e3a841c0d3e5aed4a5417f6f2ea225a2  pmc_33401696.pdf
0437b07d83da11dbc176c4289d7b68d69101cad2  pmc_33401696.pdf
a98822a0b40014b9a72a3fc106d53eb9171d45c816e33347d44cd3c53068cc25  pmc_33401696.pdf

Title:           Alkaloids Analysis of Habranthus cardenasianus (Amaryllidaceae), Anti-Cholinesterase Activity and Biomass Production by Propagation Strategies
Subject:         Plants in the Amaryllidaceae family synthesize a diversity of bioactive alkaloids. Some of these plant species are not abundant and have a low natural multiplication rate. The aims of this work were the alkaloids analysis of a Habranthus cardenasianus bulbs extract, the evaluation of its inhibitory activity against cholinesterases, and to test several propagation strategies for biomass production. Eleven compounds were characterized by GC-MS in the alkaloid extract, which showed a relatively high proportion of tazettine. The known alkaloids tazettine, haemanthamine, and the epimer mixture haemanthidine/6-epi-haemanthidine were isolated and identified by spectroscopic methods. Inhibitory cholinesterases activity was not detected. Three forms of propagation were performed: bulb propagation from seed, cut-induced bulb division, and micropropagated bulbs. Finally, different imbibition and post-collection times were evaluated in seed germination assays. The best propagation method was cut-induced bulb division with longitudinal cuts into quarters (T1) while the best conditions for seed germination were 0-day of post-collection and two days of imbibition. The alkaloids analyses of the H. cardenasianus bulbs showed that they are a source of anti-tumoral alkaloids, especially pretazettine (tazettine) and T1 is a sustainable strategy for its propagation and domestication to produce bioactive alkaloids.
Keywords:        Amaryllidaceae; bioactive alkaloids; GC-MS; propagation methods; biomass production
Author:          Daniel Zaragoza-Puchol, Javier E. Ortiz, Alejandro A. Orden, Marianela Sanchez, Jorge Palermo, Alejandro Tapia, Jaume Bastida and Gabriela E. Feresin
Creator:         LaTeX with hyperref
Producer:        pdfTeX-1.40.21
CreationDate:    Wed Jan  6 04:45:03 2021 CET
ModDate:         Wed Jan  6 04:51:12 2021 CET
Custom Metadata: no
Metadata Stream: no
Tagged:          no
UserProperties:  no
Suspects:        no
Form:            none
JavaScript:      no
Pages:           18
Encrypted:       no
Page size:       595.276 x 841.89 pts (A4)
Page rot:        0
File size:       1656024 bytes
Optimized:       no
PDF version:     1.7


name                                 type              encoding         emb sub uni object ID
------------------------------------ ----------------- ---------------- --- --- --- ---------
EUNBMT+VnURWPalladioL                Type 1            Custom           yes yes no      10  0
BQKOZC+URWPalladioL-Ital             Type 1            Custom           yes yes no      15  0
OSVIUB+URWPalladioL-Bold             Type 1            Custom           yes yes no      20  0
WDNXXL+URWPalladioL-BoldItal         Type 1            Custom           yes yes no      24  0
OJZBEA+URWPalladioL-Roma             Type 1            Custom           yes yes no      28  0
PUSVWS+TimesNewRomanPS-BoldItalicMT  Type 1C           WinAnsi          yes yes no      36  0
MCBCCI+SourceSansRoman-Regular       Type 1C           Custom           yes yes no      58  0
MCBCCM+SourceSansRoman.613wght       Type 1C           Custom           yes yes no      62  0
MCBCCO+SourceSansRoman-Bold          Type 1C           Custom           yes yes no      66  0
ZZWDNM+CMSY10                        Type 1            Builtin          yes yes no      78  0
FJMBHN+PalatinoLinotype,Italic       TrueType          WinAnsi          yes yes no      95  0
FJMBHP+PalatinoLinotype              TrueType          WinAnsi          yes yes no      98  0
FJMBJB+PalatinoLinotype,Bold         TrueType          WinAnsi          yes yes no     101  0
