The Total Amount and Composition Determination of Crocetin and Its Derivatives from Gardenia Yellow by HPLC-MS-MS

Research Article

The Total Amount and Composition Determination of Crocetin and Its Derivatives from Gardenia Yellow by HPLC-MS-MS

  • Bodi Hui 1*
  • Jin Huang 1,2

*Corresponding Author: Bodi Hui, Department of Food Science, Beijing Union University, Beijing, P. R. China. Received date: January 15, 2021; Accepted date: April 27, 2021; Published date: May 26, 2021

Citation: Bodi Hui, Jin Huang, (2021). The Total Amount and Composition Determination of Crocetin and Its Derivatives from Gardenia Yellow by HPLC-MS-MS. J. Nutrition and Food Processing, 4(3); DOI:10.31579/2637-8914/037.

Copyright: © 2021 Chen Pan, This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Received: 15 January 2021 | Accepted: 27 April 2021 | Published: 26 May 2021

Keywords: gardenia yellow; crocetin; crocetin derivative; crocin; gardenia jasminoides ellis; high performance liquid chromatography- mass spectrometry/ mass spectrometry (HPLC-MS/MS); crocetin esters; major fragmentation

Abstract

Consisting of crocetin derivatives as major color materials, gardenia yellow is a water-soluble food colorant extracted from the fruits of Gardenia jasminoides Ellis. However, as color materials, the total crocetin and its derivative amount and composition cannot be determined because the structures of some crocetin derivatives are currently unknown and the commercially-available reference sample is still short. This study aims to develop a method for the total amount and composition determinations of crocetin and its derivatives from gardenia yellow. In practice, the absorption coefficient of each crocetin derivative is calculated from that of crocetin and the negative correlation of absorbance with molecular mass of each derivative, while the molecular mass of each crocetin derivative is substituted by the m/z value of its parent ion on MS. The relative standard deviation (RSD) of reproducibility was 3.17%. The recovery of added external reference sample was ≥97±1.61%. The minimum detection limit was 7.44μg/mL.

Introduction

Practical Applications

In the industrial base, a method to determine the absorption coefficient, normally presented as a color value, of gardenia yellow is currently applied for the product quality control scheme. Under such conditions, the product quality was controlled by its color shade but not mass. It is expected that this approach will be replaced by the method developed in this study for the product quality control scheme. By this method, the color value will be substituted by the mass amount of the product for its quality control. This effort will provide a possibility to verify the biological functions and evaluate the safety of gardenia yellow.

Introduction

Gardenia yellow is a product that is usually used in foods as a colorant under the category of food additive in eastern Asian countries, especially in China, Japan and South Korea (World Health Organization). The product is usually extracted from the fruits of Gardenia jasminoides Ellis by EtOH with further purification [1-5]. Crocetin and its derivatives are the major color materials of the product [6]. Apart from crocetin, the product usually comprises more than 15 species of crocetin derivatives, mainly esters.

The root structure of the derivatives, crocetin, is a C20 carotenoid with the molecular formula and mass of C20H24O4 and 328.40 Da. Crocin was the first discovered crocetin ester (from Crocus sativus L.) in nature, and occurs in the fruits of Gardenia jasminoides Ellis with the largest amount. Its molecular structure has been well elucidated. In subsequent studies, Crocus sativus L. was the primary focus in understanding the structure of crocetin esters. In 1975, several carotenoid glycosyl esters were isolated and identified from by ultraviolet–visible spectroscopy (UV/VIS), infrared spectroscopy (IR), 1H nuclear magnetic resonance (1H-NMR) and mass spectrometry (MS) [7]. Sujata V. et al further confirmed that crocetin esters were the metabolites in the followers of Crocus sativus L. A number of crocetin esters and their geometrical isomers were then identified by HPLC-PDA-MS [8-10]. Pfister S. et al first reported the isolation and structure elucidation of crocetin esters from the fruits of Gardenia jasminoides Ellis [11]. The identification and structure elucidation of crocetin esters were then undertaken [14-16]. It is believed that a continuous effort to elucidate the structures of crocetin esters will not stop until the structures of all esters are known from the fruits of gardenia yellow. The molecular structures of crocetin, crocetin ester and crocin, the most common encountered crocetin ester in gardenia yellow are given in Figures 1(a), (b) and (c).

Figure 1. Molecular structures of crocetin (a), crocetin ester (b) and crocin (c).

Because the structures of some derivatives are still unknown and the commercially available reference samples of all the derivatives, apart from crocetin-di-β-D-gentiobiosyl ester (crocin), are lacking in the market, the amount and composition of color materials, including crocetin and its derivatives from gardenia yellow cannot be currently determined by either external or internal references. In food industrial base, the addition of gardenia yellow (as a colorant) to food media can only be undertaken by its color value but not quantity. The product quality is usually controlled by its color value but not purity. A number of attempts were made to determine the total amount and composition of crocetin derivatives from gardenia yellow [17,18]. Crocin was considered as an external reference sample in those studies. As an alternative, crocetin was also applied as an indicator to evaluate the composition of the crocetin derivatives [19,20].

