Unsaponifiable Matter in Carnuba (Cera carnuba) Wax, a Modification of the USP/NF and FCC Methods

Carnuba wax consists chiefly of myricyl cerotate (MW 817.4) and small quantities of free cerotic acid (C26H52O2, Mw 396.7) and myricyl alcohol (C30H62O, mp 90°C). Of the two common extraction solvents, ethyl ether or petroleum ether, Lewkowitsch prefers the former. Concerning separation of phases, he advocates addition of small amounts of alcohol or caustic, and he also states that formation of a flocculant layer between the aqueous layer and the solvent does not interfere with the correct estimation of the unsaponifiable matter. These statements were not corroborated in the hands of this chemist. The “Unsaponifiable Matter” in oils or and fats, which consist mainly of hydrocarbons, sterols and aliphatic alcohols of high molecular mass that are not saponifiable by alkali hydroxides but are soluble in the ordinary fat solvents, and to products of saponification that are soluble in such solvents. Carnuba wax, a rather expensive wax, may be adulterated with less expensive paraffin by dishonest merchants. ASTM has a method for determining paraffinic material in carnuba wax. It uses heptanes at its boiling point to dissolve the wax, apply it to a silica gel column, and elute only the nonpolar (i.e. alkane) material. The method has the disadvantage of using a large volume of haptane, nor is it called for by either US Pharmacopeia/National Formulary (USP/NF) or Food Chemicals Codex (FCC). The test for unsaponifiable matter on pure carnuba wax will yield a result of 50.0% - 55.0%, while a higher result will betray the presence of paraffin adulterants.

Chondrocyte Production of Pro-Inflammatory Chemokine MCP-1 (CCL-2) and Prostaglandin E-2 Is Inhibited by Avocado/Soybean Unsaponifiables, Glucosamine, Chondroitin Sulfate Combination

Osteoarthritis (OA) is a chronic, painful disease affecting articulating joints in man and animals. It is characterized by cartilage breakdown, bone remodeling, osteophyte formation and joint inflammation. Currently used non-steroidal anti-inflammatory drugs for the management of OA are known to have deleterious side effects. To address the need for alternatives, we evaluated the anti-inflammatory effects of a combination of avocado/soybean unsaponifiables (ASU), glucosamine (GLU) and chondroitin sulfate (CS) by measuring chemokine MCP-1 (monocyte chemoattractant protein 1, CCL2) and prostaglandin E-2 (PGE2) in stimulated chondrocytes. As the only cellular constituents of cartilage, chondrocytes are the source of pro-inflammatory mediators that play critical roles in the pathogenesis of OA. Chondrocytes were incubated: with: 1) control media, 2) [ASU + GLU + CS] combination, or 3) Phenylbutazone (PBZ) for 24 hours. Cells were next stimulated with IL-1β or LPS for another 24 hrs. MCP-1 and PGE2 from supernatants were quantitated by immunoassay. Another set of chondrocytes seeded in chamber slides were stimulated with IL-1β for 1 hour and then immunostained for NF-κB. Chondrocytes stimulated with IL-1β or LPS significantly increased MCP-1 and PGE2 production which were significantly reduced after treatment with [ASU + GLU + CS]. In contrast, PBZ significantly reduced PGE2 but not MCP-1 production. IL-1β stimulation induced nuclear translocation of NF-κB, which was inhibited by pre-treatment with either [ASU + GLU + CS] or PBZ. The present study provides evidence that the production of MCP-1 by chondrocytes can be inhibited by the combination of [ASU + GLU + CS] but not by PBZ. In contrast, PGE2 production was inhibited by either treatment suggesting that the production of MCP-1 and PGE2 could be independently regulated. The finding of distinct effects of [ASU + GLU + CS] on MCP-1 and PGE2 synthesis supports a scientific rationale for a multimodal treatment approach in the management of OA.

