Glycyrrhiza glabra (root)

Nomenclature

Glycyrrhiza glabra L.   Fabaceae  
Standardized common name (English): licorice  
Ayurvedic name(s): yashtimadhu  
Pinyin name(s): guang guo gan cao; gan cao (root & rhizome)

Botanical Voucher Specimen

Source: MOBOT, Tropicos.org	Source: Trish Flaster, MSc, Botanical Liaisons, LLC

Organoleptic Characteristics

Color: Unpeeled – yellowish or purplish brown to dark brown externally and yellowish internally. Peeled – pale yellow. Source: Natural Remedies Pvt Ltd Aroma/Odor: Characteristic Flavor/Taste: Sweet, sugary Source: American Herbal Products Association. March 2013. Organoleptic Analysis of Herbal Ingredients. AHPA: Silver Spring, MD

Macroscopic Characteristics

"...Root nearly cylindrical up to 2 cm in diameter, externally wrinkled with patches of cork. Fracture, coarsely fibrous in bark and splintery in wood." Source: Natural Remedies Pvt Ltd Source: Flora von Deutschland, Österreich und der Schweiz- Otto Wilhelm Thomé (1885) Foliage Source: Encyclopedia of Life http://eol.org/data_objects/2447928 Inflorescence Source: Encyclopedia of Life http://eol.org/data_objects/24932881 Source: PlantaPhile Source: PlantaPhile Dried Roots Source: Natural Remedies Pvt Ltd http://www.naturalremedy.com/ Dried Roots 2 Source: Encyclopedia of Life http://eol.org/data_objects/19163752

Microscopic Characteristics

"Transverse section of stolon more or less rounded. Phellem severeal layered with tabular cells; outer layers filled with reddish brown contents, inner colourless. Phellogen indistinct; phelloderm three to five layered, collenchymatous; some of the cells contain calcium oxalate and minute starch grains. Secondary phloem with numerous concentrically arranged bundles of phloem fibres and surrounded by a parenchymatous sheath containing prisms of calcium oxalate. Medullary rays distinct, bi-to multiseriate, parenchymatous, in continuation with those of xylem. The rays are narrower in xylem and wider in phloem region. Xylem consists of vessels, fibres and lignified wood parenchyma. The unpeeled drug shows the presence of polyhedral tubular brownish cork cells. In case of stolons, the pith is present and is parenchymatous. The root is characterized by the presence of tetrarch xylem and absence of pith. Powder: It shows plenty of starch grains, hexagonal crystals vessel elements are with reticulate wall pitting." Source: Natural Remedies Pvt Ltd "The powder is identified by (1.) The character and location of starch-grains and crystals; (2) the very numerous bast-fibers of peculiar appearance and the almost identical wood-fibers; (3) The peculiar sieve-tissue. The starch-grains are irregularly spheroidal, mostly solitary, and range from 1.5 ore 2 to 20 microns in diameter. They are contained in medullary-ray and parenchyma-cells, and are often associated in the same cell with the monoclinic prismatic crystals of calcium oxalate, sometimes also with oil-gobules. The bast- and wood-fibers are yellow, thick-walled, and doubly pointed. Part of the sieve-tubes have their cavities nearly or quite obliterated by cell-wall thickening." Source: Hare, Caspari, Rusby. National Standard Dispensatory (1905) Transverse section Source: Natural Remedies Pvt Ltd http://www.naturalremedy.com/ Starch granules and hexagonal crystals in powder Source: Natural Remedies Pvt Ltd http://www.naturalremedy.com/ Vessels with reticulate thickening in powder Source: Natural Remedies Pvt Ltd http://www.naturalremedy.com/

High Performance Liquid Chromatographic Identification

G. glabra root Licorice (root) (Glycyrrhiza glabra) Sample Preparations: Extract 1.0 g of coarsely powdered Glycyrrhiza root in 50 mL of water by boiling for about 5 minutes, and filter. Repeat for 4-5 times or until the extract is colorless. Combine the extracts, concentrate to about 100 mL, and cool to room temperature. Before injection, filter through a membrane filter of 0.45-um or finer pore size, discarding the first 5 mL of the filtrate. Column: C18, 25-cm x 4.6 mm, 5-um Mobile Phase: 0.14 g of anhydrous potassium dihydrogen phosphate in 900 mL of water, add 0.5 mL phosphoric acid, mix, complete to volume with water, and mix (Solution A); and acetonitrile (Solution B) Elution: Gradient program, see Table below Flow rate: 1.5 mL/min Detection: UV, 254 nm Injection volume: 20 uL G. glabra root G. inflata root G. uralensis root Source: Natural Remedies Pvt Ltd Table: Gradient program Time (min) Solution A (%) Solution B (%) 0-18 95-55 5-45 18-25 55-20 45-80 25-28 20 80 28-35 20-55 80-45 35-40 55-95 45-5 40-45 95 5