A method to determine crocetin, its derivative amounts and composition from gardenia yellow was developed by UV-VIS and HPLC-MS-MS in this study based on their electronic absorption spectral characteristics and molecular mass. With this method, the crocetin, its derivative amounts and composition were able to be determined without the aid of external or internal references, even though the molecular structures of some derivatives were unknown.  

In industrial base, a method to determine the absorption coefficient, normally presented as color value, of gardenia yellow is currently applied for the product quality control scheme. It is expected that this approach will be replaced by the method developed in this study for the product quality control scheme. By this method, the color value will be substituted by the mass amount of the product for its quality control. This effort will provide a possibility to verify the biological functions and evaluate the safety of gardenia yellow product.

Methods and materials

1.1 Materials

Gardenia yellow product with the label of E500 was extracted from the fruits of Gardenia jasminoides Ellis by EtOH and provided by Henan Zhongda Hengyuan Biotechnology Co. Ltd. EtOH, trifluoroacetic acid (TFA), acetic acid was of AR grade and purchased from a local supplier. Acetonitrile was of HPLC grade and purchased from Dikma Technologies (USA). The water used to prepare the HPLC mobile phase was purified by counter-current osmosis. The crocin reference sample was purchased from Sigma (C/N: 17304-1G). Purity: >95% (W/W, HPLC).

1.2   Instrumentation

UV-VIS spectrophotometer: MultiSpec1500 (Shimadzu); LC-PDA-MS-MS: Thermo U-3000-Thermo LTQ Orbitrap XL equipped with PDA, the ion source of ACPI and the central data processing unit of Xcalibur (Thermo Fisher Scientific, USA).

1.3   Methods

1.3.1 The specific absorption coefficient determination of gardenia yellow by UV-VIS

This investigation aims to provide the E1cm1%

 (440±1 nm) of gardenia yellow. In practice, 0.0502 g gardenia yellow powders were accurately (0.0001 g) weighed and then completely dissolved in water to make the total volume of 50 mL. After the solution was diluted 100 times, the absorbance (A) of the solution was measured on a UV-VIS spectrophotometer at 440±1 nm, with water used as control.  The specific absorption coefficient of the product is calculated as follow:

https://auctoresonline.org/beta/uploads/articles/external/0-94130600-1625206400.png

Where

E1cm1%

 (440±5nm) = The specific absorption coefficient is defined as the absorbance of the product solution of 1 % concentration, i. e. 1 g in 100 mL, in a 1cm path-length cuvette.

A=Actual absorbance of the product solution

C= Concentration [g/mL] of the product solution

1.3.2 The crocetin derivative separation from gardenia yellow by C18-HPLC

An aliquot of gardenia yellow aqueous solution as described in Section 1.3.1 was filtered by 0.45 µm filter before dilution and immediately used for HPLC injection.

The chromatographic profile of gardenia yellow aqueous solution was acquired on C18-HPLC under the following conditions: DiamonsilTM C18 (4.6×250 mm, 5 mm) column; Mobile phase A = H2O-CH3CN-acetic acid at 74.95:25:0.05 (V/V/V); Mobile phase B = CH3CN; Linear gradient: Mobile phase B increased from 0 to 13.1% (V/V) in 20 minutes subsequently followed by an isocratic elution for 20 minutes; Flow rate = 0.8 mL/min; Sample injection volume = 20 µL; Column temperature = 25 ℃; PDA monitoring wavelength = 440 nm. The electronic absorption spectrum of each fraction was acquired from 250 to 700 nm by PDA. The peak area of each fraction was integrated.

1.3.3 The relative peak area measurement of each fraction

The relative peak area (%) of each crocetin derivative fraction is calculated according to formula (2).

Where

Relative peak area i= Relative peak area of crocetin derivative i in total crocetin derivative

Peak area i=Peak area of crocetin derivative i

i = Fraction number of crocetin derivative

n = Total number of crocetin derivative

1.3.4 The crocetin derivative fraction identification by MS

The MS characteristics of each crocetin derivative fraction were further acquired by the online MS detector under the following conditions: Spray voltage = -4.0 Kv; Sheath gas = 30 arb; Auxiliary gas = 10 arb; Purge gas = 0 arb; Capillary temperature = 350 ºC; Negative ion detection mode; The secondary mass spectra data acquisition: Data dependency scan; CID collision energy = 35%; Fragment mass scanning range = 150-2000 (m/z). The molecular ion peak was for each crocetin derivate was carefully identified and verified. The m/z value of each molecular ion was recorded.

The crocetin derivative fractions were identified by MS and the secondary MS according to the presence of crocetin ion = 327 [m/z], as the root structure of the molecules. 

