Rapid chiral analysis based on liquid-phase cyclic chemiluminescence

DOI: 10.1039/d0sc03496g

Source and publish data:

Chemical Science p. 660 - 668 (2021)

Update date:2022-08-16

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Authors:

Zhang, Runkun

Zhong, Yanhui

Lu, Zhenyu

Chen, Yanlong

Li, Gongke

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Article abstract of DOI:10.1039/d0sc03496g

Rapid chiral analysis has become one of the important aspects of academic and industrial research. Here we describe a new strategy based on liquid-phase cyclic chemiluminescence (CCL) for rapid resolution of enantiomers and determination of enantiomeric excess (ee). A single CCL measurement can acquire multistage signals that provide a unique way to examine the intermolecular interactions between chiral hosts and chiral guests, because the lifetime (τ) of the multistage signals is a concentration-independent and distinguishable constant for a given chiral host-guest system. According to the τ values, CCL allows discrimination between a wide range of enantiomeric pairs including chiral alcohols, amines and acids by using only one chiral host. Even the chiral systems hardly distinguished by nuclear magnetic resonance and fluorescence methods can be distinguished easily by CCL. Additionally, the τ value of a mixture of two enantiomers is equal to the weighted average of each enantiomer, which can be used for the direct determination of ee without the need to separate the chiral mixture and create calibration curves. This is extremely crucial for the cases without readily available enantiomerically pure samples. This strategy was successfully applied to monitoring of the Walden inversion reaction and analysis of chiral drugs. The results were in good agreement with those obtained by high-performance liquid chromatography, indicating the utility of CCL for routine quick ee analysis. Mechanism study revealed that the τ value is possibly related to the activity of the chiral substance to catalyze a luminol-H2O2 reaction. Our research provides an unprecedented and general protocol for chirality differentiation and ee determination, which is anticipated to be a useful technology that will find wide application in chirality-related fields, particularly in asymmetric synthesis and the pharmaceutical industry.

Full text of DOI:10.1039/d0sc03496g

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DOI: 10.1039/D0SC03496G

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Rapid Chiral Analysis Based on Liquid-Phase Cyclic

Chemiluminescence

Runkun Zhang, Yanhui Zhong, Zhenyu Lu, Yanlong Chen, and Gongke Li*

Received 00th June 2020,

Accepted 00th June 2020

Rapid chiral analysis has become one of the important aspects of academic and industrial research. Here we describe a new

strategy based on liquid-phase cyclic chemiluminescence (CCL) for rapid resolution of enantiomers and determination of

enantiomeric excess (ee). A single CCL measurement can acquire multistage signals that provide a unique way to examine

the intermolecular interactions between chiral hosts and chiral guests, due to the lifetime (τ) of the multistage signals is a

concentration-independent and distinguishable constant for a given chiral host-guest system. According to the τ values, CCL

allows discrimination between a wide range of enantiomeric pairs include chiral alcohols, amines and acids by using only

one chiral host. Even the chiral systems hardly distinguished by nuclear magnetic resonance and fluorescence method can

be distinguished easily by CCL. Additionally, the τ value of the mixture of two enantiomers is equal to the weighted average

of each enantiomer, which can be used for direct determination of the ee without the need to separate the chiral mixture

and create calibration curves. It is extremely crucial for the cases without readily available enantiomerically pure samples.

This strategy was successfully applied to monitoring of Walden inversion reaction and analysis of chiral drugs. The results

were in good agreement with those measured by high-performance liquid chromatography, indicating the utility of CCL for

routine quick ee analysis. Mechanism study revealed that the τ value is possible related to the activity of chiral substance to

DOI: 10.1039/x0xx00000x

2 2

catalyze luminol-H O reaction. Our research provides an unprecedented and general protocol for chirality differentiation

and ee determination, which is anticipated to being a useful technology that find wide applications in chirality-related fields,

particularly in asymmetric synthesis and pharmaceutical industry.

Introduction

Chirality is a universal phenomenon in nature, most attracted considerable research interest as a result of their

biological species, such as amino acids, nucleic acid, proteins, advantages of rapidity, simplicity, and low cost. In such a

drugs, are chiral and intermolecular chiral-chiral interactions strategy, the molecules or materials containing the chiral unit

-3

occur ubiquitously among biological processes.