High Performance Thin Layer Chromatographic Identification

Glycyrrhiza glabra HPTLC ID - Developed, UV 366 nm Licorice (root) (Glycyrrhiza glabra) Lane Assignments Lanes, from left to right (Track, Volume, Sample): 2 μL Ammonium glyccyrhizate 2 μL Glycyrrhizic acid 2 μL Licorice root 1 (Glycyrrhiza glabra) 2 μL Licorice root 2 (Glycyrrhiza glabra) 2 μL Licorice root 3 (Glycyrrhiza glabra) 2 μL Licorice root (Glycyrrhiza glabra)  Other Notes Reference Standard Solution: 0.1 mg/mL ammonium glycyrrhizate in ethanol and water (7:3). Or 0.1 mg/mL glycyrrhizic acid in ethanol and water (7:3). Reference Sample Preparations: Mix 0.5 g of powdered sample with 10 mL of ethanol and water (7:3, v/v), sonicate for 10 minutes, centrifuge or filter the solution, and use the supernatant / filtrate. Stationary Phase: HPTLC, Silica gel 60 F254 Mobile Phase: Ethyl acetate, acetic acid, formic acid, water (15:1:1:2) Development: Saturate chamber for 20 minutes; developing distance 70 mm from lower edge of the plate; relative humidity 33%, temperature 25°. Derivatization reagent: Methanol sulfuric acid reagent- 180 mL of ice-cooled methanol are mixed with 20 mL of sulfuric acid. Detection: a. Examine under UV light at 254 nm b. Dip (time 0, speed 5) in Derivatization reagent, heat at 100°C for 10 min, leave to cool, and examine under visible light. Procedure: Reference Standard Solutions, Stationary Phase, Mobile Phase, Development, Derivatization reagent, and Detection, as described above. Test Sample Preparation: Prepare test sample as described under Reference Sample Preparations and apply 2 uL. Identification: Compare Test Sample Preparation chromatogram with chromatograms of Reference Sample Preparations. The Test Sample Preparation chromatogram is similar to that of the Reference Sample Preparations chromatograms. Additional weak zones may be present. Under UV light, the Test Sample Preparation chromatogram exhibits a quenching zone in the lower-third section of the chromatogram corresponding to the zone due to ammonium glycyrrhizate in the Reference Standard Solution chromatogram. Above it there are four quenching zones corresponding to those marked with red arrows in the Reference Sample Preparations chromatograms. After derivatization and under visible light, the Test Sample Preparation chromatogram exhibits a brown zone in the lower-third section of the chromatogram corresponding to the zone due to ammonium glycyrrhizate in the Reference Standard Solution chromatogram. Above it there are four yellow zones corresponding to those marked with black arrows in the Reference Sample Preparations chromatograms. Note: Images presented in this entry are examples and are not intended to be used as a bases for setting specifications for quality control purposes. Glycyrrhiza glabra HPTLC ID - Developed, UV 366 nm Glycyrrhiza glabra HPTLC ID - Developed, White RT Source: HPTLC Association

Supplementary Information

Other Publications

HPLC with Online Near-IR analysis, Li, et al., 2015

Online near-infrared analysis coupled with MWPLS and SiPLS models for the multi-ingredient and multi-phase extraction of licorice (Gancao),

Abstract. This study aims to analyze the active pharmaceutical ingredients (APIs) of licorice (Radix Glycyrrhizae; gancao), including glycyrrhizic acid, liquiritin, isoliquiritin and total flavonoids, in multi-ingredient and multi-phase extraction by online near-infrared technology with fiber optic probes and chemometric analysis. High-performance liquid chromatography and ultraviolet spectrophotometry determined the APIs content in different extraction phases by online near-infrared analysis, which included sample set selection by the Kennard–Stone algorithm, optimization of spectral pretreatment methods (i.e., orthogonal signal correction and wavelet denoising spectral correction), and model calibration by the partial least-squares algorithm, moving-window partial least-squares algorithm and synergy interval partial least-squares (SiPLS) algorithm. The relative errors and F values were used to assess the models in different extraction phases. The root-mean-square error of correction, root-mean-square error of cross-validation and root-mean-square error of prediction of APIs in the SiPLS model was less than 0.07. The F values of glycyrrhizic acid, liquiritin, isoliquiritin and total flavonoids were 10,765, 32,431, 649 and 6080, respectively, which were larger than 6.90 (P < 0.01). The study demonstrated the feasibility of online NIR analysis in the multi-ingredient and multi-phase extraction of APIs from licorice.