1.3.5 The specific absorbance coefficient measurement of each crocetin derivative

The specific absorption coefficient of each crocetin derivative fraction is calculated from the published absorption coefficient, A1cm1%

 = 3820 (in EtOH), of crocetin as root structure, in accordance with the negative dependence of crocetin derivative UV-VIS absorbance on its side-chain quantity and mass indicated by molecular mass as shown in formula (3). The molecular mass of each crocetin derivative is hereon substituted by its measured parent ion [M-1]- mass reading [m/z] from MS.

https://www.auctoresonline.org/images/article/210503082122_4449587.png

Where

A1cmi1%

 = Absorbance coefficient of crocetin derivative i, defined as the theoretical absorbance of a 1% (W/V) crocetin derivative i solution in a cell of 1cm path-length at monitored 440nm.

A1cmCrocetin1%

 = Absorbance coefficient of crocetin, defined as the theoretical absorbance of a 1% (W/V) crocetin solution in a cell of 1cm path-length monitored at 440 nm. Published value of 3820 (in methanol) is applied hereon.

Molecular mass crocetin= Molecular mass of crocetin. Published value of 328 is applied hereon.

Molecular mass I = Molecular mass of crocetin derivative i. Molecular mass of each crocetin derivative is hereon substituted by its parent ion mass [m/z] from MS.

i= Fraction number of crocetin derivative

1.3.6 The specific absorbance coefficient measurement of total crocetin derivative

The specific absorption coefficient of total crocetin derivative is calculated by weighted sum from the relative peak area and absorption coefficients of each crocetin derivative as the results of formulae (2) and (3), respectively, according to formula (4).

https://auctoresonline.org/beta/uploads/articles/external/0-15714700-1625206401.png

Where

Relative peak area i= Relative peak area of crocetin derivative i

A1cmTotal crocetin derivative1%

 = Absorption coefficient of total crocetin derivative, defined as the theoretical absorbance of a 1% (W/V) total crocetin derivative solution in a cell of 1cm path-length monitored at 440nm.

A1cmi1%

 =Absorbance coefficient of crocetin derivative i, defined as the theoretical absorbance of a 1% (W/V) crocetin derivative i solution in a cell of 1cm path-length monitored at 440nm.

i= Fraction number of crocetin derivative

n= Total number of crocetin derivative

1.3.7 The relative content measurement of total crocetin derivative

The relative content of total crocetin derivative in total pigment is calculated from the peak area of each fraction monitored at 440 nm according to formula (5).

https://auctoresonline.org/beta/uploads/articles/external/0-23820900-1625206401.png

Where

Relative content Total crocetin derivative= Relative content of total crocetin derivative in total pigment

Peak area i= Peak area of crocetin derivative i

Peak area j= Peak area of fraction j

i = Fraction number of crocetin derivative

n = Total number of crocetin derivative

j = Fraction number

m = Total fraction number

1.3.8 Measurement of the total crocetin derivative amount

According to Lambert-Beer law, the amount of total crocetin derivative in gardenia yellow sample is calculated from the absorbance of the gardenia yellow aqueous solution, the relative content of total crocetin derivative in total pigment, and the specific absorption coefficient of total crocetin derivative as shown by formula (6).

https://auctoresonline.org/beta/uploads/articles/external/0-26877100-1625206401.png

​​Where

Amount Total crocetin derivative[g] = Amount of total crocetin derivative in the gardenia yellow sample

A = Absorbance of sample solution

V = Solvent volume for sample originally dissolving

N = Dilution fold of sample solution

Relative content Total crocetin derivative= Relative content of total crocetin derivative in total pigment

A1cmTotal crocetin derivative1%

 = Absorption coefficient of the total crocetin derivative, defined as the theoretical absorbance of a 1% (W/V) total crocetin derivative solution in a cell of 1cm path-length monitored at 440nm.

1.3.9 The relative content measurement of total crocetin derivative

The relative content of total crocetin derivative in the gardenia yellow sample is calculated from the amount of total crocetin derivative and sample weight according to formula (7).

https://auctoresonline.org/beta/uploads/articles/external/0-32398400-1625206401.png

​Where

Content Total crocetin derivative [%] =Content of total crocetin derivative in the gardenia yellow sample

Amount Total crocetin derivative[g] = Amount of total crocetin derivative in the gardenia yellow sample

M=Mass of the gardenia yellow sample

1.3.10 The relative content measurement of each crocetin derivative

According to Lambert-Beer law, the relative content of each crocetin derivative in total crocetin derivative is calculated from the peak area and absorption coefficients of each crocetin derivative as shown by formula (8).​​ 

https://www.auctoresonline.org/images/article/210503091228_8581441.png

​​Where

Relative content i= Relative content of each crocetin derivative in total crocetin derivative

Peak area i= Peak area of crocetin derivative i

i=Fraction number of crocetin derivative

n=Total number of crocetin derivative

1.3.11 Measuring the amount of each crocetin derivative

The amount of each crocetin derivative from the gardenia yellow sample is calculated from the relative content of each crocetin derivative and the total amount of crocetin derivative according to formula (9).

https://auctoresonline.org/beta/uploads/articles/external/0-38130900-1625206401.png

​Where

Amount i= Amount of each crocetin derivative from the gardenia yellow sample

Relative content i= Relative content of each crocetin derivative in total crocetin derivative

