Name: Fast Inspection of Quality of Indigo Naturalis by Multiple Light Scattering (Video) | JoVE Journal
Uploaded: 2023-08-18T12:10:00.000+00:00
Duration: 3 min 40 s
Description: Learn rapid quality assessment of Indigo Naturalis using multiple light scattering for traditional Chinese medicine. This non-invasive method ensures authenticity without altering samples or using harmful reagents.

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5 minutes

The work wishes to acknowledge support from the National Natural Science Foundation of China (No. 82173976), National Key Research and Development Program (No. 2018YFC1707205), and State Key Laboratory of Innovative Drugs and High Energy Saving Pharmaceutical Equipment, Jiangxi University of Traditional Chinese Medicine (No. GZSYS202003).

1. Preparation of test sample
<ol>
	<li>Set up four different batches of Indigo Naturalis herbal powder for the test. Pass each sample through the seventh sieve and ninth sieve in turn and collect the sample between the seventh and ninth sieve5. 
	NOTE: The average aperture size of the seventh sieve is 125 &#181;m &#177; 5.8 &#181;m. The average aperture size of the ninth sieve is 75 &#181;m &#177; 4.1 &#181;m.</li>
	<li>Accurately weigh 0.2 g of the sample (Indigo Naturalis powder) on a weighing paper and set it aside.</li>
</ol>
2. Sample addition
<ol>
	<li>Build an iron support stand and put an iron ring with a 5 cm diameter funnel on it.</li>
	<li>Use a pipette to add 20 mL of pure water to the sample glass bottle (bottom diameter 2.6 cm, height 6 cm). Place the sample glass bottle directly under the funnel so that the lower edge of the funnel is flush with the mouth of the bottle. 
	NOTE: Clean the exterior of the sample glass vial with a clean, non-abrasive paper towel and inspect the glass surface for visible marks. If there are, change the glass bottle. Be careful not to spill when adding liquid.</li>
	<li>Release the sample at a height of 80 cm from the lower edge of the funnel so that it can slide freely along the funnel into the sample bottle.</li>
</ol>
3. Instrument operation
<ol>
	<li>Turn on the Turbiscan Lab instrument and warm it up for 30 min.</li>
	<li>Create the file. Click the Create file button in the top menu (or the New file function in the File menu) to create a new empty measurement file. Define its name and save location (by default, the data folder is located at: &#34;C:/users/admin/Formulaction/FAnalyser/Data&#34;.</li>
	<li>Click the Show Turbiscan Lab Temperature button in the top menu to set the instrument target temperature to 25 &#176;C. 
	NOTE: The instrument temperature is influenced by the room temperature, so be careful to adjust the ambient temperature.</li>
	<li>Click Program Scan in the top menu to enter the setup analysis program. Add the program to the list, and in the taskbar, add 30 s as a cycle and 21 scans to the analysis sequence. Select this analysis program for all subsequent measurements.</li>
	<li>Move the prepared sample vial into the measurement system. After setting up the program, click Start to start the measurement. 
	NOTE: Be careful not to shake the glass bottle when moving and only move it slightly.</li>
	<li>After data acquisition, click on the list of calculated parameters to automatically calculate TSI.</li>
</ol>

