particles synthesized using inexpensive TiO2 precursor

Research on Chemical Intermediates - Fe-doped TiO2, Ti1?xFexO2 (x?=?0.00, 0.01, 0.03, 0.05, 0.07, and 0.09) photocatalysts with enhanced activity were synthesized via a...     

Synthetic, X-ray Structure, Electron Paramagnetic Resonance, and Magnetic Studies of the Manganese(II) Complex of 1-Thia-4,7-diazacyclononane ([9]aneN(2)S)

Reaction of manganese(II) perchlorate hexahydrate with a methanol solution of 1-thia-4,7-diazacyclononane ([9]aneN(2)S) resulted in the isolation of the manganese(II) complex [Mn([9]aneN(2)S)(2)](ClO(4))(2). The X-ray structure of this complex is reported: crystal system orthorhombic, space group Pbam, No. 55, a = 7.937(2) ?,b = 8.811(2) ?, c = 15.531(3) ?, Z = 2, R = 0.0579. The complex is high spin (S = (5)/(2)) with an effective magnetic moment (&mgr;(eff)) 5.82 &mgr;(B) at 298 K and 5.65 &mgr;(B) at 4.2 K. Computer simulation of the Q-band EPR spectrum of [Mn([9]aneN(2)S)(2)](ClO(4))(2) yields g = 1.99 +/- 0.01, |D| = 0.19 +/- 0.005 cm(-)(1), and E/D = 0.04 +/- 0.02. For the analogous hexaamine complex [Mn([9]aneN(3))(2)](ClO(4))(2) ([9]aneN(3) = 1,4,7-triazacyclononane) analysis of the EPR spectra produced the following values: g = 1.98 +/- 0.01, |D| = 0.09 +/- 0.003 cm(-)(1), and E/D = 0.1 +/- 0.01. The spin Hamiltonian parameters for [Mn([9]aneN(2)S)(2)](ClO(4))(2) derived from the EPR spectra produced a good fit to the magnetic susceptibility data.     

Conformational equilibria of trans -3-X-cyclohexanols (X = Cl, Br, CH3 and OCH3). A low temperature NMR study and theoretical calculations

The integration of 1H and 13C NMR spectra, at - 90 degrees C in CS2/CD2Cl2 (9:1), for the trans-3-chlorocyclohexanol (1), trans-3-bromocyclohexanol (2), and trans-3-methoxycyclohexanol (4) showed that the equatorial-axial (ea) conformer occurs as ca 63, 63, and 69% in the conformational equilibrium, respectively. This corresponds to the following DeltaG(ea-ae) values (from (1)H spectrum): - 0.32 +/- 0.01, - 0.32 +/- 0.04, - 0.48 +/- 0.05 kcal mol(-1); and to (from 13C spectrum): - 0.31 +/- 0.04, - 0.35 +/- 0.05, and - 0.44 +/- 0.01 kcal mol(-1), respectively, in very good agreement within both series. Thus, although bromine is bulkier than chlorine, the 1,3-diaxial steric effects are similar in these equilibria. However, the integration of (1)H NMR spectrum for the trans-3-methylcyclohexanol (3) gave 90% of the 3ae conformer in the equilibrium, at - 90 degrees C on CS2/CD2Cl2 (9:1), corresponding to a DeltaG(ea-ae) value of 1.31 +/- 0.02 kcal mol(-1). The values obtained through the additivity rule, with data from monosubstituted cyclohexanes (DeltaG(Ad) = DeltaG(X) + DeltaG(OH)), for compounds 1, 2, and 4 (-0.37 +/- 0.15, - 0.34 +/- 0.09, and - 0.46 +/- 0.04 kcal mol(-1), respectively) are in very good agreement with the experimental values, but it is significantly smaller for compound 3 (0.79 +/- 0.02 kcal mol(-1)). Theoretical calculations through different levels of theory (HF/6-311 + g**, B3LYP/6-311 + g**, MP2/6-31 + g**, and CBS-4M) showed that CBS-4M is the best method for the study of conformational equilibria for these systems, since it provides DeltaG(ea-ae) values similar to the experimental values.     

Yttrium substituting in Mn site to improve electrochemical kinetics activity of sol-gel synthesized LiMnPO4/C as cathode for lithium ion battery

Journal of Solid State Electrochemistry - LiMn1-x Y x PO4/C (x&nbsp;=&nbsp;0, 0.01, 0.02, 0.04) materials have been prepared by a simple sol-gel method using glycollic acid as chelating...     