Amount Total crocetin derivative= Amount of total crocetin derivative in the gardenia yellow sample

i= Fraction number of crocetin derivative

1.3.12. The relative crocetin content measurement of each crocetin derivative

According to formula (10), the relative crocetin content of each crocetin derivative is calculated from the molecular masses of crocetin and its derivative, 328Da and the parent ion mass of the derivative respectively.

https://auctoresonline.org/beta/uploads/articles/external/0-40259600-1625206401.png

​Where

Relative crocetin content i= Relative crocetin content of each crocetin derivative

Crocetin molecular mass = 328Da

Crocetin derivative molecular mass i: The molecular mass of crocetin derivative i is hereon substituted by its parent ion [M-1]- mass value[m/z] from MS.

i = Fraction number of crocetin derivative

1.3.13 The relative crocetin content measurement from the total crocetin derivative

The relative crocetin content of total crocetin derivative is calculated from the relative crocetin content of each crocetin derivative and its relative peak area by weighted sum.

https://auctoresonline.org/beta/uploads/articles/external/0-44522500-1625206401.png

Where

Relative crocetin content Total crocetin derivative= Relative crocetin content of total crocetin derivative

Relative crocetin content i= Relative crocetin content of each crocetin derivative

Peak area i= Peak area of crocetin derivative i

i = Fraction number of crocetin derivative

1.3.14 The total crocetin amount measurement of gardenia yellow sample

The total crocetin amount of gardenia yellow sample is calculated from the relative crocetin content of total crocetin derivative and the amount of total crocetin derivative.

https://auctoresonline.org/beta/uploads/articles/external/0-47972800-1625206401.png

​​Where

Relative crocetin content Total crocetin derivative= Relative crocetin content of total crocetin derivative

Amount Total crocetin derivative= Amount of total crocetin derivative in the gardenia yellow sample

1.3.15 Recovery, reproducibility and minimum detection limit

The amount equal to the crocin amount from gardenia yellow product adduced from external references was added to the sample before the analysis for the recovery, reproducibility and RSD assessment of the method. Additionally, minimum detection limit is also detected with added crocin.

The final result is calculated from the arithmetic mean of parallel measurements. The absolute difference of two independent observations is not more than 5% of their arithmetic mean under the same conditions.

Results

2.1 The specific absorption coefficient determination of gardenia yellow by UV-VIS

The absorbance (A) of the diluted gardenia yellow aqueous solution at the concentration of 10.04mg/mL was 0.5064 at 440nm. The specific absorption coefficient at 440nm (E1cm1%

 ) of gardenia yellow was 504.38, calculated according to formula (1).

2.2 The C18-HPLC profile of Gardenia Yellow

Figure 2 illustrates the C18-HPLC profile of gardenia yellow. It was observed from Figure 2 that the crocetin and its derivatives were well separated under the conditions employed in this study. The crocin fraction was identified in comparison of its retention time = 7.093 min, its lmax= 440 nm and the fine structure of its electronic absorption spectrum with those of external reference sample.

Figure 2. The C18-HPLC profile of gardenia yellow. HPLC conditions: as shown in Section 1.3.2.

2.3 The fraction identification of gardenia yellow by HPLC-MS-MS

Through MS and the secondary MS, the crocetin derivative fractions were carefully identified as the root structure of the molecules from the presence of crocetin ion = 327[m/z]. Under the conditions of ionization employed in this study, the major fragmentation taken placed during the MS process was the breaking of glycoside bonds. As an example, Figure 3 gives the TIC (a) of gardenia yellow and the mass spectrum (b) and secondary mass spectrum (c) of fraction at 7.093min. The ions 1089.34(100), 975.35 and 651.25 were seen from Figure 3(b). The ion 1089.34 was identified as parent ion [M-1]-. The presence of ions 975.35 and 651.25 suggested that the parent ion consisted of gentibioside-crocetin-gentibioside as the root structure. This was further verified by the presence of ions 1089.34(100), 975.35, 651.25 and 327.15 from the secondary spectrum in Figure 3(c). The fraction would therefore be considered as a crocetin derivative.

Figure 3. MS characteristics of fraction at 7.093 min from gardenia yellow. (a)The TIC of gardenia yellow.                                      (b)The mass spectrum of fraction at 7.093 min
Figure 3(c). The secondary mass spectrum of fraction at 7.093 min. HPLC-MS-MS conditions: as shown in Sections 1.3.2 and 1.3.4.

2.4 The relative peak area of each fraction on the HPLC chromatogram of gardenia yellow

It is observed from Figure 2 that each fraction of gardenia yellow sample was readily separated under HPLC conditions as mentioned above. The peak area of each fraction was integrated. The relative peak area of each fraction was then calculated and given in Table 1.

Table 1. The chromatographic behaviors, MS characteristics and amounts of each crocetin derivative fraction from gardenia yellow.

