e-book NMR Data for Carbon-13. Part 4: Natural Products

Free download. Book file PDF easily for everyone and every device. You can download and read online NMR Data for Carbon-13. Part 4: Natural Products file PDF Book only if you are registered here. And also you can download or read online all Book PDF file that related with NMR Data for Carbon-13. Part 4: Natural Products book. Happy reading NMR Data for Carbon-13. Part 4: Natural Products Bookeveryone. Download file Free Book PDF NMR Data for Carbon-13. Part 4: Natural Products at Complete PDF Library. This Book have some digital formats such us :paperbook, ebook, kindle, epub, fb2 and another formats. Here is The CompletePDF Book Library. It's free to register here to get Book file PDF NMR Data for Carbon-13. Part 4: Natural Products Pocket Guide.

Hence the data set represents the upper limit of correlations that may be experimentally available for the constitution. All molecules were drawn in the "Alternative Constitutions" module and submitted to the server. The number of solutions suggested for Ascomycin 1 and Oroidin 2 in runs with theoretical and experimental data are shown in table 1. Ascomycin 1, Oroidin 2 and Aflatoxin B1 3 are used to evaluate the use of theoretical data. Ascomycin 1 is a well known ethyl derivative of Tacrolimus, it serves as example of a large natural product, featuring 43 Carbon atoms.

Using experimental COSY and 13 C-HMBC correlation data the structure generator comes up with structural assignments, which are reduced to one when the atom types are fixed as well. In this case NMR correlation data was able to define the constitution unambiguously. The use of theoretical COSY and 13 C-HMBC correlations leads to a total of 16 possible constitutional assignments, also predefining the atom types reduces this set to one constitutional assignment.

The experimental data set leads to , structural assignments generated, which reduce to 1, when atom types are predefined as well. Hence the structure can not be safely determined by NMR alone. The original structure determination was carried out by chemical derivatization and total synthesis [ 25 , 26 ]. The pictures change with Aflatoxin B1 3 with 17 Carbon atoms. When the atom types are predefined, COCON generates 55 constitutional assignments, compared to with experimental data. The molecule set generated contains constitutions with the element cyclobutadiene, a structural element that is very uncommon in natural products.

COCON has several built-in rules that eliminate certain constitutional elements, like cyclobutadiene, cyclopropene and peroxides. By default these rules are not used, but in this special case we observed a substantial difference in the number of results. When these rules are activated the number of solutions drops to 58 for the experimental correlation data set and 33 for the theoretical data set. All planar molecules suggested are shown in Figure 2 , the correct constitution and starting point of the analysis is 6.

For the small number of interesting constitutions a back-calculation on the carbon chemical shifts was made ChemDraw v11 , that were compared to the experimental values see table 2. The last line in the table contains the sum of the absolute chemical shift differences for all carbons, exposing molecule 6 as the one that best fits the experimental data [ 24 , 27 , 28 ]. Planar constitutions suggested for Aflatoxin B1.

Suggestions 4 - 6 are obtained using theoretical data, 5 - 10 using experimental data. Constitution 6 is the correct one. The theoretical NMR correlation dataset is the upper limit of number of correlations that are possible with a given constitution. Therefore all alternative constitutions generated with this data are "NMR-identical" with regard to correlation data.

A careful analysis of this alternatives might be used to direct further investigations needed to confirm the proposed constitution. The results obtained would direct further work towards chemical derivatization and synthesis [ 25 , 26 ] or x-ray crystallography. The results obtained for Aflatoxin B1 show nicely how carbon chemical shift prediction can be used as tool for the structure discussion, exposing one suggested constitutional assignment as best fitting. Org Lett. Prog Nucl Mag Res Sp. Magn Reson Chem. J Mol Model.

Quim Nova. J Chem Inf Model. Steinbeck C: Recent developments in automated structure elucidation of natural products. Nat Prod Rep. J Chem Inf Comp Sci. Abstr Pap Am Chem S. Internet J Chem. Angew Chem Int Edit. Anal Chim Acta. J Syn Org Chem Jpn. J Mol Struct. J Magn Reson Ser A. Eur J Org Chem. J Chem Soc Chem Comm. Samples S and R are two different candidates of known structure that are compared to a natural product NP of unknown structure. To the extent possible, the resonances of the spectra are assigned to their associated carbon atoms by standard means.