The are used as chemosensory hosts that can interact with the chiral

simultaneous determination of the absolute configuration and targets and then induce distinguishable optical signals.²

enantiopurity of chiral substrates is highly significant in the Although the great efforts and achievements that have been

-6

fields of synthetic, biological and pharmaceutical chemistry.

made in this field, some problems still remain to be solved. For

However, chiral analysis remains a great challenge due to instance, most methods suffer from narrow substrate scope

enantiomers have nearly identical physical properties.

and limitedly differentiable enantiomers. In addition, the

In the past several decades, many effective methods have output signal for chiral discrimination is strongly influenced by

been developed for the chiral analysis, such as gas the analyte concentration, this would normally need two

chromatography, high-performance liquid chromatography independent methods to simultaneously determine both

HPLC),¹

0,11

capillary electrophoresis,

(NMR)

18,19]

12-14

nuclear magnetic enantiomeric configuration and enantiopurity.

28,29

Especially,

mass these assays usually require highly pure samples to construct

curves for enantiomeric excess (ee)

It would be particularly significant to

(

,15-17

resonance

spectrometry.^[

spectroscopy

and

However, these methods are time- calibration

,30,31

consuming and technically complex, and the instrumentation determination.

required is expensive, bulky, and requires well-trained acquire a concentration-independent parameter for chiral

0,21

operators.

dichroism,²

Optical sensing methodologies, such as circular discrimination and enantiopurity determination, due to such

2-24

25,26

fluorescence

and colorimetry

have technique can simplify and speed up the measurement, and

thereby find wide applications in rapid screening of ee for

asymmetric synthesis and pharmaceutical industry etc. In this

paper, we report an unprecedented strategy of liquid-phase

cyclic chemiluminescence (CCL) that does not suffer from the

^a^.School of Chemistry, Sun Yat-sen University, Guangzhou 510275, People’s

Republic of China；E-mail: cesgkl@mail.sysu.edu.cn

Electronic Supplementary Information (ESI) available: Fabrication and working

principle of liquid-phase CCL system; method for immobilization of luminol; Fig.S1-

S10 and Table S1-S5. See DOI: 10.1039/x0xx00000x

above limitations in chiral analysis. To the best of our

knowledge, this paper is the ﬁrst to report

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Journal Name

chemiluminescence-based sensing methodology that providing by reactive oxygen species to form a 3-aminophthalate dianion

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DOI: 10.1039/D0SC03496G

a concentration-independent constant for the simultaneous in an excited state, which emitted light when returning to the

determination of enantiomeric configuration and ee value.

ground state. Specifically, a circular pipeline was designed for

As an important optical analytical technology, CCL system, and the direction of the carrier in CCL system can

chemiluminescence (CL) have been widely used for be changed automatically at the indicated cycle period. Through

33-35

pharmaceutical analysis, clinical analysis

and biological automatically changing the carrier direction at the indicated

analysis.³

6,37

However, typical CL kinetic curve lacks the period, the chemical species at different stages flowed through

characteristic features of the analytes. To overcome this the sensor cell repeatedly to induce the CL reactions proceed in

drawback, our group proposed a new concept of cyclic succession. Multistage signals that satisfy the exponential decay

chemiluminescence (CCL) for rapid discrimination of law were measured over time. The design of CCL system

structurally similar compounds. In contrast to conventional extends the observable time, and thereby the distinctions

flow CL system, the direction of the carrier in the CCL system is between different luminous reactions that usually are not easy

in a state of periodical change, which triggers a succession of CL to observe by conventional CL-based detection, but can be

reactions in a single sample introduction to produce multistage observed by CCL detection. The flow paths of liquid-phase CCL

signals. We found that the multistage signals on CCL kinetic detection at different stages are shown in the ESI (Fig. S1).

curve satisfy the first-order exponential decay law:

For chiral analysis, chiral metal-organic complexes that are

able to enhance the luminescence intensity of luminol-H reaction

were used as hosts, new complexes were formed via chiral–



2 2



I  Ae  I

(1)

where I

is the signal of n th stage reaction, and I

is the chiral interactions between chiral host and chiral guest when

background noise. t is the peak time. A is the pre-exponential chiral guests were mixed with chiral hosts. The τ values of the

factor that stands for the maximum luminescence intensity. τ CCL kinetic curves of luminol-H reaction catalyzed by the

2 2

represents the lifetime of the multistage signals, it is the time at new complexes were use for chiral analysis. The schematic

which the intensity of the assembly is reduced to 1/e times its representation of liquid-phase CCL for chiral analysis is shown

initial value, its unit is "second". We demonstrated each given in the Fig. 1.