FJMDEK+TimesNewRoman                 CID TrueType      Identity-H       yes yes yes    104  0
NKXLQD+EURM10                        Type 1            Builtin          yes yes no     114  0
MBPIJH+Arial                         CID TrueType      Identity-H       yes yes yes    128  0
MBPFNP+PalatinoLinotype,Italic       TrueType          WinAnsi          yes yes no     134  0
MBPFPA+PalatinoLinotype              CID TrueType      Identity-H       yes yes yes    137  0
MBPFPB+PalatinoLinotype              TrueType          WinAnsi          yes yes no     143  0
MBPFPD+PalatinoLinotype,Bold         TrueType          WinAnsi          yes yes no     146  0
MBPHBP+TimesNewRoman                 CID TrueType      Identity-H       yes yes yes    149  0
FJMFEO+Arial                         CID TrueType      Identity-H       yes yes yes    183  0
FJMGII+Calibri                       CID TrueType      Identity-H       yes yes yes    189  0
FJMGJI+Calibri,Bold                  CID TrueType      Identity-H       yes yes yes    195  0
FJMBHN+PalatinoLinotype,Italic       TrueType          WinAnsi          yes yes no     201  0
FJMBHP+PalatinoLinotype              TrueType          WinAnsi          yes yes no     204  0
FJMBJB+PalatinoLinotype,Bold         TrueType          WinAnsi          yes yes no     207  0
FJMDEK+TimesNewRoman                 CID TrueType      Identity-H       yes yes yes    210  0
FJMFEO+Arial                         CID TrueType      Identity-H       yes yes yes    226  0
FJMGII+Calibri                       CID TrueType      Identity-H       yes yes yes    232  0
FJMGJI+Calibri,Bold                  CID TrueType      Identity-H       yes yes yes    238  0
FJMBHN+PalatinoLinotype,Italic       TrueType          WinAnsi          yes yes no     244  0
FJMBHP+PalatinoLinotype              TrueType          WinAnsi          yes yes no     247  0
FJMBJB+PalatinoLinotype,Bold         TrueType          WinAnsi          yes yes no     250  0
FJMDEK+TimesNewRoman                 CID TrueType      Identity-H       yes yes yes    253  0
FJMGII+Calibri                       CID TrueType      Identity-H       yes yes yes    271  0
FJMGJI+Calibri,Bold                  CID TrueType      Identity-H       yes yes yes    277  0
FJMIIN+MicrosoftSansSerif            CID TrueType      Identity-H       yes yes yes    283  0
FJMIKO+Calibri,Italic                CID TrueType      Identity-H       yes yes yes    289  0
FJMBHN+PalatinoLinotype,Italic       TrueType          WinAnsi          yes yes no     295  0
FJMBHP+PalatinoLinotype              TrueType          WinAnsi          yes yes no     298  0
FJMBJB+PalatinoLinotype,Bold         TrueType          WinAnsi          yes yes no     301  0
FJMDEK+TimesNewRoman                 CID TrueType      Identity-H       yes yes yes    304  0
FJMGII+Calibri                       CID TrueType      Identity-H       yes yes yes    323  0
FJMGJI+Calibri,Bold                  CID TrueType      Identity-H       yes yes yes    329  0
FJMIKO+Calibri,Italic                CID TrueType      Identity-H       yes yes yes    335  0
FJMBHN+PalatinoLinotype,Italic       TrueType          WinAnsi          yes yes no     341  0
FJMBHP+PalatinoLinotype              TrueType          WinAnsi          yes yes no     344  0
FJMBJB+PalatinoLinotype,Bold         TrueType          WinAnsi          yes yes no     347  0
FJMDEK+TimesNewRoman                 CID TrueType      Identity-H       yes yes yes    350  0
UGLOHL+PazoMath                      Type 1            Builtin          yes yes no     373  0


Jhove (Rel. 1.28.0, 2023-05-18)
 Date: 2024-10-02 03:36:09 CEST
 RepresentationInformation: pmc_33401696.pdf
  ReportingModule: PDF-hul, Rel. 1.12.4 (2023-03-16)
  LastModified: 2023-03-31 12:08:30 CEST
  Size: 1656024
  Format: PDF
  Version: 1.7
  Status: Well-Formed and valid
  SignatureMatches:
   PDF-hul
  MIMEtype: application/pdf