2.5 Crocetin derivative fraction identification by MS

The crocetin derivative fractions were identified by MS as the root structure of the molecules according to the presence of crocetin ion = 327 m/z. The fractions containing crocetin structure are given in Table 1. The parent ion m/z value of each crocetin derivative is also recorded in Table 1.

2.6 The specific absorbance coefficient of each crocetin derivative

With reference to the published specific absorption coefficient of crocetin, the specific absorption coefficient of each crocetin derivative fraction is calculated in accordance with the negative dependence of crocetin derivative UV-VIS absorbance on its side-chain quantity and mass indicated by molecular mass given in Table 1.

2.7 The specific absorbance coefficient of total crocetin derivative

The specific absorption coefficient of the total crocetin derivative was calculated by the weighted sum from the relative peak area and the specific absorption coefficients of each crocetin derivative to be 1125.

2.8 The relative content of total crocetin derivative

The relative content of total crocetin derivative in total pigment is calculated to be 97.20%, from the peak area of each fraction monitored at 440 nm.

2.9 Total crocetin derivative amount    

The amount of total crocetin derivative in gardenia yellow sample is calculated to be 0.0225 g from the absorbance of gardenia yellow aqueous solution (measured by UV-VIS), the relative content of total crocetin derivative in total pigment and the specific absorption coefficient of total crocetin derivative.  

2.10 The relative content of total crocetin derivative from gardenia yellow

The relative content of total crocetin derivative in the gardenia yellow sample is calculated to be 44.82% (W/W) from the amount of total crocetin derivative and sample weight.

2.11 The relative content of each crocetin derivative

According to Lambert-Beer law, the relative content of each crocetin derivative in total crocetin derivative is calculated from the peak area and absorption coefficients of each crocetin derivative and illustrated in Table 1.

2.12 The amount of each crocetin derivative

The amount of each crocetin derivative from the gardenia yellow sample is calculated from the relative content of each crocetin derivative and the amount of total crocetin derivative and is given in Table 1.

2.13 The relative crocetin content of each crocetin derivative

The relative crocetin content of each crocetin derivative is calculated from the molecular masses of crocetin and its derivative, 328Da and the parent ion mass of the derivative respectively. The relative crocetin content of each crocetin derivative is given in Table 1.

2.14 The relative crocetin content of total crocetin derivative

The relative crocetin content of total crocetin derivative is calculated to be 29.35% from the relative crocetin content of each crocetin derivative and its relative peak area by weighted sum.

2.15 The total crocetin amount of gardenia yellow sample

From the relative crocetin content of total crocetin derivative and the amount of total crocetin derivative, the total crocetin amount of gardenia yellow is calculated to be 0.0066 g in the 0.0502 g gardenia yellow sample. The relative crocetin amount of gardenia yellow is 13.15% (W/W).

2.16 Precision and accuracy

To evaluate the precision and accuracy of the method as described above, 0.0115 g external reference sample of crocin (equal to the amount of crocin derivative fraction at 7.093 min from gardenia yellow sample) was added to the 0.0502 g gardenia yellow sample. The recovery, reproducibility and relative standard deviation (RSD) of added external reference sample were consequently assessed. Additionally, minimum detection limit was also detected.

It was observed that the RSD of repeated assessments for more than 6 times was less than 3.17%. This observation suggested that the method exhibited good reproducibility. Additionally, the recovery of added crocin was more than 97±1.61%. The minimum detection limit of crocin was 7.44μg/mL.

Conclusion

The major color materials from gardenia yellow are a group of crocetin derivatives. The molecular structures of some derivatives are well elucidated while those of the others are unknown and their commercial refence samples are short. The total amount and composition determinations of crocetin derivative are therefore currently difficult. The method developed in this study provides a possibility to assess each of the total derivative amounts through their absorption coefficients, although the molecular structures of some derivatives are unknown.

References

Clearly Auctoresonline and particularly Psychology and Mental Health Care Journal is dedicated to improving health care services for individuals and populations. The editorial boards' ability to efficiently recognize and share the global importance of health literacy with a variety of stakeholders. Auctoresonline publishing platform can be used to facilitate of optimal client-based services and should be added to health care professionals' repertoire of evidence-based health care resources.

img

Virginia E. Koenig

Journal of Clinical Cardiology and Cardiovascular Intervention The submission and review process was adequate. However I think that the publication total value should have been enlightened in early fases. Thank you for all.

img

Delcio G Silva Junior

Journal of Women Health Care and Issues By the present mail, I want to say thank to you and tour colleagues for facilitating my published article. Specially thank you for the peer review process, support from the editorial office. I appreciate positively the quality of your journal.

img

Ziemlé Clément Méda

Journal of Clinical Research and Reports I would be very delighted to submit my testimonial regarding the reviewer board and the editorial office. The reviewer board were accurate and helpful regarding any modifications for my manuscript. And the editorial office were very helpful and supportive in contacting and monitoring with any update and offering help. It was my pleasure to contribute with your promising Journal and I am looking forward for more collaboration.

