4f1320ac8c246a5fbb04f3380dd8e9bf pmc_33050372.pdf 7060deb46fa9bdf85c584658a28c89e01ef646c1 pmc_33050372.pdf 743eec0a7687571091723e61a92db1ccb6e8bd7faeeec905df39462de0e5d01e pmc_33050372.pdf Title: Carprofen Permeation Test through Porcine Ex Vivo Mucous Membranes and Ophthalmic Tissues for Tolerability Assessments: Validation and Histological Study Subject: Carprofen (CP), a non-steroidal anti-inflammatory drug (NSAID), is profusely used in veterinary medicine for its analgesic and anti-inflammatory activity. Some undesirable effects are associated with its systemic administration. Alternative local routes are especially useful to facilitate its administration in animals. The main aim of this paper is to validate the suitability of ex vivo permeation experiments of CP with porcine mucous membranes (buccal, sublingual and vaginal) and ophthalmic tissues (cornea, sclera and conjunctiva) intended to be representative of naïve in vivo conditions. Chromatographic analysis of CP in membrane-permeated samples and drug-retained have been validated following standard bioanalytical guidelines. Then, recovery levels of drugs in tissue samples were assessed with aqueous phosphate buffered saline (PBS) buffer to preserve the histological integrity. Finally, as a proof of concept, a series of CP permeation tests in vertical Franz diffusion cells has been performed to evaluate permeation flux and permeability constants in all tissues, followed by a histological study for critical evaluation. Furthermore, synthetic tissue retention-like samples were prepared to verify the value of this experimental study. Results show linear relationships with good determination coefficient (R2 > 0.998 and R2 > 0.999) in the range of 0.78 to 6.25 mg/mL and 3.125 mg/mL to 100 mg/mL, respectively. Low limits of quantification around 0.40 g/mL were allowed to follow permeation levels until a minimum of 0.40% of the locally-applied dose. This method showed a good accuracy and precision with values lower than 2%. After the recovery technique, reproducible values below 30% were achieved in all tissues, suggesting it is a non-damaging method with low efficiency that requires the use of further solvents to enhance the extraction percentages. After permeation and histology tests, no relevant peak interferences were detected, and no cell or tissue damage was found in any tissue. In conclusion, results demonstrate the suitability of this test to quantify the distribution of CP with good histological tolerability. Keywords: carprofen; HPLC; permeation test; histological integrity; mucous tolerability; ophthalmic tolerability Author: Lidia Gómez-Segura, Alexander Parra, Ana C. Calpena, Álvaro Gimeno and Antonio Boix-Montañes Creator: LaTeX with hyperref package Producer: pdfTeX-1.40.18 CreationDate: Sat Oct 10 09:25:23 2020 CEST ModDate: Sat Oct 10 09:25:23 2020 CEST Custom Metadata: yes Metadata Stream: no Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 17 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 4563810 