  PDFMetadata: 
   Objects: 604
   FreeObjects: 1
   IncrementalUpdates: 0
   DocumentCatalog: 
    PageLayout: SinglePage
    PageMode: UseNone
    Outlines: 
     Item: 
      Title: Introduction 
      Destination: section.1
     Item: 
      Title: Results and Discussion 
      Destination: section.2
      Children: 
       Item: 
        Title: Chemical Analysis 
        Destination: subsection.2.1
        Children: 
         Item: 
          Title: Alkaloid Profile by GC-MS Analysis 
          Destination: subsubsection.2.1.1
         Item: 
          Title: Alkaloid Isolation 
          Destination: subsubsection.2.1.2
       Item: 
        Title: AChE and Butyrylcholinesterase (BuChE) Inhibitory Activities 
        Destination: subsection.2.2
       Item: 
        Title: Propagation from Habranthus cardenasianus 
        Destination: subsection.2.3
       Item: 
        Title: Germination Tests 
        Destination: subsection.2.4
     Item: 
      Title: Material and Methods 
      Destination: section.3
      Children: 
       Item: 
        Title: Plant Material 
        Destination: subsection.3.1
       Item: 
        Title: Alkaloid Extraction and Isolation 
        Destination: subsection.3.2
       Item: 
        Title: Microplate Assay for AChE and BuChE Inhibitory Activities 
        Destination: subsection.3.3
       Item: 
        Title: Chemical Analysis 
        Destination: subsection.3.4
       Item: 
        Title: Propagation Methods from H. cardenasianus 
        Destination: subsection.3.5
        Children: 
         Item: 
          Title: Cut-Induced Bulb Division 
          Destination: subsubsection.3.5.1
         Item: 
          Title: Micropropagation of Bulbs 
          Destination: subsubsection.3.5.2
         Item: 
          Title: Seed Propagation 
          Destination: subsubsection.3.5.3
       Item: 
        Title: Evaluated Parameters 
        Destination: subsection.3.6
       Item: 
        Title: Analysis of Data 
        Destination: subsection.3.7
     Item: 
      Title: Conclusions 
      Destination: section.4
     Item: 
      Title: References
      Destination: section.4
   Info: 
    Title: Alkaloids Analysis of Habranthus cardenasianus (Amaryllidaceae), Anti-Cholinesterase Activity and Biomass Production by Propagation Strategies
    Author: Daniel Zaragoza-Puchol, Javier E. Ortiz, Alejandro A. Orden, Marianela Sanchez, Jorge Palermo, Alejandro Tapia, Jaume Bastida and Gabriela E. Feresin
    Subject: Plants in the Amaryllidaceae family synthesize a diversity of bioactive alkaloids. Some of these plant species are not abundant and have a low natural multiplication rate. The aims of this work were the alkaloids analysis of a Habranthus cardenasianus bulbs extract, the evaluation of its inhibitory activity against cholinesterases, and to test several propagation strategies for biomass production. Eleven compounds were characterized by GC-MS in the alkaloid extract, which showed a relatively high proportion of tazettine. The known alkaloids tazettine, haemanthamine, and the epimer mixture haemanthidine/6-epi-haemanthidine were isolated and identified by spectroscopic methods. Inhibitory cholinesterases activity was not detected. Three forms of propagation were performed: bulb propagation from seed, cut-induced bulb division, and micropropagated bulbs. Finally, different imbibition and post-collection times were evaluated in seed germination assays. The best propagation method was cut-induced bulb division with longitudinal cuts into quarters (T1) while the best conditions for seed germination were 0-day of post-collection and two days of imbibition. The alkaloids analyses of the H. cardenasianus bulbs showed that they are a source of anti-tumoral alkaloids, especially pretazettine (tazettine) and T1 is a sustainable strategy for its propagation and domestication to produce bioactive alkaloids.