The spectra of the synthetic samples must be recorded under the same conditions and at the same nominal probe temperature within about 1 K. The temperature of the NP spectrum is critical but does not need to be known in advance because it emerges from the analysis. Step 1 is to compare the spectra of the candidate samples R and S not to the natural product but to each other. The goals are to determine whether the candidate spectra are the same or different and, if different, to articulate the differences.

Start by subtracting the chemical shifts of R from S or the reverse and gauge the experimental uncertainty in the chemical shift values. Record duplicate spectra if needed to estimate this. If the subtractions are all zero within the expected experimental error, then the spectra are substantially identical. This means that the two candidate spectra cannot be used to assign the configuration of NP.

Further subtraction of S and R from NP is pointless, and another means of assignment is needed. If some subtractions are not zero, then group all resonances into three categories based on their differences in their chemical shifts.

The chemical shifts of each pair of resonances in S and R are either 1 the same, 2 uncertain, or 3 reliably different. Establish chemical shift limits for the three categories based on the estimated error and common sense analysis of the data. Before advancing to step 2, check the carbon atom assignments of the various resonances, which should pass the sniff test of chemical common sense. In steps 2 and 3, we compare the spectra of the candidate samples to that of the NP. This is where standard comparisons usually start; however, we are ahead of the game because we already know that our comparisons are meaningful that is, S and R do not have identical spectra and we know what to compare the reliably different resonances.

In step 2, we control for temperature and calibration differences of the candidate and natural samples. In principle, the subtractions should all be zero. In practice, there are three possible outcomes: First, if all of the values are zero or close to zero , then both the temperature and calibration errors are small. In that case, proceed directly to step 3 final comparison.

Second, if the values are small and constant either positive or negative , then there is a calibration difference between the synthetic and natural samples. Temperature effects on 13 C NMR spectra of alkanes are variable, ranging from 0 to about 20 ppb per degree K in either direction that is, upfield or downfield with increasing T , reflecting the change in conformation populations with temperature. Clearly the spectra of the candidate samples have to be collected at the same sample temperature as the spectrum of the natural product. What to do if you identify a temperature difference between your synthetic samples sample temperature known and the natural product sample sample temperature unknown?

This process will be illustrated below. If all of the subtraction values equal zero within the experimental uncertainty, then this is the natural product match. If all subtraction values equal the reliably different values identified in step 1, then this is the mismatch. No other result is possible; there cannot be subtractions that neither match zero nor mismatch reliably different. First, Kitching provided the spectrum of the isolated natural product and assigned the various resonances. This proves a key assumption of our method, that at least some of the carbons in the isomers have identical chemical shifts.

Breit concluded that assigning the configuration of 3 by the usual raw subtraction of chemical shifts of synthetic and natural samples was not possible because the spectra of 1 and 3 were too similar. The 16 R ,18 S configuration 1 was assigned to the natural product because no peaks were doubled in the spectrum of the sample of 1 with the natural product capillary inserted, whereas a lone peak was doubled in the corresponding spectrum of 3 with the capillary. The tube-in-tube experiments suggest that the stereoisomers 1 and 3 have 27 chemical shifts that are the same and only one that is significantly different; however, we show presently that more differences can be identified.

Together, the data sets provide an extremely rare perhaps the only case study where spectra of pairs of diastereomers with similar spectra have been collected in three different ways: as individual, pure samples Breit , truly mixed Kitching, but admixed with two other isomers , and artificially mixed Breit, tube-in-tube. Our goal was to do what was not possible by raw subtraction: to confidently assign the structure of the hexamethyldocosane natural product from the data sets of the individual samples alone.

Table 1 shows his complete data lists 2 along with the subtraction results. Breit is correct—the spectra are strikingly similar. Chemical shift differences in ppb and assigned carbon atoms on the structures of the hexamethyldocosane top and pentamethyldocosane bottom candidate isomers. The uncertain resonances are labeled in green, and reliably different resonances are in red. Notice how the reliably different resonances are not randomly distributed or clustered at the ends.

Instead, they cluster in the region C9—C This makes sense given the structural difference between the two isomers. Still, the six reliably different resonances suffice for the analysis. Strikingly, most of the uncertain and reliably different resonances in the hexamethyldocosane isomers 1 and 3 belong to methylene carbons. In contrast, only one of the six methines C10 RD class and one of the six methyl groups not shown because it was not assigned, UC class lights up in this analysis.

These and the remaining subtraction tables are shown in the Supporting Information. The result is a small, constant difference of about 3 ppb. The 4. Without question, the hexamethyldocosane is 1. Specifically, are the reliably different resonances identified by this method real?