GC–MS analysis of phytoconstituents present in Trigonella foenumgraecum L. seeds extract and its antioxidant activity

Trigonella foenumgraecum L. (TF) is a medicinal herb, belonging to the family Legumes. It has shown positive results in remedying hypo-cholesterolemic, anti-inflammatory, antibacterial, antioxidant, anti-lipidemia, antilithigenic, hepatoprotective, antiulcer, anticarcinogenic, antifungal and other miscellaneous pharmacological effects of fenugreek. The n-hexane extract of Trigonella foenumgraecum L. Seeds (TF) was analysed by gas chromatography-mass spectroscopy for identification and characterization of its therapeutic claim by traditional system. DPPH method was used to determine the antioxidant activity of Trigonella foenumgraecum L-seeds extract using UV spectrophotometer at a wavelength of 518 nm as it is one of the most sorted methods for antioxidant activity. The major compounds discovered in Trigonella foenumgraecum L. seeds extract are Linoleic acid (48.01%); 9,12-Octadecadienoic acid (Z,Z)-, 2,3-dihydroxypropyl ester (24.65%); 2-[4-Methyl-6-(2,6,6-trimethylcyclohex-1-enyl)hexa-1,3,5-trienyl]cyclohex-1-en-carboxaldehyde(1.88%); Nonane dioic acid, bis (2-ethylhexyl) ester (1.09%); Bis (2-ethylhexyl) ester of azelaic acid (11.97%); Elemicin (0.51%); cis-Linoleic acid methyl ester (0.76%); Linoleic acid chloride(0.57%); Ethyl oleate(0.18%); Isopropyl linoleate (0.38%); Dihydrovallesiachotamine (0.06%); 4-(2,2-Dimethyl-6-methylenecyclohexyl) butanal(0.12%); Citronellyl myristate (0.09%); Rhaphidecursinol B(0.25%); 5-Methoxygalbelgin (0.07%); Vitamin E (0.31%); 1,1,1,5,5,5-hexafluoro-4-{[3-(trifluoromethyl) phenyl]imino}-2-pentanone (0.76%); γ-Sitosterol (0.66%); 1-(1,5-Dimethylhexyl)-3a,12a-dimethyltetradecahydro-1H-cyclopenta[a]cyclopropa[e]phenanthren-7-ol (0.16%) and (9Z)-9-Octadecenyl (9Z)-9-hexadecenoate (1.57%). The results showed potential antioxidant activity of n-hexane extract of Trigonella foenumgraecum L. seeds by showing significant reduction in free radical against DPPH. The hexane extract of Trigonella foenumgraecum L. seeds comprises various non-water-soluble (nonpolar) constituents. These compounds were established qualitatively via GC-MS evaluation. The free radical scavenging activity of the plant extract was established owning to the presence of compounds such as terpenes, vitamin E, and unsaturated fatty acids.     

Determination of antioxidant additives in foodstuffs by direct measurement of gold nanoparticle formation using resonance light scattering detection

The capability of antioxidant compounds to reduce gold(III) to gold nanoparticles has been kinetically studied in the presence of cetyltrimethylammonium bromide using stopped-flow mixing technique and resonance light scattering as detection system. This study has given rise to a simple and rapid method for the determination of several synthetic and natural antioxidants used as additives in foodstuff samples. The formation of AuNPs was monitored by measuring the initial reaction-rate of the system in about 5s, using an integration time of 0.1s. Dynamic ranges of the calibration graphs and detection limits, obtained with standard solutions of the analytes, were (μmolL?1): gallic acid (0.04-0.59, 0.01), propyl gallate (0.04-1.41, 0.01), octyl gallate (0.03-0.35, 0.08), dodecyl gallate (0.02-0.30, 0.007), butylated hydroxyanisol (0.07-0.39, 0.009), butylated hydroxytoluene (0.04-0.32, 0.01), ascorbic acid (0.11-1.72, 0.03) and sodium citrate (0.07-1.29, 0.02). The regression coefficients were higher than 0.994 in all instances. The precision of the method, expressed as RSD%, was established at two concentration levels of each analyte, with values ranging between 0.6 and 4.8%. The practical usefulness of the developed method was demonstrated by the determination of several antioxidant additives in foodstuff samples, which were extracted, appropriately diluted and assayed, obtaining recoveries between 95.4 and 99.5%. The results obtained were validated using two reference methods.     

Paper electrophoretic technique in the study of metal complexes

A new method, paper electrophoresis, involving the use of a ionophoretic technique is described for the study of equilibria in binary complex system in solution. The stability constants of ML and ML₂ complex species of metal(II)-α-aminobutyric acid and metal(II) — homoserine were found to be [(8.09 ± 0.03, 6.89 ± 0.09) (3.58 ± 0.07, 2.67 ± 0.11) (7.66 ± 0.06, 6.13 ± 0.02)]; [(7.80 ± 0.07, 6.45 ± 0.02) (2.93 ± 0.04, 1.97 ± 0.01) (7.41 ± 0.011, 4.67 ± 0.06)] for copper(II), manganese(II) and uranyl(II) complexes, respectively at an ionic strength 0.1 M and 35°C. Published in Russian in Zhurnal Neorganicheskoi Khimii, 2009, Vol. 54, No. 7, pp. 1196–1199. The article is published in the original.