CCL reaction has a unique constant of τ. The distinguishable τ

values allow facile identification of various analytes include

homologous series and structural isomers. Recently, we further

demonstrated that the multistage signals of multicomponent

mixture also satisfy the first-order exponential decay law with

their unique τ vaues.³This allows facile identification of mixed

samples consisting of complex components, such as liquor,

beer, toner, and baby powder. However, our previous

researches were limited to studying the gaseous-phase CL

emitted on nanosized catalysts, and a wider adaptation for the

application of CCL has not been demonstrated yet. Following

our research interest in demonstrating the generality of CCL and

exploring its further application, here, we extend the CCL

strategy to liquid-phase reactions via immobilizing luminous

2 2

reagent, and metal-organic complex-catalyzed luminol-H O

reaction system was used as basic model to probe the

application of CCL in chiral analysis. The most distinct advantage

of liquid-phase CCL is that it provides a concentration-

independent constant (τ value) to discriminate a wide range of

enantiomers, and allows direct determination of the

enantiopurity without the need to separate the chiral mixture,

and thereby the use of enantiomerically pure samples for

creating calibration curves can be avoided.

Fig. 1. The schematic representation of liquid-phase CCL for chiral analysis.

Results and discussion

Resolution of enantiomers

Experimental section

The fabrication of liquid-phase CCL system can be seen in

the ESI. In the present work, immobilized luminol (5-amino-2,3-

dihydro-1,4-phthalazinedione, immobilized method was

detailed in the ESI) was packed into a homemade flow-through

2 2

reactor to fabricate a sensor cell. H O was used as a carrier and

oxidant, and it was catalyzed by a sample containing a metal-

organic complex to produce reactive oxygen species.

Subsequently, immobilized luminol in a sensor cell was oxidized

To examine the feasibility of our strategy for chiral analysis,

three pairs of commercially available chiral metal-organic complexes

(

R1 and S1, R2 and S2, R3 and S3, the detail information of these

chiral complexes can be seen in Table S1 of the ESI) were used as

hosts. The original CCL kinetic curves of luminol-H system

2 2

catalyzed by these chiral metal-organic complexes are shown in the

ESI (Fig. S2). It shows that the two enantiomers of one chiral

substrate of metal-organic complex have the same τ value when

without adding chiral guests. This because the differences between

| J. Name., 2012, 00, 1-3

This journal is © The Royal Society of Chemistry 20xx

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enantiomers are difficult to observe under normal circumstance, but constant that characteristic of a given chiral system, w _V h _i _e ic _w h _A f _r _t u _i _c n _l _e c _O ti _n o _l _i n _n _e s

these differences become much more marked in a chiral as characteristic labelling for directly dete^Drm^O^Ii^:n¹i⁰n^.g¹⁰³th⁹e^/^D⁰id^Se^Cn⁰t³it⁴y⁹⁶o^Gf

environment. Thus, chiral guests were added into the solutions of enantiomers.

chiral hosts. We found that additions of (R)-(+)-1-phenylethanol and

(

S)-(-)-1-phenylethanol into R1 outputting τ values of 64.6 and 39.0 s

Fig. 2a), respectively. τ values of 35.2 and 51.6 s were observed

when added (R)-(+)-1-phenylethanol and (S)-(-)-1-phenylethanol into

S1 (Fig. 2b).

We further investigated the effects of the concentration of

guest on response behaviors of host at a much broader

concentration range. We have plotted the τ values of R1 and S1

versus the increasing concentrations of (R)-(+)-1-phenylethanol and

(S)-(-)-1-phenylethanol in Fig. 2c and Fig. 2d, respectively. We found

that the τ value is independent of the concentration of the chiral

guest in the range of 0.005 to 0.5 mmol/L but is only dependent on

the chiral configuration. In addtion, the τ value also is independent

of the H

discussion can be seen in the ESI). These results demonstrate that Fig. 2. (a) The CCL kinetic curves of luminol-H

each chiral system has a unique and constant τ value, and the phenylethanol enantiomers; (b) the CCL kinetic curves of luminol-H

2 2

O concentration within a certain range (Fig. S3, detailed

system catalyzed by R1 with 1-

system

2 2

distinction in τ value confirms the chiral discrimination. Quite catalyzed by S1 with 1-phenylethanol enantiomers; (c) Plots of τ value for R1 (0.1

different from other sensing methodologies based on intensity mmol/L) in the presence of varying concentrations of 1-phenylethanol; (d) Plots of

quenching/enhancement or spectral changes, the parameter (τ τ value for S1 (0.1 mmol/L) in the presence of varying concentrations of 1-

value) of CCL for chiral analysis is a concentration-independent phenylethanol. The concentration of H

2 2

O is 0.05 mmol/L.