bytes Optimized: no PDF version: 1.5 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- ZZDTXJ+URWPalladioL-Ital Type 1 Custom yes yes no 159 0 QVKUYQ+URWPalladioL-Bold Type 1 Custom yes yes no 160 0 HCFYFJ+VnURWPalladioL-Bold Type 1 Custom yes yes no 161 0 TLDMFY+URWPalladioL-Roma Type 1 Custom yes yes no 162 0 OFLXAT+Rpxr Type 1 Builtin yes yes no 163 0 OVNEXW+EURM10 Type 1 Builtin yes yes no 164 0 CTCXDQ+TimesNewRomanPS-BoldItalicMT Type 1C WinAnsi yes yes no 169 0 MCBCCI+SourceSansRoman-Regular Type 1C Custom yes yes no 186 0 MCBCCM+SourceSansRoman.613wght Type 1C Custom yes yes no 187 0 MCBCCO+SourceSansRoman-Bold Type 1C Custom yes yes no 188 0 SPXLNH+PazoMath Type 1 Builtin yes yes no 234 0 RYNHBZ+Pxsy Type 1 Builtin yes yes no 265 0 ABCDEE+Palatino Linotype TrueType WinAnsi yes yes no 272 0 ABCDEE+Palatino Linotype,Italic TrueType WinAnsi yes yes no 273 0 ABCDEE+Palatino Linotype,Bold TrueType WinAnsi yes yes no 274 0 Times New Roman TrueType WinAnsi no no no 275 0 ABCDEE+Palatino Linotype CID TrueType Identity-H yes yes yes 276 0 ABCDEE+Palatino Linotype TrueType WinAnsi yes yes no 298 0 ABCDEE+Palatino Linotype,Italic TrueType WinAnsi yes yes no 299 0 ABCDEE+Palatino Linotype,Bold TrueType WinAnsi yes yes no 300 0 Times New Roman TrueType WinAnsi no no no 301 0 GBLQZB+Rpxi Type 1 Builtin yes yes no 321 0 TGMEOR+Rpcxr Type 1 Builtin yes yes no 322 0 YOPLSB+VnURWPalladioL Type 1 Custom yes yes no 341 0 WBVKDJ+URWPalladioL-BoldItal Type 1 Custom yes yes no 365 0 XIRVNA+EURB10 Type 1 Builtin yes yes no 366 0 MFIGJL+Rpxb Type 1 Builtin yes yes no 367 0 ABCDEE+Palatino Linotype TrueType WinAnsi yes yes no 386 0 ABCDEE+Palatino Linotype,Italic TrueType WinAnsi yes yes no 387 0 ABCDEE+Palatino Linotype,Bold TrueType WinAnsi yes yes no 388 0 Times New Roman TrueType WinAnsi no no no 389 0 ABCDEE+Palatino Linotype CID TrueType Identity-H yes yes yes 390 0 ABCDEE+Palatino Linotype,BoldItalic TrueType WinAnsi yes yes no 391 0 ABCDEE+Palatino Linotype TrueType WinAnsi yes yes no 415 0 ABCDEE+Palatino Linotype,Italic TrueType WinAnsi yes yes no 416 0 ABCDEE+Palatino Linotype,Bold TrueType WinAnsi yes yes no 417 0 Times New Roman TrueType WinAnsi no no no 418 0 ABCDEE+Palatino Linotype CID TrueType Identity-H yes yes yes 419 0 ABCDEE+Palatino Linotype,BoldItalic TrueType WinAnsi yes yes no 420 0 ABCDEE+Palatino Linotype TrueType WinAnsi yes yes no 456 0 ABCDEE+Palatino Linotype,Italic TrueType WinAnsi yes yes no 457 0 ABCDEE+Palatino Linotype,Bold TrueType WinAnsi yes yes no 458 0 Times New Roman TrueType WinAnsi no no no 459 0 ABCDEE+Palatino Linotype CID TrueType Identity-H yes yes yes 460 0 ABCDEE+Palatino Linotype,BoldItalic TrueType WinAnsi yes yes no 461 0 ABCDEE+Palatino Linotype TrueType WinAnsi yes yes no 500 0 ABCDEE+Palatino Linotype,Italic TrueType WinAnsi yes yes no 501 0 ABCDEE+Palatino Linotype,Bold TrueType WinAnsi yes yes no 502 0 Times New Roman TrueType WinAnsi no no no 503 0 ABCDEE+Palatino Linotype CID TrueType Identity-H yes yes yes 504 0 ABCDEE+Palatino Linotype