    Keywords: Amaryllidaceae; bioactive alkaloids; GC-MS; propagation methods; biomass production
    Creator: LaTeX with hyperref
    Producer: pdfTeX-1.40.21
    CreationDate: Wed Jan 06 04:45:03 CET 2021
    ModDate: Wed Jan 06 04:51:12 CET 2021
   ID: 0xe6fdcced4a16908a3a92baff0e0b507e, 0xe6fdcced4a16908a3a92baff0e0b507e
   Filters: 
    FilterPipeline: FlateDecode
   Fonts: 
    Type1: 
     Font: 
      BaseFont: NKXLQD+EURM10
      FontSubset: true
      FirstChar: 14
      LastChar: 22
      FontDescriptor: 
       FontName: NKXLQD+EURM10
       Flags: Symbolic
       FontBBox: -32, -243, 1060, 720
       FontFile: true
     Font: 
      BaseFont: OSVIUB+URWPalladioL-Bold
      FontSubset: true
      FirstChar: 3
      LastChar: 134
      FontDescriptor: 
       FontName: OSVIUB+URWPalladioL-Bold
       Flags: Symbolic
       FontBBox: -152, -301, 1000, 935
       FontFile: true
      EncodingDictionary: 
       Differences: true
     Font: 
      BaseFont: UGLOHL+PazoMath
      FontSubset: true
      FirstChar: 83
      LastChar: 83
      FontDescriptor: 
       FontName: UGLOHL+PazoMath
       Flags: Symbolic
       FontBBox: -40, -283, 878, 946
       FontFile: true
     Font: 
      BaseFont: WDNXXL+URWPalladioL-BoldItal
      FontSubset: true
      FirstChar: 72
      LastChar: 117
      FontDescriptor: 
       FontName: WDNXXL+URWPalladioL-BoldItal
       Flags: Symbolic
       FontBBox: -170, -300, 1073, 935
       FontFile: true
      EncodingDictionary: 
       Differences: true
     Font: 
      BaseFont: EUNBMT+VnURWPalladioL
      FontSubset: true
      FirstChar: 129
      LastChar: 226
      FontDescriptor: 
       FontName: EUNBMT+VnURWPalladioL
       Flags: Symbolic
       FontBBox: -166, -283, 1021, 943
       FontFile: true
      EncodingDictionary: 
       Differences: true
     Font: 
      BaseFont: OJZBEA+URWPalladioL-Roma
      FontSubset: true
      FirstChar: 2
      LastChar: 241
      FontDescriptor: 
       FontName: OJZBEA+URWPalladioL-Roma
       Flags: Symbolic
       FontBBox: -166, -283, 1021, 943
       FontFile: true
      EncodingDictionary: 
       Differences: true
     Font: 
      BaseFont: ZZWDNM+CMSY10
      FontSubset: true
      FirstChar: 0
      LastChar: 112
      FontDescriptor: 
       FontName: ZZWDNM+CMSY10
       Flags: Symbolic
       FontBBox: -29, -960, 1116, 775
       FontFile: true
     Font: 
      BaseFont: BQKOZC+URWPalladioL-Ital
      FontSubset: true
      FirstChar: 2
      LastChar: 122
      FontDescriptor: 
       FontName: BQKOZC+URWPalladioL-Ital
       Flags: Symbolic
       FontBBox: -170, -305, 1010, 941
       FontFile: true
      EncodingDictionary: 
       Differences: true
   Pages: 
    Page: 
     Sequence: 1
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 35, 759, 179, 795
      Annotation: 
       Subtype: Link
       Rect: 515, 759, 560, 795
      Annotation: 
       Subtype: Link
       Rect: 165, 641, 176, 654
      Annotation: 
       Subtype: Link
       Rect: 359, 641, 370, 654
      Annotation: 
       Subtype: Link
       Rect: 479, 641, 490, 654
      Annotation: 
       Subtype: Link
       Rect: 203, 628, 215, 642
      Annotation: 
       Subtype: Link
       Rect: 425, 628, 436, 642
      Annotation: 
       Subtype: Link
       Rect: 143, 337, 154, 346
      Annotation: 
       Subtype: Link
       Rect: 35, 325, 154, 334
      Annotation: 
       Subtype: Link
       Rect: 35, 315, 95, 322
      Annotation: 
       Subtype: Link
       Rect: 35, 162, 97, 184
      Annotation: 
       Subtype: Link
       Rect: 123, 89, 154, 98
      Annotation: 
       Subtype: Link
       Rect: 35, 77, 154, 86
      Annotation: 
       Subtype: Link
       Rect: 35, 66, 49, 74
      Annotation: 
       Subtype: Link
       Rect: 549, 99, 556, 108
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 375, 74, 382, 82
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 117, 17, 277, 28
      Annotation: 
       Subtype: Link
       Rect: 399, 17, 558, 28
      Annotation: 
       Subtype: Link
       Rect: 35, 449, 87, 468
    Page: 
     Sequence: 2
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 281, 724, 288, 733
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 384, 674, 391, 683
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 408, 636, 415, 645
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 462, 586, 469, 595
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 469, 586, 476, 595
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 417, 548, 424, 557