Fig.3. The τ values of different chiral host-guest systems. The concentration of all hosts and guests is 0.1 mmol/L, the concentration of H

2 2

O is 0.05 mmol/L. The unit of

τ is second.

This journal is © The Royal Society of Chemistry 20xx

J. Name., 2013, 00, 1-3 | 3

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The reproducibility of the immobilized luminol was investigated by

Noteworthy, each chiral host-guest system has a corresponding

measure of (R)-(+)-1-phenylethano using S1. We found that the τ value, that is: different hosts show different values

background noise increases and luminescence intensity (A) guest, and the same host exhibits different values of τ to different

tic

line

DOI: 10.1039/D0SC03496G

decreases gradually with increasing testing number (ESI, Fig. S4), but guests. The above results demonstrate that each chiral host-guest

the τ could remain stable within 25 replicates with a RSD of 3.7% (ESI, system has a corresponding exponential decay equation with a

Fig. S5a). It means that τ has a good stability, possibly due to τ unique τ value. Thus, we can establish a database of τ values of

depends on the rate of luminescence depletion that related to the various chiral host-guest systems in future, which provides

characteristics of the reaction, but not the apparent intensity. The τ facilitation for quick recognition of unknown chiral compounds. For

values measured by 5 different batches of immobilized luminol example, if the τ values of an analyte were measured to be 64.6 and

vary between 4.2% (ESI, Fig. S5b). The sufficient stability of τ within 35.2 s when using R1 and S1 as hosts, respectively, the identity of the

the same batch and between different batches of immobilized analyte could be confirmed to be (R)-(+)-1-phenylethanol by

luminol make CCL is qualified for chiral differentiation. It is worth searching database.

noting that the A value is easily fluctuant and the τ value depends on

the decay rate of multistage signal, and thereby the multistage signal satisfactory enantio-differentiating ability of our strategy. Usually,

can be normalized for nonlinear curve fitting to obtain τ value. we can discriminate most enantiomeric pairs by only using one chiral

Fig. 3 illustrates the broad substrate applicability, and the

An ideal method for chiral sensing should be capable of host. However, the τ values of some chiral compounds on certain

discrimination of different kinds of chiral molecules accompanied host are close. For example, although host S2 allows discrimination

with the resolution of the two enantiomers of one chiral substrate.

between enantiomers of 1-phenylethanol or 2-pentanol, it unable to

However, current methods usually suffer from narrow substrate resolve (S)-(-)-1-phenylethanol (τ=40.4 s) and (S)-(+)-2-pentanol

scope and limitedly differentiable enantiomers. We next turned (τ=40.5 s). Our aim is to realize unambiguous discrimination among

attention to demonstrate the ability of CCL to discriminate a wide a large variety of chiral compounds, not just limits to the two

range of enantiomers with broad substrate applicability. The chiral enantiomers of one chiral substrate. In consideration of the cross-

metal-organic complexes shown in the ESI (Table S1) were used as reactivity of the hosts, we combined all the hosts (R1, S1, R2, S2, R3

hosts to sensing two pairs of chiral alcohols, chiral amines and chiral and S3) to construct a sensor array, expecting to realize

acid, respectively. As expected, each enantiomer can be correlated unambiguously discriminate various chiral compounds. Three

to a distinct τ value when it is mixed with a given chiral host (Fig. 3), concentrations (low: 0.005 mmol/L, medium: 0.05 mmol/L and high:

enabling easy discrimination of chiral compounds. For example, 0.1 mmol/L) of 12 chiral compounds were tested by the sensor array,

when chiral complex R1 was used as a chiral host, τ values of 64.6 the resulting τ values are summarized in the ESI (Table S2). For easier

and 39.0 s were observed for (R)-(+)-1-phenylethanol and (S)-(-)-1- visualization, the τ value of each response is depicted in the form of

phenylethanol; 52.7 and 38.2 s for (R)-(-)-2-pentanol and (S)-(+)-2- a heat map according to the scale bar on the right. As shown in Fig.

pentanol; 58.3 and 46.5 s for (R)-(-)-2-phenylglycinol and (S)-(+)-2- 4a, each of the chiral guests creates a unique multidimensional

phenylglycinol; 12.1 and 21.4 s for (R)-3-amino-3-phenylpropan-1-ol response pattern, and different concentrations of the same chiral

and (S)-3-amino-3-phenylpropan-1-ol; 49.6 and 73.9 s for D-(-)- guest induce similar response pattern. Linear discriminant analysis

tartaric acid and L-(+)-tartaric acid; 31.6 and 23.7 s for D-(-)-mandelic (LDA) was then used to further digitize and visualize the response

acid and L-(+)-mandelic acid, respectively. Satisfactory resolution of patterns (6 host units × 12 chiral guest × 3 concentrations). Fig. 4b

the two enantiomers of one chiral substrate is easily achieved by shows the 3-D discrimination plot constructed by the first three

simply comparing the τ values.

factors. The first three factors account for 85.5% of the total variance.