TrueType WinAnsi yes yes no 543 0 ABCDEE+Palatino Linotype,Italic TrueType WinAnsi yes yes no 544 0 ABCDEE+Palatino Linotype,Bold TrueType WinAnsi yes yes no 545 0 Times New Roman TrueType WinAnsi no no no 546 0 ABCDEE+Palatino Linotype CID TrueType Identity-H yes yes yes 547 0 ABCDEE+Palatino Linotype TrueType WinAnsi yes yes no 573 0 ABCDEE+Palatino Linotype,Italic TrueType WinAnsi yes yes no 574 0 ABCDEE+Palatino Linotype,Bold TrueType WinAnsi yes yes no 575 0 Times New Roman TrueType WinAnsi no no no 576 0 ABCDEE+Palatino Linotype CID TrueType Identity-H yes yes yes 577 0 ABCDEE+Palatino Linotype TrueType WinAnsi yes yes no 603 0 ABCDEE+Palatino Linotype,Italic TrueType WinAnsi yes yes no 604 0 ABCDEE+Palatino Linotype,Bold TrueType WinAnsi yes yes no 605 0 Times New Roman TrueType WinAnsi no no no 606 0 ABCDEE+Palatino Linotype CID TrueType Identity-H yes yes yes 607 0 ABCDEE+Palatino Linotype TrueType WinAnsi yes yes no 634 0 ABCDEE+Palatino Linotype,Italic TrueType WinAnsi yes yes no 635 0 ABCDEE+Palatino Linotype,Bold TrueType WinAnsi yes yes no 636 0 Times New Roman TrueType WinAnsi no no no 637 0 ABCDEE+Palatino Linotype CID TrueType Identity-H yes yes yes 638 0 ABCDEE+Palatino Linotype TrueType WinAnsi yes yes no 679 0 ABCDEE+Palatino Linotype,Italic TrueType WinAnsi yes yes no 680 0 ABCDEE+Palatino Linotype,Bold TrueType WinAnsi yes yes no 681 0 Times New Roman TrueType WinAnsi no no no 682 0 ABCDEE+Palatino Linotype CID TrueType Identity-H yes yes yes 683 0 Jhove (Rel. 1.28.0, 2023-05-18) Date: 2024-10-02 04:18:18 CEST RepresentationInformation: pmc_33050372.pdf ReportingModule: PDF-hul, Rel. 1.12.4 (2023-03-16) LastModified: 2023-03-31 11:43:27 CEST Size: 4563810 Format: PDF Version: 1.5 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 0 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: ViewerPreferences: HideToolbar: false HideMenubar: false HideWindowUI: false FitWindow: true CenterWindow: false DisplayDocTitle: false NonFullScreenPageMode: UseNone Direction: L2R ViewArea: CropBox ViewClip: CropBox PrintArea: CropBox PageClip: CropBox PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: ぉのぴひはつふっぴどはの〠 Item: Title: きちぴづひどちぬび〠ちのつ〠きづぴとはつび〠 Children: Item: Title: ぃとづねどっちぬび〠ちのつ〠げづちでづのぴび〠 Item: Title: あどはぬはでどっちぬ〠きちぴづひどちぬび〠 Item: Title: あふててづひび〠ちのつ〠こはぬふぴどはのび〠 Item: Title: ぃちぬどぢひちぴどはの〠ぃふひぶづ〠こぴちのつちひつび〠〨ぃぃこ〩〬〠けふちぬどぴべ〠ぃはのぴひはぬ〠こぴちのつちひつび〠〨けぃこ〩〠ちのつ〠うはひぴどてどづつ〠こちねばぬづび〠〨うこ〩〠 Item: Title: ぃちひばひはてづの〠こはぬふぢどぬどぴべ〠 Item: Title: えぐがぃ〭さざ〠 Item: Title: ぁのちぬべぴどっちぬ〠きづぴとはつ〠ざちぬどつちぴどはの〠 Children: Item: Title: こばづっどてどっどぴべ〓こづぬづっぴどぶどぴべ〠 Item: Title: がどのづちひどぴべ〠ちのつ〠げちのでづ〠 Item: Title: いづぴづひねどのちぴどはの〠がどねどぴび〠 Item: Title: ぁっっふひちっべ〠ちのつ〠ぐひづっどびどはの〠 Item: Title: こぴちぢどぬどぴべ〠はて〠こぴちのつちひつび〠 Item: Title: ぁばばぬどっちぢどぬどぴべ〠はて〠ぴとづ〠きづぴとはつ〠 Children: Item: Title: ぐづひねづちぴどはの〠ごづびぴ〠ぷどぴと〠いどててふびどはの〠ぃづぬぬび〠 Item: Title: げづっはぶづひべ〠てひはね〠ぐはひっどのづ〠きふっはふび〠きづねぢひちのづび〠はひ〠くばとぴとちぬねどっ〠ごどびびふづび〠 Item: Title: いひふで〠げづぴづのぴどはの〠どのびどつづ〠ぴとづ〠ぐづひねづちぴどはの〠きづねぢひちのづ〠 Item: Title: ごひちのびねづねぢひちのづ〠うぬふへ〠ふのつづひ〠こどのに〠ぃはのつどぴどはのび〠 Item: Title: えどびぴはぬはでどっちぬ〠ぉのぴづでひどぴべ〠はて〠ぴとづ〠ぐづひねづちぴどはの〠きづねぢひちのづび〠 Item: Title: げづびふぬぴび〠 Children: Item: Title: ぁのちぬべぴどっちぬ〠きづぴとはつ〠ざちぬどつちぴどはの〠 Item: Title: きづぴとはつ〠ぁばばぬどっちぢどぬどぴべ〠 Children: Item: Title: ぐづひねづちぴどはの〠ぅへばづひどねづのぴ〠 Item: Title: いひふで〠げづっはぶづひべ〠ちのつ〠げづぴづのぴどはの〠どの〠きづねぢひちのづび〠 Item: Title: えどびぴはぬはでどっちぬ〠ぉのぴづでひどぴべ〠はて〠ぴとづ〠ぐづひねづちぴどはの〠きづねぢひちのづび〠 Item: Title: いどびっふびびどはの〠 Children: Item: Title: ぁのちぬべぴどっちぬ〠きづぴとはつ〠ざちぬどつちぴどはの〠 Item: Title: きづぴとはつ〠ぁばばぬどっちぢどぬどぴべ〠 Children: Item: Title: ぐづひねづちぴどはの〠ぅへばづひどねづのぴび〠 Item: Title: いひふで〠げづっはぶづひべ〠ちのつ〠いひふで〠げづぴづのぴどはの〠どの〠きづねぢひちのづび〠 Item: Title: えどびぴはぬはでどっちぬ〠ぉのぴづでひどぴべ〠はて〠ぴとづ〠ぐづひねづちぴどはの〠きづねぢひちのづび〠 Item: Title: ぃはのっぬふびどはのび〠 Item: Title: げづてづひづのっづび Filters: FilterPipeline: FlateDecode Fonts: Type1: Font: BaseFont: QVKUYQ+URWPalladioL-Bold FontSubset: true FirstChar: 2 LastChar: 241 FontDescriptor: FontName: QVKUYQ+URWPalladioL-Bold Flags: Symbolic FontBBox: -152, -301, 1000, 935 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: HCFYFJ+VnURWPalladioL-Bold FontSubset: true FirstChar: 129 LastChar: 226 FontDescriptor: FontName: HCFYFJ+VnURWPalladioL-Bold Flags: Symbolic FontBBox: -152, -301, 1000, 935 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: GBLQZB+Rpxi FontSubset: true FirstChar: 11 LastChar: 11 FontDescriptor: FontName: GBLQZB+Rpxi Flags: Symbolic FontBBox: -162, -280, 856, 733 FontFile: true Font: BaseFont: TLDMFY+URWPalladioL-Roma FontSubset: true FirstChar: 2 LastChar: 246 FontDescriptor: FontName: TLDMFY+URWPalladioL-Roma Flags: Symbolic FontBBox: -166, -283, 1021, 943 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: TGMEOR+Rpcxr FontSubset: true FirstChar: 169 LastChar: 174 FontDescriptor: FontName: TGMEOR+Rpcxr Flags: Symbolic FontBBox: -212, -286, 1344, 973 FontFile: true Font: BaseFont: OFLXAT+Rpxr FontSubset: true FirstChar: 11 LastChar: 62 FontDescriptor: FontName: OFLXAT+Rpxr Flags: Symbolic FontBBox: -40, -283, 904, 728 FontFile: true Font: BaseFont: OVNEXW+EURM10 FontSubset: true FirstChar: 22 LastChar: 22 FontDescriptor: FontName: OVNEXW+EURM10 Flags: Symbolic FontBBox: -32, -243, 1060, 720 FontFile: true Font: BaseFont: RYNHBZ+Pxsy FontSubset: true FirstChar: 0 LastChar: 21 FontDescriptor: FontName: RYNHBZ+Pxsy Flags: Symbolic FontBBox: -21, -994, 1292, 912 FontFile: true Font: BaseFont: SPXLNH+PazoMath FontSubset: true FirstChar: 87 LastChar: 87 FontDescriptor: FontName: SPXLNH+PazoMath Flags: Symbolic FontBBox: -40, -283, 878, 946 FontFile: true Font: BaseFont: WBVKDJ+URWPalladioL-BoldItal FontSubset: true FirstChar: 112 LastChar: 112 FontDescriptor: FontName: WBVKDJ+URWPalladioL-BoldItal Flags: Symbolic FontBBox: -170, -300, 1073, 935 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: XIRVNA+EURB10 FontSubset: true FirstChar: 22 LastChar: 22 FontDescriptor: FontName: XIRVNA+EURB10 Flags: Symbolic FontBBox: -30, -247, 1178, 723 FontFile: true Font: BaseFont: MFIGJL+Rpxb FontSubset: true FirstChar: 47 LastChar: 47 FontDescriptor: FontName: MFIGJL+Rpxb Flags: Symbolic FontBBox: -57, -266, 932, 729 FontFile: true Font: BaseFont: YOPLSB+VnURWPalladioL