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<li>Liu, X. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Establishment+and+application+of+a+rapid+quality+inspection+method+for+Indigo+Naturalis+based+on+quantitative+portrayal+of+water+testing+process%5BTitle%5D)+AND+%22Acta+Pharmacologica+Sinica%22%5BJournal%5D)">Establishment and application of a rapid quality inspection method for Indigo Naturalis based on quantitative portrayal of water testing process.</a> Acta Pharmacologica Sinica. 57 (11), 3411-3418 (2022).</li>
<li>Mengual, O., Meunier, G., Cayre, I., Puech, K., Snabre, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Characterisation+of+instability+of+concentrated+dispersions+by+a+new+optical+analyser%3A+the+TURBISCAN+MA+1000%5BTitle%5D)+AND+%22Colloids+and+Surfaces+A%3A+Physicochemical+and+Engineering+Aspects%22%5BJournal%5D)">Characterisation of instability of concentrated dispersions by a new optical analyser: the TURBISCAN MA 1000.</a> Colloids and Surfaces A: Physicochemical and Engineering Aspects. 152 (1), 111-123 (1999).</li>
<li>Olatunji, O. N., Du, J., Hintz, W., Tomas, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Application+of+particle+sedimentation+analysis+in+sterically-stabilized+TiO2+particles+stability+assessment%5BTitle%5D)+AND+%22Advanced+Powder+Technology%22%5BJournal%5D)">Application of particle sedimentation analysis in sterically-stabilized TiO2 particles stability assessment.</a> Advanced Powder Technology. 27 (4), 1325-1336 (2016).</li>
<li>Ferrentino, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Fining+of+red+wine+monitored+by+multiple+light+scattering%5BTitle%5D)+AND+%22Journal+of+Agricultural+and+Food+Chemistry%22%5BJournal%5D)">Fining of red wine monitored by multiple light scattering.</a> Journal of Agricultural and Food Chemistry. 65 (27), 5523-5530 (2017).</li>
<li>Ramezani, M., Ferrentino, G., Morozova, K., Scampicchio, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Multiple+light+scattering+measurements+for+online+monitoring+of+milk+fermentation%5BTitle%5D)+AND+%22Foods%22%5BJournal%5D)">Multiple light scattering measurements for online monitoring of milk fermentation.</a> Foods. 10 (7), 1582(2021).</li>
<li>Yang, H. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+new+approach+to+evaluate+the+particle+growth+and+sedimentation+of+dispersed+polymer+microsphere+profile+control+system+based+on+multiple+light+scattering%5BTitle%5D)+AND+%22Powder+Technology%22%5BJournal%5D)">A new approach to evaluate the particle growth and sedimentation of dispersed polymer microsphere profile control system based on multiple light scattering.</a> Powder Technology. 315, 477-485 (2017).</li>
<li>Zhang, X. X., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Treatment+of+non-high-risk+acute+promyelocytic+leukemia+with+realgar-indigo+naturalis+formula+%28RIF%29+and+all-trans+retinoid+acid+%28ATRA%29%3A+study+protocol+for+a+randomized+controlled+trial%5BTitle%5D)+AND+%22Trials%22%5BJournal%5D)">Treatment of non-high-risk acute promyelocytic leukemia with realgar-indigo naturalis formula (RIF) and all-trans retinoid acid (ATRA): study protocol for a randomized controlled trial.</a> Trials. 21 (1), 7(2020).</li>
<li>Sugimoto, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Clinical+efficacy+and+safety+of+oral+Qing-Dai+in+patients+with+ulcerative+colitis%3A+a+single-center+open-label+prospective+study%5BTitle%5D)+AND+%22Digestion%22%5BJournal%5D)">Clinical efficacy and safety of oral Qing-Dai in patients with ulcerative colitis: a single-center open-label prospective study.</a> Digestion. 93 (3), 193-201 (2016).</li>
<li>Lin, Y. K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Clinical+assessment+of+patients+with+recalcitrant+psoriasis+in+a+randomized%2C+observer-blind%2C+vehicle-controlled+trial+using+indigo+naturalis%5BTitle%5D)+AND+%22Archives+of+Dermatology%22%5BJournal%5D)">Clinical assessment of patients with recalcitrant psoriasis in a randomized, observer-blind, vehicle-controlled trial using indigo naturalis.</a> Archives of Dermatology. 144 (11), 1457-1464 (2008).</li>
<li>Sun, Q., Leng, J., Tang, L., Wang, L., Fu, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+Comprehensive+review+of+the+chemistry%2C+pharmacokinetics%2C+pharmacology%2C+clinical+applications%2C+adverse+events%2C+and+quality+control+of+indigo+naturalis%5BTitle%5D)+AND+%22Frontiers+in+Pharmacology%22%5BJournal%5D)">A Comprehensive review of the chemistry, pharmacokinetics, pharmacology, clinical applications, adverse events, and quality control of indigo naturalis.</a> Frontiers in Pharmacology. 12, 664022(2021).</li>
<li>Yang, Y. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Investigation+and+analysis+of+the+commodity+quality+of+Indigo+Naturalis+herbs+in+Beijing+area%5BTitle%5D)+AND+%22Lishizhen+Medicine+and+Materia+Medica+Research%22%5BJournal%5D)">Investigation and analysis of the commodity quality of Indigo Naturalis herbs in Beijing area.</a> Lishizhen Medicine and Materia Medica Research. 23 (07), 1787-1788 (2012).</li>
<li>Yao, Z. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Comparative+study+of+thirty-eight+batches+of+indigo+naturalis%5BTitle%5D)+AND+%22Journal+of+Chengdu+University+of+TCM%22%5BJournal%5D)">Comparative study of thirty-eight batches of indigo naturalis.</a> Journal of Chengdu University of TCM. 34 (02), 86-88 (2011).</li>
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</ol>