img

Mina Sherif Soliman Georgy

We would like to thank the Journal of Thoracic Disease and Cardiothoracic Surgery because of the services they provided us for our articles. The peer-review process was done in a very excellent time manner, and the opinions of the reviewers helped us to improve our manuscript further. The editorial office had an outstanding correspondence with us and guided us in many ways. During a hard time of the pandemic that is affecting every one of us tremendously, the editorial office helped us make everything easier for publishing scientific work. Hope for a more scientific relationship with your Journal.

img

Layla Shojaie

The peer-review process which consisted high quality queries on the paper. I did answer six reviewers’ questions and comments before the paper was accepted. The support from the editorial office is excellent.

img

Sing-yung Wu

Journal of Neuroscience and Neurological Surgery. I had the experience of publishing a research article recently. The whole process was simple from submission to publication. The reviewers made specific and valuable recommendations and corrections that improved the quality of my publication. I strongly recommend this Journal.

img

Orlando Villarreal

Dr. Katarzyna Byczkowska My testimonial covering: "The peer review process is quick and effective. The support from the editorial office is very professional and friendly. Quality of the Clinical Cardiology and Cardiovascular Interventions is scientific and publishes ground-breaking research on cardiology that is useful for other professionals in the field.

img

Katarzyna Byczkowska

Thank you most sincerely, with regard to the support you have given in relation to the reviewing process and the processing of my article entitled "Large Cell Neuroendocrine Carcinoma of The Prostate Gland: A Review and Update" for publication in your esteemed Journal, Journal of Cancer Research and Cellular Therapeutics". The editorial team has been very supportive.

img

Anthony Kodzo-Grey Venyo

Testimony of Journal of Clinical Otorhinolaryngology: work with your Reviews has been a educational and constructive experience. The editorial office were very helpful and supportive. It was a pleasure to contribute to your Journal.

img

Pedro Marques Gomes

Dr. Bernard Terkimbi Utoo, I am happy to publish my scientific work in Journal of Women Health Care and Issues (JWHCI). The manuscript submission was seamless and peer review process was top notch. I was amazed that 4 reviewers worked on the manuscript which made it a highly technical, standard and excellent quality paper. I appreciate the format and consideration for the APC as well as the speed of publication. It is my pleasure to continue with this scientific relationship with the esteem JWHCI.

img

Bernard Terkimbi Utoo

This is an acknowledgment for peer reviewers, editorial board of Journal of Clinical Research and Reports. They show a lot of consideration for us as publishers for our research article “Evaluation of the different factors associated with side effects of COVID-19 vaccination on medical students, Mutah university, Al-Karak, Jordan”, in a very professional and easy way. This journal is one of outstanding medical journal.

img

Prof Sherif W Mansour

Dear Hao Jiang, to Journal of Nutrition and Food Processing We greatly appreciate the efficient, professional and rapid processing of our paper by your team. If there is anything else we should do, please do not hesitate to let us know. On behalf of my co-authors, we would like to express our great appreciation to editor and reviewers.

img

Hao Jiang

As an author who has recently published in the journal "Brain and Neurological Disorders". I am delighted to provide a testimonial on the peer review process, editorial office support, and the overall quality of the journal. The peer review process at Brain and Neurological Disorders is rigorous and meticulous, ensuring that only high-quality, evidence-based research is published. The reviewers are experts in their fields, and their comments and suggestions were constructive and helped improve the quality of my manuscript. The review process was timely and efficient, with clear communication from the editorial office at each stage. The support from the editorial office was exceptional throughout the entire process. The editorial staff was responsive, professional, and always willing to help. They provided valuable guidance on formatting, structure, and ethical considerations, making the submission process seamless. Moreover, they kept me informed about the status of my manuscript and provided timely updates, which made the process less stressful. The journal Brain and Neurological Disorders is of the highest quality, with a strong focus on publishing cutting-edge research in the field of neurology. The articles published in this journal are well-researched, rigorously peer-reviewed, and written by experts in the field. The journal maintains high standards, ensuring that readers are provided with the most up-to-date and reliable information on brain and neurological disorders. In conclusion, I had a wonderful experience publishing in Brain and Neurological Disorders. The peer review process was thorough, the editorial office provided exceptional support, and the journal's quality is second to none. I would highly recommend this journal to any researcher working in the field of neurology and brain disorders.

img

Dr Shiming Tang

Dear Agrippa Hilda, Journal of Neuroscience and Neurological Surgery, Editorial Coordinator, I trust this message finds you well. I want to extend my appreciation for considering my article for publication in your esteemed journal. I am pleased to provide a testimonial regarding the peer review process and the support received from your editorial office. The peer review process for my paper was carried out in a highly professional and thorough manner. The feedback and comments provided by the authors were constructive and very useful in improving the quality of the manuscript. This rigorous assessment process undoubtedly contributes to the high standards maintained by your journal.

img

Raed Mualem

International Journal of Clinical Case Reports and Reviews. I strongly recommend to consider submitting your work to this high-quality journal. The support and availability of the Editorial staff is outstanding and the review process was both efficient and rigorous.