FontSubset: true FirstChar: 129 LastChar: 243 FontDescriptor: FontName: YOPLSB+VnURWPalladioL Flags: Symbolic FontBBox: -166, -283, 1021, 943 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: ZZDTXJ+URWPalladioL-Ital FontSubset: true FirstChar: 2 LastChar: 121 FontDescriptor: FontName: ZZDTXJ+URWPalladioL-Ital Flags: Symbolic FontBBox: -170, -305, 1010, 941 FontFile: true EncodingDictionary: Differences: true Pages: Page: Sequence: 1 Annotations: Annotation: Subtype: Link Rect: 75, 757, 183, 793 Annotation: Subtype: Link Rect: 474, 757, 519, 793 Annotation: Subtype: Link Rect: 176, 619, 187, 633 Annotation: Subtype: Link Rect: 359, 619, 370, 633 Annotation: Subtype: Link Rect: 224, 608, 235, 619 Annotation: Subtype: Link Rect: 466, 446, 518, 465 Annotation: Subtype: Link Rect: 161, 38, 240, 50 Annotation: Subtype: Link Rect: 410, 38, 519, 50 Page: Sequence: 2 Annotations: Annotation: Subtype: Link Rect: 115, 696, 122, 705 ActionDest: 779 Annotation: Subtype: Link Rect: 254, 655, 261, 665 ActionDest: 779 Annotation: Subtype: Link Rect: 500, 642, 507, 651 ActionDest: 779 Annotation: Subtype: Link Rect: 508, 642, 515, 651 ActionDest: 779 Annotation: Subtype: Link Rect: 149, 628, 156, 638 ActionDest: 779 Annotation: Subtype: Link Rect: 209, 628, 216, 638 ActionDest: 779 Annotation: Subtype: Link Rect: 216, 628, 223, 638 ActionDest: 779 Annotation: Subtype: Link Rect: 434, 628, 441, 638 ActionDest: 779 Annotation: Subtype: Link Rect: 441, 628, 448, 638 ActionDest: 810 Annotation: Subtype: Link Rect: 396, 615, 408, 624 ActionDest: 810 Annotation: Subtype: Link Rect: 408, 615, 420, 624 ActionDest: 810 Annotation: Subtype: Link Rect: 450, 615, 462, 624 ActionDest: 810 Annotation: Subtype: Link Rect: 465, 615, 477, 624 ActionDest: 810 Annotation: Subtype: Link Rect: 503, 615, 515, 624 ActionDest: 810 Annotation: Subtype: Link Rect: 146, 601, 158, 611 ActionDest: 810 Annotation: Subtype: Link Rect: 191, 601, 203, 611 ActionDest: 810 Annotation: Subtype: Link Rect: 326, 588, 338, 597 ActionDest: 810 Annotation: Subtype: Link Rect: 372, 574, 384, 584 ActionDest: 810 Annotation: Subtype: Link Rect: 451, 561, 463, 570 ActionDest: 810 Annotation: Subtype: Link Rect: 466, 561, 478, 570 ActionDest: 810 Annotation: Subtype: Link Rect: 273, 534, 285, 543 ActionDest: 810 Annotation: Subtype: Link Rect: 504, 534, 516, 543 ActionDest: 810 Annotation: Subtype: Link Rect: 117, 493, 129, 502 ActionDest: 810 Annotation: Subtype: Link Rect: 149, 399, 161, 408 ActionDest: 810 Annotation: Subtype: Link Rect: 164, 399, 176, 408 ActionDest: 810 Page: Sequence: 3 Annotations: Annotation: Subtype: Link Rect: 192, 699, 199, 711 ActionDest: 262 Annotation: Subtype: Link Rect: 337, 688, 349, 698 ActionDest: 842 Annotation: Subtype: Link Rect: 351, 688, 363, 698 ActionDest: 842 Annotation: Subtype: Link Rect: 421, 686, 428, 698 ActionDest: 262 Annotation: Subtype: Link Rect: 448, 686, 455, 698 ActionDest: 262 Page: Sequence: 4 Annotations: Annotation: Subtype: Link Rect: 242, 356, 254, 365 ActionDest: 810 