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 286, 523, 293, 532
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 216, 498, 228, 507
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 490, 485, 497, 495
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 238, 460, 245, 469
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 189, 410, 201, 419
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 202, 410, 214, 419
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 253, 398, 260, 407
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 260, 398, 272, 407
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 273, 398, 285, 407
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 288, 398, 300, 407
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 461, 372, 473, 382
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 268, 322, 280, 331
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 473, 322, 485, 331
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 485, 322, 497, 331
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 232, 310, 244, 319
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 247, 310, 259, 319
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 373, 310, 385, 319
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 388, 310, 400, 319
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 329, 297, 341, 306
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 333, 285, 345, 294
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 226, 259, 238, 269
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 221, 247, 233, 256
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 531, 235, 543, 244
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 544, 235, 556, 244
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 407, 197, 419, 206
       ActionDest: 380
    Page: 
     Sequence: 3
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 437, 697, 444, 706
       ActionDest: 82
      Annotation: 
       Subtype: Link
       Rect: 268, 685, 275, 694
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 360, 634, 367, 643
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 390, 572, 402, 581
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 405, 572, 417, 581
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 318, 127, 325, 138
       ActionDest: 82
      Annotation: 
       Subtype: Link
       Rect: 512, 104, 524, 112
       ActionDest: 383
    Page: 
     Sequence: 4
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 414, 697, 425, 706
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 318, 634, 330, 643
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 203, 609, 210, 618
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 211, 609, 223, 618
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 200, 569, 207, 581
       ActionDest: 85
      Annotation: 
       Subtype: Link
       Rect: 467, 572, 474, 581
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 474, 572, 486, 581
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 544, 237, 556, 246
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 544, 187, 556, 196
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 378, 162, 390, 171
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 390, 162, 402, 171
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 505, 137, 517, 146
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 297, 86, 309, 95
       ActionDest: 383
    Page: 
     Sequence: 5
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 480, 685, 492, 694
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 374, 659, 386, 669
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 512, 609, 524, 618
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 237, 584, 249, 593
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 249, 584, 261, 593
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 260, 559, 272, 568
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 500, 496, 512, 505
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 245, 471, 257, 480
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 326, 471, 338, 480
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 487, 433, 499, 443
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 233, 305, 241, 317
       ActionDest: 118
      Annotation: 
       Subtype: Link
       Rect: 257, 283, 269, 292
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 225, 257, 232, 267
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 232, 258, 244, 267