Primary Separation of Antioxidants (Unsaponifiables) the Wet Biomass Microalgae <i>Chlamydomonas</i>sp. and Production of the Biodiesel

This work studies the saponification which directs the wet biomass of algae Chlamydomonas sp. like a previous stage to production of biodiesel. This stage allows the obtainment of fatty acids to produce biodiesel, instead of the gross lipid fraction. In addition of the fatty acids, utilizing the same process one can also obtain the fraction unsaponifiable, these are soluble in apolar solvents and contain mainly carotenoids that can take action as antioxidants and photoprotectors, as they reduce the oxidation of unsaturated fatty acids. The saponification direct and extraction of fatty acids from biomass is faster and reduces the time and cost of operation. The separation of unsaponifiable matter from the biomass humid of microalgae Chlamydomonas sp., was held according to the method AOCS (Ca 6a-40), using extraction Liquid-liquid with hexane as solvent. Subsequently, phase hydroalcoholic or from soap, containing fatty acids, was acidified by addition of H2SO4 and the fatty acids were recovered by the addition of hexane. After acidulation of the soap, necessary for obtaining of the fatty acids was performed the stage of esterification for obtaining of biodiesel. The operating conditions were: molar ratio fatty acid:methanol (1:10), as catalyst 8% H2SO4 calculated in relation to the mass of fatty acid, 200℃ and reaction time of 90 minutes. The content of methyl esters was 96.8% determined by gas chromatography according to standard EN14103. The quality of biodiesel produced from wet biomass of Chlamydomonas sp. is according to the specification established by standard EN 14214 and RANP No. 14. For the identification of the composition the unsaponifiable fraction was used the method of High Performance Liquid Chromatography (HPLC). The composition of the material unsaponifiable found was of: Carotenoids total (0.76%);Lutein (0.45%);Zeaxanthin (0.07%);α-carotene (0.05%);β-carotene (0.11%);13 cisβ-carotene (0.05%) and 9-cisβ-carotene (0.03%).

Comparative Studies on Egyptian and Libyan Roselle Seeds as a Source of Lipid and Protein

Proceeding from the fact that the seeds of Roselle plant are full of nutritional constituents, however in Egypt and Libya, they are often discarded as a by-product, this study aims to evaluate the nutritional composition of Roselle seeds grown in Egypt and Libya as a source of oil and protein besides making a comparison between whole chemical composition of Roselle seeds grown in both countries. Ground of whole Egyptian and Libyan Roselle seeds powder contained high amount of protein (31.02% ± 0.93% and 28.67% ± 0.45%), crude fat (21.6% ± 0.66% and 16.94% ± 0.86%) and total ash (6.89% ± 0.11% and 5.60% ± 0.10%), respectively. However, Egyptian seeds have moisture content, protein, crude fat and total ash higher than Libyan seeds. Crude oil from Egyptian seeds had high refractive index and iodine value in comparison with crude oil from Libyan seeds. There were no remarkable differences between both seeds in acidity percent, unsaponifiable matters percent and saponification value. Linoleic, oleic and palmitic acids were the major fatty acid constituents in Egyptian Roselle seeds. Meanwhile linolenic, linoleic, oleic, stearic, palmitoleic and palmitic acids were the major fatty acid constituents in Libyan Roselle seeds. Crude oil from Egyptian seeds had higher percent of unsaturated fatty acids than crude oil from Libyan seeds. Unsaponifiable matters constituents for extracted oil from Egyptian seeds were free from n-pentacosane (C25) and rich in n-hexacosane (C26). Oil from both seeds had the same content of Beta sito-sterol and stigma-sterol. Both seeds were rich in glutamic acid, aspartic acid, arginine and leucine. Libyan seeds were rich in essential amino acids in comparison with Egyptian seeds. Finally nutritional comparison of Roselle seeds variation depends on the variety, location and environmental conditions during cultivation. Roselle seeds are a good source for extraction of oil and protein. Protein from Roselle seeds could be used as a supplement material for poor food in lysine.