Page: Sequence: 5 Annotations: Annotation: Subtype: Link Rect: 113, 659, 125, 668 ActionDest: 810 Annotation: Subtype: Link Rect: 369, 659, 381, 668 ActionDest: 810 Annotation: Subtype: Link Rect: 157, 646, 169, 655 ActionDest: 810 Annotation: Subtype: Link Rect: 439, 646, 451, 655 ActionDest: 810 Annotation: Subtype: Link Rect: 280, 560, 294, 572 ActionDest: 231 Annotation: Subtype: Link Rect: 112, 495, 124, 504 ActionDest: 842 Annotation: Subtype: Link Rect: 109, 427, 131, 437 ActionDest: 361 Annotation: Subtype: Link Rect: 473, 382, 495, 394 ActionDest: 338 Annotation: Subtype: Link Rect: 374, 290, 386, 299 ActionDest: 810 Annotation: Subtype: Link Rect: 121, 263, 142, 272 ActionDest: 338 Annotation: Subtype: Link Rect: 482, 207, 493, 216 ActionDest: 810 Annotation: Subtype: Link Rect: 494, 207, 506, 216 ActionDest: 842 Annotation: Subtype: Link Rect: 503, 124, 515, 133 ActionDest: 842 Annotation: Subtype: Link Rect: 225, 97, 239, 106 ActionDest: 338 Page: Sequence: 6 Annotations: Annotation: Subtype: Link Rect: 279, 613, 291, 622 ActionDest: 842 Annotation: Subtype: Link Rect: 125, 500, 132, 512 ActionDest: 262 Annotation: Subtype: Link Rect: 250, 378, 265, 390 ActionDest: 318 Page: Sequence: 7 Annotations: Annotation: Subtype: Link Rect: 109, 740, 131, 754 ActionDest: 352 Annotation: Subtype: Link Rect: 462, 659, 474, 668 ActionDest: 842 Annotation: Subtype: Link Rect: 151, 386, 158, 398 ActionDest: 378 Annotation: Subtype: Link Rect: 186, 307, 193, 317 ActionDest: 361 Page: Sequence: 8 Annotations: Annotation: Subtype: Link Rect: 130, 286, 137, 298 ActionDest: 378 Annotation: Subtype: Link Rect: 155, 286, 162, 298 ActionDest: 448 Annotation: Subtype: Link Rect: 129, 259, 136, 271 ActionDest: 448 Annotation: Subtype: Link Rect: 435, 232, 442, 244 ActionDest: 448 Page: Sequence: 9 Annotations: Annotation: Subtype: Link Rect: 112, 403, 119, 412 ActionDest: 448 Annotation: Subtype: Link Rect: 101, 180, 108, 190 ActionDest: 492 Annotation: Subtype: Link Rect: 122, 165, 129, 176 ActionDest: 492 Page: Sequence: 10 Annotations: Annotation: Subtype: Link Rect: 433, 317, 440, 329 ActionDest: 492 Annotation: Subtype: Link Rect: 443, 317, 455, 329 ActionDest: 672 Page: Sequence: 11 Page: Sequence: 12 Annotations: Annotation: Subtype: Link Rect: 457, 565, 463, 576 ActionDest: 492 Annotation: Subtype: Link Rect: 292, 524, 299, 536 ActionDest: 532 Annotation: Subtype: Link Rect: 266, 497, 273, 509 ActionDest: 532 Annotation: Subtype: Link Rect: 139, 470, 146, 482 ActionDest: 532 Annotation: Subtype: Link Rect: 127, 443, 139, 455 ActionDest: 532 Annotation: Subtype: Link Rect: 328, 416, 340, 428 ActionDest: 672 Annotation: Subtype: Link Rect: 318, 299, 330, 308 ActionDest: 810 Annotation: Subtype: Link Rect: 330, 299, 342, 308 ActionDest: 810 Annotation: Subtype: Link Rect: 393, 163, 405, 173 ActionDest: 810 Page: Sequence: 13 Annotations: Annotation: Subtype: Link Rect: 349, 700, 361, 709 ActionDest: 842 Annotation: Subtype: Link Rect: 285, 643, 292, 655 ActionDest: 378 Annotation: Subtype: Link Rect: 404, 629, 411, 641 ActionDest: 378 Annotation: Subtype: Link Rect: 430, 548, 437, 560 ActionDest: 448 Annotation: Subtype: Link Rect: 281, 521, 288, 533 ActionDest: 448 Annotation: Subtype: Link Rect: 504, 427, 518, 439 ActionDest: 318 Annotation: Subtype: Link Rect: 200, 375, 212, 385 ActionDest: 842 Annotation: Subtype: Link Rect: 388, 348, 399, 358 ActionDest: 842 Annotation: Subtype: Link Rect: 135, 294, 147, 304 ActionDest: 842 Annotation: Subtype: Link Rect: 150, 295, 162, 304 ActionDest: 842 Annotation: Subtype: Link Rect: 368, 281, 380, 290 ActionDest: 842 Annotation: Subtype: Link Rect: 381, 281, 393, 290 ActionDest: 842 Annotation: Subtype: Link Rect: 273, 200, 285, 209 ActionDest: 842 Annotation: Subtype: Link Rect: 317, 173, 329, 182 ActionDest: 842 Page: Sequence: 14 Annotations: Annotation: Subtype: Link Rect: 112, 702, 124, 711 ActionDest: 842 Annotation: Subtype: Link Rect: 190, 672, 197, 686 ActionDest: 492 Annotation: Subtype: Link Rect: 255, 648, 267, 657 ActionDest: 842 Annotation: Subtype: Link Rect: 274, 605, 281, 617 ActionDest: 492 Annotation: Subtype: Link Rect: 503, 567, 515, 576 ActionDest: 842 Annotation: Subtype: Link Rect: 228, 540, 240, 549 ActionDest: 842 Annotation: Subtype: Link Rect: 325, 513, 337, 522 ActionDest: 842 Annotation: Subtype: Link Rect: 286, 389, 298, 398 ActionDest: 842 Annotation: Subtype: Link Rect: 299, 389, 311, 398 ActionDest: 842 Annotation: Subtype: Link Rect: 132, 359, 139, 371 ActionDest: 492 Annotation: Subtype: Link Rect: 142, 359, 149, 371 ActionDest: 532 Annotation: Subtype: Link Rect: 465, 359, 472, 371 ActionDest: 492 Annotation: Subtype: Link Rect: 255, 346, 262, 358 ActionDest: 492 Annotation: Subtype: Link Rect: 416, 346, 423, 358 ActionDest: 492 Annotation: Subtype: Link Rect: 207, 319, 214, 331 ActionDest: 532 Annotation: Subtype: Link Rect: 174, 292, 181, 304 ActionDest: 532 Annotation: Subtype: Link Rect: 360, 292, 367, 304 ActionDest: 532 Annotation: Subtype: Link Rect: 505, 292, 512, 304 ActionDest: 532 Annotation: Subtype: Link Rect: 131, 251, 138, 263 ActionDest: 532 Annotation: Subtype: Link Rect: 141, 251, 153, 263 ActionDest: 672 Annotation: Subtype: Link Rect: 116, 238, 123, 250 ActionDest: 532 Annotation: Subtype: Link Rect: 279, 224, 291, 236 ActionDest: 532 Annotation: Subtype: Link Rect: 386, 224, 398, 236 ActionDest: 532 Annotation: Subtype: Link Rect: 300, 211, 312, 223 ActionDest: 532 Annotation: Subtype: Link Rect: 277, 197, 288, 209 ActionDest: 672 Page: Sequence: 15 Annotations: Annotation: Subtype: Link Rect: 341, 664, 518, 675 Annotation: Subtype: Link Rect: 223, 286, 260, 296 Annotation: Subtype: Link Rect: 267, 286, 303, 296 Annotation: Subtype: Link Rect: 417, 260, 454, 271 Annotation: Subtype: Link Rect: 461, 260, 497, 271 Annotation: Subtype: Link Rect: 228, 182, 265, 193 Annotation: Subtype: Link Rect: 241, 143, 279, 154 Annotation: Subtype: Link Rect: 285, 143, 321, 154 Annotation: Subtype: Link Rect: 159, 106, 196, 116 Annotation: Subtype: Link Rect: 202, 106, 238, 116 