Fig. 4. (a) Response patterns of 6 chiral hosts to 12 guests at different concentrations. 0.005, 0.05 and 0.1 strand for the concentrations of the guests, the unit is mmol/L;

b) LDA 2D plot derived from the colour response pattern. A: (R)-(+)-1-phenylethanol; B: (S)-(-)-1-phenylethanol; C: (R)-(-)-2-pentanol; D: (S)-(+)-2-pentanol; E: (R)-(-)-2-

phenylglycinol; F: (S)-(+)-2-phenylglycinol; G: (R)-3-amino-3-phenylpropan-1-ol; H: (S)-3-amino-3-phenylpropan-1-ol; I: D-(-)-tartaric acid; J: L-(+)-tartaric acid; K: D-(-)-

(

2 2

mandelic acid; L: L-(+)-mandelic acid. The concentration of all host is 0.1 mmol/L, the concentration of H O is 0.05 mmol/L.

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Chemical Science

The same colored symbol within the LDA graphical output assigns to percentages of 1-phenylethanol enantiomers, and the trends of the

specific chiral guests. These chiral guests are classified into 12 groups τ value of the mixture (τ ) versus the percentages Vie

with 100% accuracy, and small spatial distance are observed within phenylethanol and (S)-(-)-1-phenylethanol were plotted in Fig. 5,

the same group. This demonstrates the satisfactory discrimination respectively. Linear-regression analysis shows that the τ is linearly

capacity of the sensor array based on CCL strategy. related to the percentages of the enantiomers in the mixture.

DOI: 10.1039/D0SC03496G

NMR spectroscopy and fluorescence spectroscopy are common Further study found that the slope of the linear regression equation

technique used for chiral discrimination. However, we found that the is equal to difference between the τ values of the two enantiomers,

R1 or S1 with 1-phenylethanol enantiomers (ESI, Fig. S6). The same and the intercept equals the τ value of one of the enantiomers. The

chiral systems also cannot be resolved by fluorescent method, due

τ of the mixture composed by arbitrary proportions of enantiomers

to R1, S1 and 1-phenylethanol are non-fluorescent substances. These can be expressed as:

results demonstrate that CCL can distinguish some chiral host-guest

_m ₍_R₎(₍_R₎₍_S₎) ₍_S₎    (1₍_R₎)₍_S₎

(2)

(R) (R)

systems, which would be difficult for NMR and fluorescence

spectroscopy. Compared with the method for chiral discrimination

Or can be presented in the following form:

_m  ( ₍_R₎) ₍_R₎ (1 )   

using QDs enhanced-CL system, the commercial chiral metal-

organic complexes used in the present work are easy available. In

addition, they allow facile discrimination between a wide range of

enantiomeric pairs. It is well known that static injection CL can

provide the full intensity kinetic curve. However, we found that the

CL kinetic curves obtained by static injection lack enough

discriminability to differentiate chiral compounds, mainly due to the

short lifetime of the CL kinetic curve. The τ values of different kinetic

curve obtained by static injection are in the in the range from 1.0 to

(3)

(S )

(R)

(S ) (S )

Where ω(R) and ω(S) are the percentages of the two enantiomers

in the mixture, and τ(R) and τ(S) stand for the τ values of the two

enantiomers, respectively. Eqs 2 and 3 indicate that the τ

is the

weighted average of the two enantiomers, which is in accordance

with the gaseous-phase CCL detection. According to eqs 2 and 3,

(R) and ω(S) in the mixture can be presented by:

^m -₍_S₎

.6 s (ESI, Fig. S7), the narrow τ-range means low resolution. The

₍_R₎

₍_S₎



(4)

(5)

detailed discussion also can be seen in the ESI. Thus, CCL will be a

powerfully complementary technology for chiral discrimination.

₍_R₎₍_S₎

₍_R₎

-_m

^⁽R)

₍_S₎

Determination of ee

The ee (%) is defined as:⁴⁶

Determination of ee is performed routinely in many scientific

fields, especially in asymmetric synthesis and pharmaceutical

ee  ₍_R₎₍_S₎

(6)

1-44

industry.