The authors have no conflicts of interest to disclose.

According to traditional Chinese medicine, Indigo Naturalis has the power to clear heat and detoxification, cool blood, eliminate spots, purge fire, and arrest convulsion. Based on randomized double-blind controlled clinical trials11,12,13, Indigo Naturalis is effective in the treatment of psoriasis, ulcerative colitis, and acute promyelocytic leukemia in addition to its traditional treatment of cough and phlegm, hemorrhagic sym...

In traditional Chinese medicine, during the course of disease treatment, the clinical effectiveness of medications and the safety of the course of treatment are directly impacted by the quality of Chinese herbal medicine. Utilizing cutting-edge identification technology, one can assess the efficacy of Chinese herbal medicine and guarantee user safety. The Chinese herbal medicine water test method refers to immersing the herbs in water or solvent, then quickly and precisely determining the medicine&#39;s authenticity by observing the changes in color, size, and shape1.
It was originally a good choice for the identification of Chinese medicine. However, the disadvantage of the traditional water test method is that the accuracy and sensitivity of distinguishing the authenticity of Chinese medicine are low because of the subjectivity of observation by the naked eye2. One of the key medicinal materials used in the water test method is Indigo Naturalis, considered an effective remedy for pediatric hyperthermia, psoriasis, leukemia, and ulcerative colitis3. The genuine Indigo Naturalis floats on the surface of water, and the water does not turn dark blue after shaking. However, the fake Indigo Naturalis has particles that sink, and the water will turn dark blue after shaking4. Its principle is due to hydrophobic and easily floatable indigo, indirubin, and other organic components of high-quality Indigo Naturalis. On the contrary, due to low organic matter, a large amount of lime, and heavy texture, some particles doped with fake Indigo Naturalis will sink quickly5. However, this method is only a simple qualitative identification, and it limits the rapid identification of the authenticity of Chinese herbal medicine and fails to reveal the changes of Indigo Naturalis in water.
Multiple light scattering technology is a technology that can measure multi-angle light signal scanning based on a laser passing through the sample. The incident light will be scattered when it penetrates the sample or encounters particles. If the scattered light penetrates through the sample, a transmission light signal is formed; If the sample concentration is high, the light will be reflected by the particles, forming a backscattering light signal. The changes in light intensity reflect the changes in particle concentration and particle size in liquid preparation6. The multiple light scattering instruments can quickly and accurately analyze phenomena such as emulsification, flocculation, precipitation, and rupture of suspension, emulsion, and foam liquid by multiple light scattering technology, as well as quantitatively analyze characteristics such as the rate of occurrence of the above phenomena.
Multiple light scattering technology has demonstrated significant benefits in particle stability monitoring7, red wine clarification8, and milk fermentation quality control9. Using this technology, the traditional water test method of Indigo Naturalis may be intuitive, quantitative, and scientific. Therefore, based on the principle of multiple light scattering technology, this study established a rapid evaluation method for the quality of Indigo Naturalis, taking the Turbiscan stability index (TSI) of the sample as the index of quality control (Figure 1).