The last column of Table 2 lists the ppb differences in the doubled resonances in Kitching mixture sample. The magnitudes of the differences match within about 2 ppb. This cannot be coincidence. This simply shows that having more reliably different resonances increases the confidence of the method. It is not important to find all of the real differences, only that the differences found are reliable. This success was enabled in no small part by the high quality of the published data sets from both Breit and Kitching.

Breit has already assigned 1 securely by the tube-in-tube method, so here it is the validation of the method that counts. Strikingly, the new method of controlled subtraction of the data was comparable in effectiveness to recording spectra of true mixtures in identifying different doubled resonances and was considerably better than the tube-in-tube method. We speculate that this may be due to the poorer field inhomogeneity inevitable with two tubes. Regardless, the results show in hindsight that no mixing of the candidate and natural samples either real or artificial is needed.

By careful, controlled comparison of published data sets, we could identify differences in chemical shift at the low ppb level, close to the experimental level of resolution. The upshot is a secure structure confirmation without access to a natural sample and without new experiments.

The Manchester group has recently communicated the synthesis of the candidate isomers 2 and 4 of the pentamethyldocosane natural product. Complete experimental information and compound characterization data are found in the Supporting Information. The two epimers 2 and 4 were made convergently with a late stage formation of the C 11 —C 12 bond. The synthesis of the requisite C 1 —C 11 fragment is based on chemistry developed for the stereoselective synthesis of hydrocarbons with methyl substituents disposed at 1,3,5-positions down an aliphatic chain.

Such reactions are selective for the formation of E -1,5- anti -products when mediated by the low valence bismuth species formed by reduction of bismuth III iodide with zinc powder. The synthesis of the left-half C1—C11 is summarized in Scheme 1. Ozonolysis of the tert -butyldimethylsilyl ether 6 11 of S -citronellol with a reductive workup gave S tert -butyldimethylsilyloxymethylhexanol 7.

the background to C NMR spectroscopy

This synthesis of 9 is straightforward and scalable. Reagents and conditions: i TBSCl, imid. The bismuth-mediated reaction of aldehyde 9 with the pentenyl bromide 10 9 gave a major product identified on the basis of precedent, 10 as the 3 E ,6 S -2,6- anti -2,4,8-trimethylundecenol The 3 E ,6 S -undecenol 11 was converted into its 3 E ,6 R -epimer 15 by treatment with 4-nitrobenzoic acid under Mitsunobu conditions followed by hydrolysis of the intermediate 4-nitrobenzoate Reductive cleavage of the benzyl ether 11 followed by selective silylation of the primary alcohol 12 gave the triisopropylsilyl ether Structures were assigned to these products by analogy with our earlier work 9 and were confirmed by comparison of the 13 C NMR spectra of later intermediates, in particular of the 2,4,6,8-tetramethylundecanol 20 , with published data.

The remaining methyl group was introduced by treatment of the toluene p -sulfonate 17 , prepared from the alcohol 16 , with a higher order methyl cuprate. Desilylation gave the alcohol 20 that was converted into the iodide 22 via the toluene p -sulfonate The syntheses of the enantiomeric C 12 —C 22 fragments and completion of the syntheses of the epimeric 4,6,8,10,pentamethyldocosanes 2 and 4 are outlined in Scheme 2.

Alkylation of butyl phenyl sulfone using the R - and S -iodides R - 23 and S - 23 16 that are available from the corresponding enantiomers of citronellol gave the dodecenyl sulfones 24 and 31 as mixtures of epimers at C 4. The alkenes in 24 and 31 were ozonized with a reductive workup to give the alcohols 25 and 32 , and then reduction by sodium amalgam gave the enantiomeric 4-methyldecanols S - and R - Reductive removal of the phenylsulfonyl group gave the 4,6,8,10,pentamethyldocosanes 2 and 4.

Again there are data sets of the natural product and the four-isomer mixture from Kitching. To control for both temperature effects and inherent reproducibility, we recorded four new 13 C NMR spectra on synthetic samples 2 and 4 two each, 1 week apart at MHz in CDCl 3 with a nominal probe temperature of K.

  1. Trends in composite materials and their design : special topic volume with invited peer reviewed papers only.
  2. Lectures on Cauchys Problem in Linear Partial Differential Equations?
  3. Help & contacts.
  4. NMR Spectroscopy!
  5. benchtop NMR | Magritek.