Chiral separation is the most common method for ee

by integrating eqs 3, 4 and 5, the ee can be calculated by:

determination. However, chiral separation is cost-intensive and

time-consuming process due to the long analysis time and the use of

2 - -₍_S₎

(R)

1,45

ee 

(7)

expensive chiral column.

Simple and inexpensive techniques for

^⁽R)

₍_S₎

rapid determination of ee is attractive for their potential applications

in academic and industrial fields. In order to probe the application

value of our strategy in rapid determination of ee, R1 and S2 was

used to measure a series of mixtures consisting of different

Eqs 4, 5 and 7 indicate that if the τ values of the enantiomers

are known in advance (through searching database), the percentages

Fig. 5. The relationship between τ value and the percentages of 1-phenylethanol enantiomers using R1 (a) and S1 (b) as hosts.

the two enantiomers and the ee value can be directly determined by in the ESI compare the calculated versus the actual ee values,

measuring the τ value of the mixture without the need to use excellent linear relationships (regression coefficients exceed 0.998)

enantiomerically pure samples for creating calibration curves. Fig. S8 with slopes close to 1 were obtained. The absolute errors between

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Journal Name

the calculated and actual ee values ranged from -6.9% to 4.6% (ESI, shown in Fig. 6. The general shape of the curves obtained by the two

View Article Online

DOI: 10.1039/D0SC03496G

Table S3).

methods is surprisingly similar, showing the conversion of (S)-(-)-1-

phenylethanolfollows first order reaction kinetics. The relative

content of (S)-(-)-1-phenylethanol steadily decreases, while the yield

of (R)-(+)-1-phenylethanol steadily increases with the conversion of

Applications

To validate the utility of CCL in enantiopurity determination, an

amberlyst-catalyzed Walden inversion was selected as a prototype

reaction because of its industrial application in production of

pharmaceutical and agrochemical building blocks. 1.0 g of

amberlyst-H was added to 100 mL solution of (S)-(-)-1-phenylethanol

(

S)-(-)-1-phenylethanol. Complete substrate conversion within 240

min was observed. The half-lives calculated by CCL (t1/2(CCL)) and HPLC

(t₁_/₂₍_H_P_L_C₎) are in good agreement. The ee values at different reaction

times measured by the two methods are listed in the ESI (Table S5).

The CCL determined ee values correlated well with chiral HPLC

results with absolute errors ranging from -7.1% to 7.0%%.

at 0.2 mmol/L, duplicate the above solutions were heated to 55 C

and 65 C under stirring condition, respectively. At various times, 2

It is worth noting that during the processes of the above assays,

the analytical time of each HPCL measurement was 13 min (ESI, Fig.

S9), the total time spent on HPCL detections was about 377 min (13

min of each detection × 7 standard solutions + 13 min of each

detection × 22 samples). Only about 5 min of each CCL measurement,

moreover, chiral separation and calibration curve are not necessary

for CCL measurement, the total analytical time of CCL detections was

mL of the reaction mixture aliquots were withdrawn and added to

cuvettes containing 2 mL of deionized water or chiral host (S1 at 0.2

mmol/L). A specific chiral separation column was used for HPLC

measurement, the enantiopurities were calculated according to the

calibration curves constructed in advance (the detail can be seen in

the ESI, Table S4, Fig. S9 and Fig. S10). For CCL measurement, the

percentages and the ee values of the 1-phenylethanol enantiomers

were directly calculated by eqs 4, 5 and 7 according to the τ values of

the aliquots. The reaction curves monitored by the two methods are

10 min (5 min of each detection × 22 samples). The above results

demonstrate CCL represent a fast and reliable tool that can reduce

the cost and time of chiral analysis.

Fig. 6. The reaction curves of conversion of (S)-(-)-1-phenylethanol at 55 C (a) and 65 C (b) monitored by CCL and HPLC. ω(S)(CCL) and ω(S)(HPLC) stand for the content of (S)-

(-)-1-phenylethanol measured by HPLC and CCL, respectively.