img

Andreas Filippaios

Thank you very much for publishing my Research Article titled “Comparing Treatment Outcome Of Allergic Rhinitis Patients After Using Fluticasone Nasal Spray And Nasal Douching" in the Journal of Clinical Otorhinolaryngology. As Medical Professionals we are immensely benefited from study of various informative Articles and Papers published in this high quality Journal. I look forward to enriching my knowledge by regular study of the Journal and contribute my future work in the field of ENT through the Journal for use by the medical fraternity. The support from the Editorial office was excellent and very prompt. I also welcome the comments received from the readers of my Research Article.

img

Dr Suramya Dhamija

Dear Erica Kelsey, Editorial Coordinator of Cancer Research and Cellular Therapeutics Our team is very satisfied with the processing of our paper by your journal. That was fast, efficient, rigorous, but without unnecessary complications. We appreciated the very short time between the submission of the paper and its publication on line on your site.

img

Bruno Chauffert

I am very glad to say that the peer review process is very successful and fast and support from the Editorial Office. Therefore, I would like to continue our scientific relationship for a long time. And I especially thank you for your kindly attention towards my article. Have a good day!

img

Baheci Selen

"We recently published an article entitled “Influence of beta-Cyclodextrins upon the Degradation of Carbofuran Derivatives under Alkaline Conditions" in the Journal of “Pesticides and Biofertilizers” to show that the cyclodextrins protect the carbamates increasing their half-life time in the presence of basic conditions This will be very helpful to understand carbofuran behaviour in the analytical, agro-environmental and food areas. We greatly appreciated the interaction with the editor and the editorial team; we were particularly well accompanied during the course of the revision process, since all various steps towards publication were short and without delay".

img

Jesus Simal-Gandara

I would like to express my gratitude towards you process of article review and submission. I found this to be very fair and expedient. Your follow up has been excellent. I have many publications in national and international journal and your process has been one of the best so far. Keep up the great work.

img

Douglas Miyazaki

We are grateful for this opportunity to provide a glowing recommendation to the Journal of Psychiatry and Psychotherapy. We found that the editorial team were very supportive, helpful, kept us abreast of timelines and over all very professional in nature. The peer review process was rigorous, efficient and constructive that really enhanced our article submission. The experience with this journal remains one of our best ever and we look forward to providing future submissions in the near future.

img

Dr Griffith

I am very pleased to serve as EBM of the journal, I hope many years of my experience in stem cells can help the journal from one way or another. As we know, stem cells hold great potential for regenerative medicine, which are mostly used to promote the repair response of diseased, dysfunctional or injured tissue using stem cells or their derivatives. I think Stem Cell Research and Therapeutics International is a great platform to publish and share the understanding towards the biology and translational or clinical application of stem cells.

img

Dr Tong Ming Liu

I would like to give my testimony in the support I have got by the peer review process and to support the editorial office where they were of asset to support young author like me to be encouraged to publish their work in your respected journal and globalize and share knowledge across the globe. I really give my great gratitude to your journal and the peer review including the editorial office.

img

Husain Taha Radhi

I am delighted to publish our manuscript entitled "A Perspective on Cocaine Induced Stroke - Its Mechanisms and Management" in the Journal of Neuroscience and Neurological Surgery. The peer review process, support from the editorial office, and quality of the journal are excellent. The manuscripts published are of high quality and of excellent scientific value. I recommend this journal very much to colleagues.

img

S Munshi

Dr.Tania Muñoz, My experience as researcher and author of a review article in The Journal Clinical Cardiology and Interventions has been very enriching and stimulating. The editorial team is excellent, performs its work with absolute responsibility and delivery. They are proactive, dynamic and receptive to all proposals. Supporting at all times the vast universe of authors who choose them as an option for publication. The team of review specialists, members of the editorial board, are brilliant professionals, with remarkable performance in medical research and scientific methodology. Together they form a frontline team that consolidates the JCCI as a magnificent option for the publication and review of high-level medical articles and broad collective interest. I am honored to be able to share my review article and open to receive all your comments.

img

Tania Munoz

“The peer review process of JPMHC is quick and effective. Authors are benefited by good and professional reviewers with huge experience in the field of psychology and mental health. The support from the editorial office is very professional. People to contact to are friendly and happy to help and assist any query authors might have. Quality of the Journal is scientific and publishes ground-breaking research on mental health that is useful for other professionals in the field”.

