Chemical Education Today
Reports from Other Journals
Research Advances by Angela G. King
Spinning Natural Proteins into Fibers In the future, you may snuggle up in warm, cozy sweats made of chicken feathers or jeans made of wheat, enjoying comfortable, durable new fabrics that are “green” and environmentally friendly. Researchers in Australia are reporting that new advances are paving the way for such exotic new materials—made from agricultural waste or byproducts—to hit store shelves as environmentally friendly alternatives to the estimated 38 million tons of synthetic fabrics produced worldwide each year. They review research on the development of these next generation eco-friendly fibers, which will produce fabrics with a conventional feel. In their recent review, Andrew Poole, Jeffrey Church, and Mickey Huson note that scientists first produced commercial fabrics made of nontraditional materials—including milk, peanut, and corn proteins—almost 50 years ago. Although these so-called “regenerated” fabrics had the look and feel of conventional, protein-based fabrics such as wool and silk, they tended to perform poorly when wet. This problem, combined with the advent of petroleum-based synthetic fibers, caused the production of these unusual fabrics to stop, the researchers say. Amid concerns about the environment and consumer demand for eco-friendly products, renewable fabrics made from nontraditional agricultural materials are now poised to make a comeback, the scientists report. Promising fabric sources include agricultural proteins, such as keratin from scrap chicken feathers, and gluten from wheat (Figure 1). The scientists describe advances in nanotechnology and chemical crosslinking that can improve the strength and biodegradability of these fabrics, paving the way for commercial production of eco-friendly clothing, furniture upholstery, and other products.
But advances in science make new types of fibers appropriate for biomedical applications as well. Scientists in Israel are reporting the first successful spinning of a key natural protein into strong nano-sized fibers about 1/50,000th the width of a human hair. The advance could lead to a new generation of stronger, longer-lasting biocompatible sutures and bandages to treat wounds. Eyal Zussman and colleagues point out that researchers have tried for years to develop wound-repair materials from natural proteins, hoping that such fibers would be more compatible with body tissue than existing materials. Scientists recently focused on producing these fibers through “electrospinning”, a high-tech weaving process that uses electrical charges to draw out nano-sized fibers from a liquid. But the approach has achieved poor results until now. In the new study, the scientists describe a novel method for producing electrospun polymers using bovine serum albumin
Figure 1. Renewable fabrics made from fibers of nontraditional agricultural materials are now poised to make a comeback due to concerns about the environment and consumer demand for ecofriendly products. For instance, feather keratins (left) can be digested
or solubilized (middle) and used as biopolymer films, coatings, compostable packaging, and composite materials, or wet-spun into regenerated keratin fibers (right). Images courtesy of CSIRO. Reprinted with permission from Biomacromolecules 2009, 10, 1–8.
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Figure 2. Scanning electron microscope (SEM) images of a crosssection of electrospun BAS fibers (left) and optical image of electrospun ribbon made of as-spun BAS nanofibers (right). Image courtesy American Chemical Society.
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Chemical Education Today
(BSA), a globular protein found in cow’s blood. BSA is similar to serum albumin, one of the most abundant proteins in the human body. The method involves removing the many natural disulfide bonds in BSA by adding β-mercaptoethanol. That reduction results in a thinner, more spinnable protein solution. Using electrospinning, the protein solution then results in strong fibers that are easily spun into suture-like threads or thick-shapeable mats resembling conventional wound dressings (Figure 2). The new, extended structures produce keratin or elastin-like fibers and are stabilized by the spontaneous reforming of strong inter- and intramolecular disulfide bonds. When the research team added a sulfhydryl blocking agent (IAA) to the solution after the original disulfides had been reduced, the result was fibers that were more ductile, but inferior in terms of stress–strain characteristics and stiffness. Scientists also studied the effects of pH on fiber properties and found that basic solutions favor reduction of the original disulfide bonds, which opens more sulfhydryl sites for the formation of new bridges and enhances mechanical properties of the resulting fibers. On the other hand, acidic conditions cause BSA to have an overall positive charge and adopt an expanded conformation called the E form, which reduces its tendency to aggregate. Solutions near the isoelectric point of the protein (pH = 5.4) are unfavorable because BSA molecules tend to clump together, interfering with electrospinning and restricting orientation. The importance of the disulfide bridges was established by analysis with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS–PAGE) and mass spectrometry. X-ray crystallography analysis indicated that the nanofibers and cast films are semicrystalline in nature. This approach is being followed by the groups of Zvi Nevo and Abraham Katzir at Tel Aviv University, the researchers said, noting that the new method also can be applied to other types of natural proteins. More Information 1. Poole, Andrew J.; Church, Jeffrey S.; Huson, Mickey G. Environmentally Sustainable Fibers from Regenerated Protein. Biomacromolecules 2009, 10, 1–8. 