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 245, 257, 257, 267
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 257, 257, 269, 267
       ActionDest: 383
    Page: 
     Sequence: 6
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 363, 405, 375, 414
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 541, 151, 548, 163
       ActionDest: 173
      Annotation: 
       Subtype: Link
       Rect: 238, 141, 250, 150
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 367, 128, 378, 137
       ActionDest: 380
    Page: 
     Sequence: 7
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 201, 198, 208, 210
       ActionDest: 173
      Annotation: 
       Subtype: Link
       Rect: 341, 200, 353, 209
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 545, 175, 556, 184
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 464, 150, 476, 159
       ActionDest: 380
    Page: 
     Sequence: 8
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 361, 720, 368, 732
       ActionDest: 173
      Annotation: 
       Subtype: Link
       Rect: 322, 710, 334, 719
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 449, 710, 461, 719
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 544, 697, 556, 706
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 505, 619, 512, 631
       ActionDest: 216
      Annotation: 
       Subtype: Link
       Rect: 336, 584, 348, 593
       ActionDest: 383
    Page: 
     Sequence: 9
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 420, 722, 432, 731
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 442, 694, 449, 706
       ActionDest: 311
      Annotation: 
       Subtype: Link
       Rect: 489, 657, 496, 670
       ActionDest: 311
      Annotation: 
       Subtype: Link
       Rect: 246, 632, 253, 644
       ActionDest: 311
      Annotation: 
       Subtype: Link
       Rect: 284, 607, 291, 619
       ActionDest: 311
      Annotation: 
       Subtype: Link
       Rect: 439, 594, 446, 608
       ActionDest: 311
      Annotation: 
       Subtype: Link
       Rect: 458, 556, 465, 570
       ActionDest: 311
    Page: 
     Sequence: 10
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 529, 336, 536, 348
       ActionDest: 216
      Annotation: 
       Subtype: Link
       Rect: 284, 326, 296, 335
       ActionDest: 380
    Page: 
     Sequence: 11
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 195, 634, 202, 644
       ActionDest: 357
      Annotation: 
       Subtype: Link
       Rect: 330, 634, 342, 643
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 385, 597, 396, 606
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 471, 597, 483, 606
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 263, 115, 270, 127
       ActionDest: 357
      Annotation: 
       Subtype: Link
       Rect: 549, 102, 556, 114
       ActionDest: 357
      Annotation: 
       Subtype: Link
       Rect: 544, 93, 556, 101
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 549, 67, 556, 77
       ActionDest: 357
    Page: 
     Sequence: 12
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 407, 707, 415, 719
       ActionDest: 357
      Annotation: 
       Subtype: Link
       Rect: 365, 697, 377, 706
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 392, 685, 399, 694
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 356, 634, 368, 643
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 224, 482, 231, 492
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 191, 395, 203, 404
       ActionDest: 380
    Page: 
     Sequence: 13
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 545, 719, 557, 728
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 284, 707, 296, 716
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 463, 390, 470, 402
       ActionDest: 82
    Page: 
     Sequence: 14
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 262, 707, 274, 716
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 274, 707, 286, 716
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 287, 707, 299, 716
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 299, 707, 311, 716
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 438, 505, 450, 515
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 544, 267, 556, 276
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 282, 179, 294, 188
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 295, 179, 307, 188
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 307, 179, 319, 188
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 484, 116, 496, 125