img

George Varvatsoulias

Dear editorial department: On behalf of our team, I hereby certify the reliability and superiority of the International Journal of Clinical Case Reports and Reviews in the peer review process, editorial support, and journal quality. Firstly, the peer review process of the International Journal of Clinical Case Reports and Reviews is rigorous, fair, transparent, fast, and of high quality. The editorial department invites experts from relevant fields as anonymous reviewers to review all submitted manuscripts. These experts have rich academic backgrounds and experience, and can accurately evaluate the academic quality, originality, and suitability of manuscripts. The editorial department is committed to ensuring the rigor of the peer review process, while also making every effort to ensure a fast review cycle to meet the needs of authors and the academic community. Secondly, the editorial team of the International Journal of Clinical Case Reports and Reviews is composed of a group of senior scholars and professionals with rich experience and professional knowledge in related fields. The editorial department is committed to assisting authors in improving their manuscripts, ensuring their academic accuracy, clarity, and completeness. Editors actively collaborate with authors, providing useful suggestions and feedback to promote the improvement and development of the manuscript. We believe that the support of the editorial department is one of the key factors in ensuring the quality of the journal. Finally, the International Journal of Clinical Case Reports and Reviews is renowned for its high- quality articles and strict academic standards. The editorial department is committed to publishing innovative and academically valuable research results to promote the development and progress of related fields. The International Journal of Clinical Case Reports and Reviews is reasonably priced and ensures excellent service and quality ratio, allowing authors to obtain high-level academic publishing opportunities in an affordable manner. I hereby solemnly declare that the International Journal of Clinical Case Reports and Reviews has a high level of credibility and superiority in terms of peer review process, editorial support, reasonable fees, and journal quality. Sincerely, Rui Tao.

img

Rui Tao

Clinical Cardiology and Cardiovascular Interventions I testity the covering of the peer review process, support from the editorial office, and quality of the journal.

img

Khurram Arshad

Clinical Cardiology and Cardiovascular Interventions, we deeply appreciate the interest shown in our work and its publication. It has been a true pleasure to collaborate with you. The peer review process, as well as the support provided by the editorial office, have been exceptional, and the quality of the journal is very high, which was a determining factor in our decision to publish with you.

img

Gomez Barriga Maria Dolores

The peer reviewers process is quick and effective, the supports from editorial office is excellent, the quality of journal is high. I would like to collabroate with Internatioanl journal of Clinical Case Reports and Reviews journal clinically in the future time.

img

Lin Shaw Chin

Clinical Cardiology and Cardiovascular Interventions, I would like to express my sincerest gratitude for the trust placed in our team for the publication in your journal. It has been a true pleasure to collaborate with you on this project. I am pleased to inform you that both the peer review process and the attention from the editorial coordination have been excellent. Your team has worked with dedication and professionalism to ensure that your publication meets the highest standards of quality. We are confident that this collaboration will result in mutual success, and we are eager to see the fruits of this shared effort.

img

Maria Dolores Gomez Barriga

Dear Dr. Jessica Magne, Editorial Coordinator 0f Clinical Cardiology and Cardiovascular Interventions, I hope this message finds you well. I want to express my utmost gratitude for your excellent work and for the dedication and speed in the publication process of my article titled "Navigating Innovation: Qualitative Insights on Using Technology for Health Education in Acute Coronary Syndrome Patients." I am very satisfied with the peer review process, the support from the editorial office, and the quality of the journal. I hope we can maintain our scientific relationship in the long term.

img

Dr Maria Dolores Gomez Barriga

Dear Monica Gissare, - Editorial Coordinator of Nutrition and Food Processing. ¨My testimony with you is truly professional, with a positive response regarding the follow-up of the article and its review, you took into account my qualities and the importance of the topic¨.

img

Dr Maria Regina Penchyna Nieto

Dear Dr. Jessica Magne, Editorial Coordinator 0f Clinical Cardiology and Cardiovascular Interventions, The review process for the article “The Handling of Anti-aggregants and Anticoagulants in the Oncologic Heart Patient Submitted to Surgery” was extremely rigorous and detailed. From the initial submission to the final acceptance, the editorial team at the “Journal of Clinical Cardiology and Cardiovascular Interventions” demonstrated a high level of professionalism and dedication. The reviewers provided constructive and detailed feedback, which was essential for improving the quality of our work. Communication was always clear and efficient, ensuring that all our questions were promptly addressed. The quality of the “Journal of Clinical Cardiology and Cardiovascular Interventions” is undeniable. It is a peer-reviewed, open-access publication dedicated exclusively to disseminating high-quality research in the field of clinical cardiology and cardiovascular interventions. The journal's impact factor is currently under evaluation, and it is indexed in reputable databases, which further reinforces its credibility and relevance in the scientific field. I highly recommend this journal to researchers looking for a reputable platform to publish their studies.

img

Dr Marcelo Flavio Gomes Jardim Filho

Dear Editorial Coordinator of the Journal of Nutrition and Food Processing! "I would like to thank the Journal of Nutrition and Food Processing for including and publishing my article. The peer review process was very quick, movement and precise. The Editorial Board has done an extremely conscientious job with much help, valuable comments and advices. I find the journal very valuable from a professional point of view, thank you very much for allowing me to be part of it and I would like to participate in the future!”

img

Zsuzsanna Bene

Dealing with The Journal of Neurology and Neurological Surgery was very smooth and comprehensive. The office staff took time to address my needs and the response from editors and the office was prompt and fair. I certainly hope to publish with this journal again.Their professionalism is apparent and more than satisfactory. Susan Weiner

img

Dr Susan Weiner