2. Dror, Yael; Ziv, Tamar; Makarov, Vadim; Wolf, Hila; Admon, Arie; Zussman, Eyal. Nanofibers Made of Globular Proteins. Biomacromolecules 2008, 9, 2749–2754. 3. http://www.csiro.au/science/From-Feathers-To-Fibres.html (accessed May 2009) will give the reader more information on fibers made from nontraditional agricultural sources. 4. This Journal has previously published upper-level lab experiments involving BSA. See J. Chem. Educ. 2006, 83, 294–295; 2004, 81, 395–397; 2002, 79, 115–116; and 1991, 68, 262–264. 5. More information on Poole and Zussman’s work, including videos of electrospinning, is available online. See http://www.csiro. au/people/Andrew.Poole.html and http://hitech.technion.ac.il/~eyal/, respectively (both sites accessed May 2009). 6. Research Advances has previously featured additional work on new fiber-related research. See J. Chem. Educ. 2007, 84, 746–748.
New Way To Remove Unwanted Heparin from Blood Scientists in Poland report the development of a potential new way to quickly remove the anticoagulant heparin from patients’ blood in order to avoid unwanted side effects that the current use of that blood thinner can cause. Heparin is a mixture of highly sulfated glycosaminoglycan (GAG) polysaccharides produced and stored in animal tissue mast cells. It has the highest negative charge density of any known biological molecule (~2.7 negative charges per disaccharide unit). It is widely prescribed as an anticoagulant because of its ability to enhance antithrombin’s inactivation of thrombin and factor Xa, two key coagulation enzymes. In a new study, Krzysztof Szczubiałka and colleagues point out that doctors often want to remove heparin from the blood of patients undergoing surgery or other procedures immediately after completing the procedure. Leaving the heparin alone could lead to unwanted bleeding. Doctors now eliminate heparin by giving patients protamine sulfate, a protein drug that stops heparin’s anticoagulant effects. However, they are seeking a better drug because protamine carries a risk of serious side effects. The scientists describe development of a potential new approach to heparin removal that involves use of microscopic polymeric beads made from modified chitosan, a material obtained from shellfish. The interaction between heparin and chitosan has been previously well documented, and the scientists used this background to develop a system where the sequestering microspheres were constructed from materials that are both inexpensive and nontoxic, opening the door for widespread medical applications. The hydrogel microspheres (ChGp, Figure 3) were produced by crosslinking chitosan with genipin. They swell in solutions with pH below 6.5 and shrink at more basic pH levels. The microspheres were spectrophotometrically shown to bind heparin in a pH-dependent manner that was also dependent on weight of the ChGp microspheres. Binding was
Figure 3. Optical microscope image of the ChGp microspheres. Reprinted with permission from Biomacromolecules 2008, 9, 3127– 3132. Copyright 2008 American Chemical Society.
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Figure 4. Schematic for removing heparin from blood with chitosan micro spheres. Reprinted with permission from Biomacromolecules 2008, 9, 3127– 3132. Copyright 2008 American Chemical Society.
Figure 5. Time dependence of relative heparin concentration (c0 = 200 μg/mL, V = 5 mL) after the addition of 40 mg (•) ChGp and (■) ChGpGl microspheres at pH 7.4. Reprinted with permission from Biomacromolecules 2008, 9, 3127–3132. Copyright 2008 American Chemical Society.
favored at lower pH ranges, but the researchers found that at pH 7.4, typical of blood, heparin binding could be facilitated by the use of glycidyltrimethylammonium chloride (GTMAC) for quaternization of the microspheres (ChGpGl), thereby positively charging the surface of microspheres (Figure 4). The charge is maintained even at basic pH. In laboratory tests, the beads reduced concentrations of heparin to nearly zero within 10 minutes (Figure 5). More Information 1. Kaminski, Kamil; Zazakowny, Karolina; Szczubiałka, Krzysztof; Nowakowska, Maria. pH-Sensitive Genipin-Cross-Linked Chito-
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san Microspheres for Heparin Removal. Biomacromolecules 2008, 9, 3127–3132. 2. This Journal has previously published an experiment on measuring heparin concentrations. See J. Chem. Educ. 1990, 67, 446–448. 3. J. Chem. Educ. 1990, 67, 938–942 will give readers access to more chemistry of chitosan and the related molecule, chitin. 4. http://www.fda.gov/cder/drug/infopage/heparin/ is the FDA site for more information on heparin, while http://www. absoluteastronomy.com/topics/Heparin has basic facts and definitions related to heparin and its use (both sites accessed May 2009).