<table><tbody><tr><td>Analytical balance (1/10,000)</td><td>Sartorious, Germany</td><td>BSA224S&amp;nbsp;</td><td>www.sartorius.com.cn</td></tr><tr><td>Funnel</td><td>Chengdu Kelong Chemical Co. LTD</td><td>Diameter 5 cm</td><td>www.cdkelongchem.com</td></tr><tr><td>Indigo Naturalis S1</td><td>Xianyou, Fujian</td><td>20210501</td><td></td></tr><tr><td>Indigo Naturalis S2</td><td>Yaan, Sichuan</td><td>20201102</td><td></td></tr><tr><td>Indigo Naturalis S3</td><td>Xianyou, Fujian</td><td>20161012</td><td></td></tr><tr><td>Indigo Naturalis S4</td><td>Xianyou, Fujian</td><td>20180305</td><td></td></tr><tr><td>&amp;nbsp;Iron ring</td><td>Chengdu Kelong Chemical Co. LTD</td><td>/</td><td>www.cdkelongchem.com</td></tr><tr><td>Iron stand</td><td>Chengdu Kelong Chemical Co. LTD</td><td>/</td><td>www.cdkelongchem.com</td></tr><tr><td>Mili-Q ultra-pure water meter</td><td>Milipore, USA</td><td>Mili-Q</td><td>www.merckmillipore.com</td></tr><tr><td>Ninth sieve</td><td>Chengdu Kelong Chemical Co. LTD</td><td>Average aperture size 75 &amp;micro;m</td><td>www.cdkelongchem.com</td></tr><tr><td>Sample bottle</td><td>French Formulaction Company</td><td>Bottom diameter 2.6 cm, height 6 cm</td><td>www.formulaction.com</td></tr><tr><td>Seventh sieve</td><td>Chengdu Kelong Chemical Co. LTD</td><td>Average aperture size 125 &amp;micro;m</td><td>www.cdkelongchem.com</td></tr><tr><td>Turbisoft Lab multiple light scattering instrument</td><td>French Formulaction Company</td><td>Turbisoft Lab 2.3.1.125 Fanalyser 1.3.5</td><td>www.formulaction.com</td></tr><tr><td>Weighing paper</td><td>Chengdu Kelong Chemical Co. LTD</td><td>/</td><td>www.cdkelongchem.com</td></tr></tbody></table>

fast inspection of quality of indigo naturalis by multiple light scattering

Fast Inspection of Quality of Indigo Naturalis by Multiple Light Scattering

Quality control of Chinese herbal medicine is a crucial component of Chinese herbal medicine research and development. Faced with the challenges of modernization and internationalization of Chinese herbal medicine, it is urgent to establish thorough and effective procedures for quality identification of Chinese herbal medicine, and there is an urgent need for new analytical and testing techniques that are efficient, accurate, and environmentally friendly. 
Multiple light scattering is a cutting-edge and analytical method that can accurately and rapidly assess the quality of Chinese herbal medicine without altering the nature or state of the sample or using organic reagents. Indigo Naturalis is considered a good remedy for pediatric hyperthermia, psoriasis, leukemia, and ulcerative colitis. In this study, the process of addition of Indigo Naturalis powder in water was recorded precisely using a multiple light scattering instrument. 
The qualitative and quantitative measurements of the instrument can be used to accurately capture the overall trajectory and sinking behavior of Indigo Naturalis powder into water and to establish a rapid evaluation method for the quality of Indigo Naturalis with the transmission and backscattering spectrograms of the sample as qualitative indicators and stability index as a quantitative indicator. The analytical technique based on multiple light scattering provides a fast, accurate, green, and environmentally friendly method for the quality evaluation of Indigo Naturalis and supports the development and transformation of high-quality Indigo Naturalis.

Figure 2A-D correspond to S1, S2, S3, and S4 of Indigo Naturalis, respectively. A is of high-quality Indigo Naturalis, showing the same light transmittance at any height within 0-10 min, which is very stable. B is the common Indigo Naturalis, and its light transmittance fluctuates slightly with the change of time and is generally stable. C and D are fake and inferior products. There may be two conditions in the transmission spectrograms of fake Indigo Natura...

Here, we present a quality evaluation method based on a multiple light scattering technique to evaluate the quality of Indigo Naturalis.

Quality control of Chinese herbal medicine is a crucial component of Chinese herbal medicine research and development. Faced with the challenges of ...

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Learn rapid quality assessment of Indigo Naturalis using multiple light scattering for traditional Chinese medicine. This non-invasive method ensures authenticity without altering samples or using harmful reagents.

Fast Inspection of Quality of Indigo Naturalis by Multiple Light Scattering

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