These data are fully tabulated in the Supporting Information alongside those of the published spectra of isomers 2 and the natural product. Comparing the spectra of the same samples recorded 1 week apart gave an idea of the random error of the experiments, which was strikingly low. We work first with the new data sets at K.

5.6B: \(^{13}C\)-NMR in isotopic labeling studies

The step 1 subtraction revealed that the differences between the candidate isomers of 2 and 4 are considerably smaller than those of 1 and 3. As before, all of the resonances with the differences exceeding 2. This is sensible. The largest difference of all, Also as before, the uncertain resonances are just to the left and right of the reliably different resonances along the chain. This time, only half 5 of the reliably different and uncertain resonances belong to methylene groups.

The other half are methyl groups 3 and methines 2. Validation that these tiny subtraction differences are real and not error again comes from the relevant mixture sample of Kitching. The pentamethyldocosane mixture is again four isomers, epimers at C16 the relevant center and C Thus, Kitching again correctly identified the peaks that differ in the natural product and its C16 epimer. At MHz, Kitching did not observe doubling of any of the three peaks that differ by 3—5 ppb in our spectra.

We suspect that these differences are meaningful because they show up reliably when subtracting the duplicate spectra recorded at MHz. This error level is crucial; if it is raised to just 10 ppb 0. Indeed, the spectra are so similar that even small differences in sample temperatures as low as 2—3 K can complicate comparisons because the magnitudes of temperature effects on chemical shifts can exceed the actual differences see below. These data were corrected for small calibration errors see Supporting Information , though again the correction is not crucial to the outcome.

Then in step 3 we compared the subtraction data for the seven reliably different resonances, as shown in Table 3. The results of subtractions NP — 2 and NP — 4 are shown in columns 3 and 4. In NP — 2 all of the values are zero or nearly zero; this is the match. In NP — 4 , the values are very close to the reliably different values established in step 1 column 2 ; this is the mismatch.

Clearly the natural product is 2 , not 4. Accordingly, the hexa- and pentamethyldocosane natural products 1 and 2 have the same configuration at C The usual way to compare spectra of two very similar samples to an unknown that is presumed to match one and not the other is to subtract pairs of the same chemical shifts in different samples , as we did in Table 2. We have previously suggested that subtraction of pairs of different chemical shifts in the same sample from each other is also a valuable tool.

Because two chemical shifts are subtracted, it gives the largest possible ppb difference in the spectrum, and most importantly, any calibration error cancels in the subtraction. So this method works without standardization of the spectra being compared. This difference is The natural product difference is Such subtractions of the absolute values give two large numbers that when compared have a small difference.

An automated framework for NMR chemical shift calculations of small organic molecules

Because only the difference is meaningful, it may be more convenient to look at the difference of differences in values of the chemical shifts. In other words, subtract the differences of C13—C9 in 2 and 4 from each other. This is the mismatch result, rounded to 19 ppb for comparison with the natural product.

As usual, the match result is zero. Likewise, for hexamethyldocosane, the same two carbons give an almost double difference of differences of 36 ppb. Subtracting the natural product value from 1 gives 0 ppb match result and subtracting the NP value from 3 gives 36 ppb mismatch result. When comparing pairs of spectra with such tiny chemical shift differences, it is crucial to account for possible differences in sample temperature. We show this by comparing the newly recorded sets of spectra of 2 and 4 to the original published spectra.

However, the published spectrum of 4 did not match the new duplicate spectra of 4 nearly as well as the duplicates matched each other. Disconcertingly, according to the raw subtraction analysis subtract all chemical shifts and calculate a standard deviation , the published spectrum of 4 is actually closer to the published spectrum of the natural product than to the new spectra of the same sample.

This is disconcerting because 4 is not the natural product. We deduced that the original sample temperature of isomer 2 matched the sample temperature of the new spectra of 2 , while the sample temperature of isomer 4 did not.

13C NMR Lecture

To retrospectively estimate the sample temperature of the published spectrum of 4 , we recorded pairs of 13 C NMR spectra of both candidate isomers at K 5 K lower sample temperature , 1 week apart. Again, the spectra of the same samples recorded 1 week apart were identical down to the 1—2 ppb level. Differences in chemical shifts for spectra of 4 recorded with sample temperatures 5 K apart. Most of the other shifts were positive that is, downfield shift with increasing temperature , but the two end methyl groups C1 and C22 and one of the methylene groups C21 experienced small negative shifts.