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Reports from Other Journals Two Food Additives Have Previously Unrecognized, Estrogen-Like Effects Scientists in Italy report the development and successful use of a fast new method to identify food additives that act as so-called “xenoestrogens”—substances with estrogen-like effects that are stirring international health concerns (literally, “foreign estrogens”). They used the method in a large-scale screening of additives that discovered two additives with previously unrecognized xenoestrogen effects. In the study, Pietro Cozzini and colleagues cite increasing concern about identifying these substances and about the possible health effects. Synthetic chemicals that mimic natural estrogens have been linked to a range of human health effects, from reduced sperm counts in men, to an increased risk of breast cancer in women. The xenoestrogens act as estrogen mimics, binding to one of two subtypes (α and β) of estrogen receptors (ERs), ligand-activated transcription factors, which trigger a biochemical cascade resulting in the transcription of certain genes. Part of the challenge in using computational screening to identify xenoestrogens is the flexibility of the ERα ligand-binding domain (LBD). Two conformations of the LBD, closed and open, are known and the protein can adopt either, depending on whether the LBD is occupied by an agonist, which stabilizes the closed conformation and seals the binding site, or an antagonist, which promotes the open conformation. The scientists examined a food additive database and employed a combination of molecular modeling and in vitro studies, and ultimately verified that the new method could identify xenoestrogens. They began by using the molecular modeling program Sybyl and using a computational screening approach to identify 31 potential ERα ligands through a docking–scoring procedure. Applying a stringent filter, based on both the calculated ligand volume buried within the LBD and the visual inspection of the chemical plausibility of the proposed ligand conformation, reduced the number of possible xenoestrogens from 31 down to 13. Of the 13 candidates identified from the food additive database, four had previously been identified as having estrogenic activity; the remaining nine were assayed in vitro to determine binding affinity to ERα and any biological effects. Through that work, the research team identified two previously unrecognized xenoestrogens. One was propyl gallate, a preservative used to prevent fats and oils from spoiling. The other was 4-hexylresorcinol, used to prevent discoloration in shrimp and other shellfish (Figure 6). Both were active at nM concentrations. “Some caution should be issued for the use
Figure 6. Scientists have identified two food additives with previously unrecognized, estrogen-like effects. One of the additives, 4-hexylresorcinol, is used to prevent discoloration in shrimp and other shellfish. Image courtesy National Cancer Institute, Renée Comet.
of propyl gallate and 4-hexylresorcinol as food additives”, the researchers recommend in the study. More Information 1. Amadasi, Alessio; Mozzarelli, Andrea; Meda, Clara; Maggi, Adriana; Cozzini, Pietro. Identification of Xenoestrogens in Food Additives by an Integrated in Silico and in Vitro Approach. Chem. Res. Toxicol. 2009, 22, 52–63. 2. This Journal has previously published work on food additives, including J. Chem. Educ. 1984, 61, 332–334. JCE has also published a problem-based learning experiment on measuring xenoestrogens in drinking water. See O’Hara, P. B. Pesticides in Drinking Water: ProjectBased Learning within the Introductory Chemistry Curriculum. J. Chem. Educ. 1999, 76, 1673–1677. 3. Additional information on exposure to xenoestrogens can be found in many scholarly journals, including Olea-Serrano, F. Exposure to Xenoestrogens in Children. Toxicol. Indust. Health 1999, 15, 152.
Supporting JCE Online Material http://www.jce.divched.org/Journal/Issues/2009/Sep/abs1006.html Abstract and keywords Full text (PDF) with links to cited URLs and JCE articles
Angela G. King is Senior Lecturer in Chemistry at Wake Forest University, P.O. Box 7486, Winston-Salem, NC 27109;
[email protected].
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