Analysis of ee of chiral drugs plays an important role in and Fig. S13). For CCL measurement, the ee values were directly

pharmaceutical industry and clinical pharmacy, due to only one of caudated by eq 7 according to the τ values of samples. The results

the two enantiomers is pharmaceutically active ingredient in many obtained by the two methods are in good agreement with absolute

cases Ibuprofen is one of the most potent anti-inflammatory drugs errors ranging from -4.2% to 4.0%(Table 1). This example illustrates

used for the treatment of acute and chronic pain, fever and the promising application of our strategy in analysis of chiral drugs.

inflammatory diseases. It has been proved that (S)-(+)-ibuprofen is

almost entirely responsible for the anti-inflammatory activity. In

order to further explore of utility of our strategy in enantiopurity

determination, we used S1 as host to analyze ibuprofen samples.

First, the standard samples of (R)-(-)-ibuprofen and (S)-(+)-ibuprofen

were measured by CCL, the τ values of them are found to be 44.6 and

Table 1 Determination of ee Values of Ibuprofen Samples by CCL and

HPLC

ee (%) measured

by CCL

-4.5 ± 2.7

5.6 ± 3.8

-1.6 ± 6.0

-5.6 ± 2.7

-42.1 ± 3.5

ee (%) measured by Absolute error

Samples

HPLC

4.2 ± 1.9

1.6 ± 5

(%)

-0.3

6.1 s, respectively (ESI, Fig. S11). Next, samples containing

-1.7 ± 8.2

-1.4 ± 0.32

-45.6 ± 2.3

0.1

-4.2

3.5

ibuprofen were dissolved in ethanol, then was diluted with deionized

water. Finally, the prepared solutions were simultaneously analyzed

by CCL and HPLC. Similarly, a specific chiral column was used and

aSamples 1 and 2 are two commercially available ibuprofen tablets; samples 3 and

calibration curves were constructed in advance for HPLC 4 are two commercially available ibuprofen capsules; sample 5 was prepared by

determination (the detail can be seen in the ESI, Table S6, Fig. S12 mixing analytical grade (R)-(-)-ibuprofen with (S)-(+)-ibuprofen at random.

| J. Name., 2012, 00, 1-3

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Mechanism study

One of the major approaches to chirality sensing is based on the

formation of diastereomeric complexes generated by intermolecular

_c_h_i_r_a_l_–_c_h_i_r_a_l_i_n_t_e_r_a_c_t_i_o_n_s_b_e_t_w_e_e_n_c_h_i_r_a_l_h_o_s_t_a_n_d_c_h_i_r_a_l_g_u_e_s_t_.₅,21,50,51

brings the hydroxyl radical close to nitrogen atom in the hosts,

forming hydrogen bond with different spatial structures, and thereby

the new adducts exhibit differential Raman bands (Video S1).

It is well known that different complexes show different

View Article Online

DOI: 10.1039/D0SC03496G

activities to catalyze luminol-H

difference in τ values among different chrial systems may relatied to

their catalytic activitie. Thus, the reaction kinetics of luminol-H

2 2

O reaction, we speculate taht the

In order to explore the work mechanism of CCL for chiral

discrimination, the density functional theory and the B3LYP method

in the Gaussian 09 program were employed to gain insight into the

possible modes of interactions between R1 (or S1) and 1-

phenylethanol enantiomers. Geometry optimizations were

performed using the basis set composed of LANL2DZ on cobalt atom

and 6-31G on the rest of the atoms, followed by frequency analysis

to insure all local minima with real frequencies only. The optimized

geometries for the new complexes are presented in Fig. 7, which

points out that 1-phenylethanol enantiomers can be linked to chiral

cobalt complexes (R1 and S1) via hydrogen bond to form adducts.

The binding geometries in the formative adducts are different, and

their binding energies are higher than -4 kcal/mol, indicating that

2 2

reactions catalyzed by different chrial systems were investigated by

using HPLC to monitor the consumption rate of luminol, and the

results are shown in the ESI (Fig. S15). In order to facilitate the

comparison, the reaction rate was expressed by lifetime using τ value.

The results showed that luminol-H

4.7 min) and S1 (τ = 15.2 min) have almost similar τ value. The τ

values of luminol-H reactions catalyzed by R1 with (R)-(+)-1-

2 2

O reactions catalyzed by R1 (τ =

2 2

phenylethanol, R1 with (S)-(-)-1-phenylethanol, S1 with (R)-(+)-1-

phenylethanol, and S1 with (S)-(-)-1-phenylethanol are 28.9, 19.2,

7.8 and 22.4 min. A high level of consistency exists between the τ

value of reaction kinetics monitored by HPLC and τ value of CCL

kinetic curve, which confirm our hypothesis. According to the above

results, we conclude that new adducts with different spatial

structures can be formed via intermolecular chiral–chiral interaction

between the chiral hosts and chiral guests. These new adducts show

stable complexes with different spatial structures were formed.