Annotation: Subtype: Link Rect: 352, 79, 390, 90 Page: Sequence: 16 Annotations: Annotation: Subtype: Link Rect: 449, 727, 486, 738 Annotation: Subtype: Link Rect: 100, 689, 137, 699 Annotation: Subtype: Link Rect: 480, 663, 516, 673 Annotation: Subtype: Link Rect: 99, 651, 135, 661 Annotation: Subtype: Link Rect: 171, 612, 208, 622 Annotation: Subtype: Link Rect: 214, 612, 250, 622 Annotation: Subtype: Link Rect: 99, 573, 135, 583 Annotation: Subtype: Link Rect: 401, 546, 438, 557 Annotation: Subtype: Link Rect: 444, 546, 480, 557 Annotation: Subtype: Link Rect: 421, 521, 459, 531 Annotation: Subtype: Link Rect: 465, 521, 501, 531 Annotation: Subtype: Link Rect: 252, 496, 290, 506 Annotation: Subtype: Link Rect: 296, 496, 332, 506 Annotation: Subtype: Link Rect: 100, 457, 137, 467 Annotation: Subtype: Link Rect: 279, 405, 317, 415 Annotation: Subtype: Link Rect: 309, 353, 347, 364 Annotation: Subtype: Link Rect: 353, 353, 389, 364 Annotation: Subtype: Link Rect: 269, 328, 306, 338 Annotation: Subtype: Link Rect: 313, 328, 349, 338 Annotation: Subtype: Link Rect: 159, 289, 196, 299 Annotation: Subtype: Link Rect: 202, 289, 238, 299 Annotation: Subtype: Link Rect: 357, 262, 393, 273 Annotation: Subtype: Link Rect: 461, 249, 499, 260 Annotation: Subtype: Link Rect: 97, 224, 513, 234 Annotation: Subtype: Link Rect: 281, 185, 519, 196 Annotation: Subtype: Link Rect: 97, 172, 177, 183 Annotation: Subtype: Link Rect: 407, 146, 519, 157 Annotation: Subtype: Link Rect: 97, 133, 278, 144 Annotation: Subtype: Link Rect: 136, 95, 174, 105 Page: Sequence: 17 Annotations: Annotation: Subtype: Link Rect: 277, 714, 315, 725 Annotation: Subtype: Link Rect: 381, 637, 419, 648 Annotation: Subtype: Link Rect: 231, 599, 269, 609 Annotation: Subtype: Link Rect: 149, 560, 187, 570 Annotation: Subtype: Link Rect: 97, 533, 396, 544 Annotation: Subtype: Link Rect: 294, 495, 331, 506 Annotation: Subtype: Link Rect: 337, 495, 373, 506 Annotation: Subtype: Link Rect: 250, 469, 519, 480 Annotation: Subtype: Link Rect: 97, 456, 115, 467 Annotation: Subtype: Link Rect: 477, 430, 514, 441 Annotation: Subtype: Link Rect: 243, 404, 281, 415 Annotation: Subtype: Link Rect: 287, 404, 323, 415 Annotation: Subtype: Link Rect: 408, 353, 445, 364 Annotation: Subtype: Link Rect: 451, 353, 487, 364 Annotation: Subtype: Link Rect: 245, 327, 282, 338 Annotation: Subtype: Link Rect: 353, 301, 390, 312 Annotation: Subtype: Link Rect: 480, 262, 516, 273 Annotation: Subtype: Link Rect: 480, 236, 516, 247 Annotation: Subtype: Link Rect: 366, 198, 403, 209 Annotation: Subtype: Link Rect: 410, 198, 446, 209 Annotation: Subtype: Link Rect: 229, 172, 266, 183 Annotation: Subtype: Link Rect: 272, 172, 308, 183 Annotation: Subtype: Link Rect: 76, 78, 166, 109 Annotation: Subtype: Link Rect: 234, 73, 407, 83 Checksum: ad0ee59e Type: CRC32 Checksum: 4f1320ac8c246a5fbb04f3380dd8e9bf Type: MD5 Checksum: 7060deb46fa9bdf85c584658a28c89e01ef646c1 Type: SHA-1 Checksum: 743eec0a7687571091723e61a92db1ccb6e8bd7faeeec905df39462de0e5d01e Type: SHA-256