These kinds of shifts are typical for other alkanes. This means that the published spectrum of 4 must have been recorded at about K. This exercise shows how to estimate an unknown sample temperature retrospectively given available candidate sample spectra. Here we show the scatter plots from raw subtraction that is, the current standard method of data analysis. The results when the new spectrum of 2 recorded at K is subtracted from the NP spectrum at K are shown with the blue diamonds. This is the case where the spectra of the correct candidate isomer and the natural product are compared, but the spectra are recorded at different temperatures right structure, wrong temperature.

Overwhelming effect of sample temperature. The scatter plot for the wrong structure at the right temperature red squares looks much better than the plot for the right structure at the wrong temperature blue diamonds. Now compare this to the subtraction of the wrong candidate isomer 4 from the natural product with spectra recorded at identical temperatures, shown with the red boxes.

This is the case where the temperature matches, but the structures do not right temperature, wrong structure. In other words, the usual raw subtraction process assigns the wrong structure to the natural product because it fails to account for the temperature effects. Chemical shift differences as a function of temperature are not random errors and therefore cannot be expected to cancel in any raw subtraction analysis.

In summary, we have postulated, validated, and then applied a systematic process for comparing 13 C NMR spectra of two or more candidate samples of known structure with the spectrum of another compound of unknown structure for example, a natural product. The process is designed for cases where the spectra of the candidates are very similar, perhaps even identical. In such cases, raw subtraction with scatter plot visualization is the current standard for assignment. This method always come down on the side of one candidate or the other. Indeed, even if the candidate samples have identical spectra, an assignment will still result simply because of error.

Accordingly, when spectra are similar, there is a low level of confidence that an assignment by the raw subtraction method is correct. In the new method, the spectra of the candidates are first compared to each other to firmly establish that they are different and then to articulate the differences. These differences should be sensible based on the structures being compared. The assignment does not rely on a statistical analysis such as determination of a standard deviation.

Instead, all of the key resonances of the natural product are expected to match one candidate isomer and not to match the other. The result is both an assignment of the natural product structure and a high level of confidence that the assignment is correct. The method reveals differences that are comparable to those revealed by recording spectra of mixed samples, a gold-standard process that is rarely practical either because no natural sample is available or because the synthetic and natural samples are simply too precious to mix.

Such small differences are not usually considered meaningful and are typically ascribed to random error. However, provided that sample temperature differences are corrected for, these small differences are demonstrably real and therefore reliable.


1H and 13C NMR for the Profiling of Natural Product Extracts: Theory and Applications

NMR probes around the world probably have ambient temperatures in the range of at least — K, a 10 K range. When spectra recorded at different sample temperatures are subtracted, real temperature effects which are not random are easily mistaken for random error. Given the advantages over the standard raw subtraction analysis, we suggest that this systematic process should become standard practice for assignment in cases where two or more candidate structures for a natural product can reasonably be expected to have very similar or identical spectra.

Finally, we caution that both or all of the candidate samples have to be made for the secure assignment. One cannot simply make and then match one candidate to the natural product structure. This is an uncontrolled process. How do we know which resonances are the same or reliably different if we do not know whether the candidate and the natural product are the same or different? Only by circular reasoning. Only when the other candidates 1 and 3 are introduced does it become clear that 2 and 4 are the mismatches and 1 and 3 are the matches.

As we have stated previously, 4d assigning structures by proof has the same pitfalls as proving mechanisms. In formal logic, a structure or mechanism can only be disproved, never proved. The final proof comes from disproof of all sensible structure candidates or mechanism candidates but one.

Imidazole mg, 7. The reaction mixture was stirred at room temperature for 16 h, diluted with ether 25 mL and partitioned between ether 20 mL and saturated aqueous sodium hydrogen carbonate. The organic layer was washed with brine 20 mL and dried Na 2 SO 4. C 12 H 25 OSi requires M , Ozone from a generator was bubbled through a solution of the alkene 6 1. After concentration under reduced pressure, the residue was partitioned between saturated aqueous sodium hydrogen carbonate 20 mL and ether 30 mL.

The organic layer was washed with brine 20 mL , dried Na 2 SO 4 and concentrated under reduced pressure. Iodine 4. Saturated aqueous sodium sulfite 30 mL and ether 50 mL were added, and the mixture was partitioned between water and ether. The organic layer was washed with brine 50 mL , dried Na 2 SO 4 and concentrated under reduced pressure.

Chromatography eluting with light petroleum gave 4 S tert -butyldimethylsilyloxymethyliodohexane 3. A portion of this iodide 3.