2 2

different catalytic activities for luminol-H O reaction, the distinction

in the catalytic activities can be reflected in the decay speeds of the

multistage signals measured by CCL, expressing as distinguishable τ

values.

Compared with the static injection CL, it seems that CCL

detection triggers the whole reaction proceed through a staged

process, which expands the differences relating to reaction kinetics

after multistage reaction. In addtion, additional features, such as

diffusion, convection and interaction between the sample and resin,

can probably do occur in CCL system. The difference in these features

may also affect the observed analytical signal. Therefore, extensive

work is still needed to further explore the mechanism.

Conclusion

In conclusion, we have described a brand-new optical strategy

of liquid-phase CCL for chiral analysis. This strategy provides a

concentration-independent parameter (τ value) to discriminate a

large variety of chiral substrates. Importantly, our strategy allows

rapid determination of ee value without the needs for chiral

separation and calibration curve, thereby paving the way for

rapid ee analysis. Further development can focus on establishing a

Fig. 7. The optimized structures of R1 with (R)-(+)-1-phenylethanol (a), R1 with

S)-(-)-1-phenylethanol (b), S1 with (R)-(+)-1-phenylethanol (c) and S1 with (S)-(-)-

(

-phenylethanol (d).

In order to further explore the interaction between R1 (or S1) standard database of τ value to realize the quickly and accurately

and 1-phenylethanol enantiomers, the mixtures of the R1 and (R)-(+)- recognize unknown chiral compounds. Mechanism study revealed

-phenylethanol (R1+R), R1 and (S)-(-)-1-phenylethanol (R1+S), S1 that that new adducts with different spatial structures can be formed

via intermolecular chiral–chiral interactions between the chiral hosts

and chiral guests. The new adducts exhibits different catalytic activity

for luminol-H O reaction, resulting in distinguishable τ values of the

2 2

and (R)-(+)-1-phenylethanol (S1+R), as well as S1 and (S)-(-)-1-

phenylethanol (S1+S), were measured by Surface-enhanced Raman

spectroscopy (SERS), density functional theory at the B3LYP/6-

multistage signals measured by CCL. Since CCL allows perception of

intermolecular interactions through outputting distinguishable τ

values. we are currently exploring this strategy to discriminate of

chiral biological substrates, and expecting to find applications in the

study of protein structure, molecular dynamics, drug design etc.

based on host-guest interactions. Further studies on these directions

are currently underway in our laboratory and will be reported in due

course. We believe the combination of our strategy and diversified

1++G(d,p) level was employed to calculate the Raman bands of the

new adducts. As Fig. S14 in ESI shows, the Raman spectra of R1+R

-1

and R1+S exhibit features at 1296 and 1278 cm , respectively.

Similarly, characteristic bands at 1281.1 and 1296.7 are observed for

S1+R and S1+S, respectively. This further demonstrates the

formation of different adducts. Molecular modeling revealed that

the stretching vibration of C-O in 1-phenylethanol enantiomers

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Journal Name

chiral hosts will offer a powerful platform to address some challenges

of chirality analysis.

Int. Ed., 2020, doi.org/10.1002/anie.202001680._V _i _e _w _A _r _t _i _c _l _e _O _n _l _i _n _e

T. R. Schulte, J. J. Holstein and G. H. Clever, Angew. Chem. Int.

DOI: 10.1039/D0SC03496G

Ed. 2019, 58, 5562−5566.

F. Y. Thanzeel, A. Sripada and C. Wolf, J. Am. Chem. Soc. 2019,

41, 16382−16387.

Conflicts of interest

There are no conflicts to declare.

Y. Sasaki, S. Kojima, V. Hamedpour, R. Kubota, S. Takizawa, I.

Yoshikawa, H. Houjou, Y. Kubo and T. Minami, Chem. Sci. 2020,

1, 3790−3796.

A.Akdeniz, T. Minami, S. Watanabe, M. Yokoyama, T. Ema and

P. Anzenbacher Jr, Chem. Sci. 2016, 7, 2016−2022.

Acknowledgements

F. F. Xie, Q. Bai, X. M. Jiang, X. S. Yu, Z. N. Xia and W. L. Wei, Acs

Appl. Mater. Interfaces 2018, 10, 11872−11879.

This work was supported by the National Natural Science

Foundation of China (Nos. 21605163, 21675178 and 21976213), the

Research and Development Plan for Key Areas of Food Safety in

Guangdong Province of China (No. 2019B020211001), and National

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Table of Contents

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