       ActionDest: 383
    Page: 
     Sequence: 15
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 310, 559, 322, 568
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 305, 521, 317, 530
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 215, 421, 227, 430
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 227, 421, 239, 430
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 239, 421, 251, 430
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 252, 421, 264, 430
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 221, 224, 233, 233
       ActionDest: 380
      Annotation: 
       Subtype: Link
       Rect: 233, 224, 245, 233
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 246, 224, 258, 233
       ActionDest: 383
      Annotation: 
       Subtype: Link
       Rect: 258, 224, 270, 233
       ActionDest: 383
    Page: 
     Sequence: 16
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 403, 463, 415, 472
       ActionDest: 383
    Page: 
     Sequence: 17
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 519, 637, 557, 648
      Annotation: 
       Subtype: Link
       Rect: 59, 626, 95, 636
      Annotation: 
       Subtype: Link
       Rect: 420, 602, 458, 613
      Annotation: 
       Subtype: Link
       Rect: 212, 580, 250, 590
      Annotation: 
       Subtype: Link
       Rect: 256, 580, 292, 590
      Annotation: 
       Subtype: Link
       Rect: 156, 557, 194, 567
      Annotation: 
       Subtype: Link
       Rect: 355, 533, 392, 544
      Annotation: 
       Subtype: Link
       Rect: 404, 510, 442, 521
      Annotation: 
       Subtype: Link
       Rect: 127, 488, 164, 498
      Annotation: 
       Subtype: Link
       Rect: 306, 464, 343, 475
      Annotation: 
       Subtype: Link
       Rect: 59, 419, 97, 429
      Annotation: 
       Subtype: Link
       Rect: 410, 361, 448, 372
      Annotation: 
       Subtype: Link
       Rect: 394, 349, 432, 360
      Annotation: 
       Subtype: Link
       Rect: 59, 315, 97, 325
      Annotation: 
       Subtype: Link
       Rect: 104, 281, 141, 291
      Annotation: 
       Subtype: Link
       Rect: 404, 257, 442, 268
      Annotation: 
       Subtype: Link
       Rect: 517, 245, 555, 256
      Annotation: 
       Subtype: Link
       Rect: 407, 199, 444, 210
      Annotation: 
       Subtype: Link
       Rect: 278, 176, 316, 187
      Annotation: 
       Subtype: Link
       Rect: 59, 97, 97, 107
      Annotation: 
       Subtype: Link
       Rect: 103, 97, 139, 107
      Annotation: 
       Subtype: Link
       Rect: 95, 62, 132, 72
      Annotation: 
       Subtype: Link
       Rect: 139, 62, 175, 72
    Page: 
     Sequence: 18
     Annotations: 
      Annotation: 
       Subtype: Link
       Rect: 95, 710, 133, 720
      Annotation: 
       Subtype: Link
       Rect: 59, 664, 97, 674
      Annotation: 
       Subtype: Link
       Rect: 351, 652, 388, 663
      Annotation: 
       Subtype: Link
       Rect: 390, 629, 428, 640
      Annotation: 
       Subtype: Link
       Rect: 301, 607, 339, 617
      Annotation: 
       Subtype: Link
       Rect: 467, 594, 504, 605
      Annotation: 
       Subtype: Link
       Rect: 519, 583, 557, 594
      Annotation: 
       Subtype: Link
       Rect: 59, 572, 95, 582
      Annotation: 
       Subtype: Link
       Rect: 371, 548, 407, 559
      Annotation: 
       Subtype: Link
       Rect: 519, 514, 557, 525
      Annotation: 
       Subtype: Link
       Rect: 296, 491, 334, 502
      Annotation: 
       Subtype: Link
       Rect: 59, 469, 97, 479
      Annotation: 
       Subtype: Link
       Rect: 266, 445, 303, 456
      Annotation: 
       Subtype: Link
       Rect: 95, 423, 133, 433
      Annotation: 
       Subtype: Link
       Rect: 514, 399, 551, 410
      Annotation: 
       Subtype: Link
       Rect: 491, 376, 528, 387
      Annotation: 
       Subtype: Link
       Rect: 231, 330, 268, 341
      Annotation: 
       Subtype: Link
       Rect: 309, 307, 346, 318
      Annotation: 
       Subtype: Link
       Rect: 451, 295, 560, 306
      Annotation: 
       Subtype: Link
       Rect: 56, 284, 312, 295
      Annotation: 
       Subtype: Link
       Rect: 232, 261, 270, 272
      Annotation: 
       Subtype: Link
       Rect: 338, 238, 423, 249
  Checksum: 3ff8d8ac
   Type: CRC32
  Checksum: e3a841c0d3e5aed4a5417f6f2ea225a2
   Type: MD5
  Checksum: 0437b07d83da11dbc176c4289d7b68d69101cad2
   Type: SHA-1
  Checksum: a98822a0b40014b9a72a3fc106d53eb9171d45c816e33347d44cd3c53068cc25
   Type: SHA-256